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REVIEW Transgenic and physiological mouse models give insights into different aspects of amyotrophic lateral sclerosis Francesca De Giorgio 1, *, Cheryl Maduro 1, *, Elizabeth M. C. Fisher 1, ‡ and Abraham Acevedo-Arozena 2, ‡ ABSTRACT A wide range of genetic mouse models is available to help researchers dissect human disease mechanisms. Each type of model has its own distinctive characteristics arising from the nature of the introduced mutation, as well as from the specific changes to the gene of interest. Here, we review the current range of mouse models with mutations in genes causative for the human neurodegenerative disease amyotrophic lateral sclerosis. We focus on the two main types of available mutants: transgenic mice and those that express mutant genes at physiological levels from gene targeting or from chemical mutagenesis. We compare the phenotypes for genes in which the two classes of model exist, to illustrate what they can teach us about different aspects of the disease, noting that informative models may not necessarily mimic the full trajectory of the human condition. Transgenic models can greatly overexpress mutant or wild- type proteins, giving us insight into protein deposition mechanisms, whereas models expressing mutant genes at physiological levels may develop slowly progressing phenotypes but illustrate early-stage disease processes. Although no mouse models fully recapitulate the human condition, almost all help researchers to understand normal and abnormal biological processes, providing that the individual characteristics of each model type, and how these may affect the interpretation of the data generated from each model, are considered and appreciated. KEY WORDS: Amyotrophic lateral sclerosis, ALS, Transgenic, Knock-in, ENU, Gene targeted Introduction Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder first described in 1869 by Jean-Martin Charcot (Charcot and Joffroy, 1869). It has a mean incidence of ∼2/ 100,000 worldwide and a prevalence of ∼6/100,000 in Europe (Costa and de Carvalho, 2016; Marin et al., 2016), with a lifetime risk of ∼1 in 300 in Western populations (Brown and Al-Chalabi, 2017). ALS patients typically present a focal onset, starting as unilateral limb weakness or bulbar impairment. Clinical symptoms usually start in mid-life and are a consequence of the dysfunction and death of motor neurons (MNs) in the primary motor cortex, brainstem and spinal cord, which causes spasticity, weakness and muscle wasting, gradually leading to paralysis and death from respiratory failure, typically less than 5 years from diagnosis (Huynh et al., 2016; Van Damme et al., 2017). There are no effective treatments for ALS apart from daily care and support to counteract the symptoms. Currently, there are only two US Food and Drug Administration (FDA)- and European Medicines Agency (EMA)-approved neuroprotective drugs that increase the lifespan of some patients by a few months: Riluzole, which blocks excessive glutamatergic neurotransmission, and Edaravone, which prevents oxidative stress damage. Although 90% of ALS patients have sporadic (sALS) disease without apparent family history, ∼5-10% of cases are familial (fALS), usually showing monogenic autosomal dominant inheritance (Brown and Al-Chalabi, 2017). In 1993, the first causative gene for ALS was discovered, encoding the enzyme Cu/Zn superoxide dismutase 1 (SOD1) (Rosen et al., 1993). Research shows that SOD1-ALS accounts for ∼20% of fALS and ∼2% of sALS, with >150 mutations identified throughout the coding region and causing an unknown toxic gain of function (GOF) (Saccon et al., 2013; Kaur et al., 2016). SOD1 is ubiquitously expressed and important for the removal of free radicals, although it likely has other non-canonical roles; for example, as a transcriptional regulator under oxidative stress, possibly as an RNA-binding protein and a signalling molecule (Bunton-Stasyshyn et al., 2015). Since the discovery of SOD1’s association with ALS, mutations in more than 20 genes were found to be causative, most with an autosomal-dominant pattern of transmission, together with >30 potential disease-modifying genes (Li and Wu, 2016). Causative genes include the chromosome 9 open reading frame 72 (C9ORF72), in which an intronic hexanucleotide repeat expansion gives rise to ALS. This mutation is the most common cause of fALS, and is found in up to 40% of fALS and ∼9% of sALS in Caucasians (DeJesus- Hernandez et al., 2011; Renton et al., 2011; Goldstein et al., 2018). Other well known ‘ALS genes’ include TAR DNA-binding protein (TARDBP; encoding TDP-43), found in ∼5% of fALS and ∼2% of sALS, and fused in sarcoma (also known as FUS RNA-binding protein; FUS), found in ∼6% of fALS and ∼1% of sALS (Ingre et al., 2015; Tarlarini et al., 2015). TDP-43 and FUS are RNA-binding heterogeneous nuclear ribonucleoproteins (hnRNPs) mainly localised in the nucleus, and are involved in mRNA splicing, gene transcription and microRNA maturation, mRNA shuttling from the nucleus to the cytoplasm and stress granule formation. Cytoplasmic mislocalisation and nuclear depletion of TDP-43 is a key feature of most ALS cases and may contribute to disease pathogenesis (Guerrero et al., 2016). Protein aggregates containing truncated hyperphosphorylated and/or ubiquitinated TDP-43 are found within MNs in >95% of ALS-affected brains and spinal cords (Chou et al., 2018), and can occur in other neurological disorders, including Alzheimer’s, Parkinson’s and Huntington’s diseases, highlighting 1 Department of Neuromuscular Diseases, UCL Institute of Neurology, and MRC Centre for Neuromuscular Disease, University College London, Queen Square, London WC1N 3BG, UK. 2 Unidad de Investigació n Hospital Universitario de Canarias, Fundació n Canaria de Investigació n Sanitaria and Instituto de Tecnologı ́ as Biomé dicas (ITB), La Laguna, 38320 Tenerife, Spain. *These authors contributed equally to this work ‡ Authors for correspondence ([email protected]; [email protected]) E.M.C.F., 0000-0003-2850-9936; A.A.-A., 0000-0001-6127-7116 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution and reproduction in any medium provided that the original work is properly attributed. 1 © 2019. Published by The Company of Biologists Ltd | Disease Models & Mechanisms (2019) 12, dmm037424. doi:10.1242/dmm.037424 Disease Models & Mechanisms

Transgenic and physiological mouse models give insights into ......REVIEW Transgenic and physiological mouse models give insights into different aspects of amyotrophic lateral sclerosis

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  • REVIEW

    Transgenic and physiological mouse models give insights intodifferent aspects of amyotrophic lateral sclerosisFrancesca De Giorgio1,*, Cheryl Maduro1,*, Elizabeth M. C. Fisher1,‡ and Abraham Acevedo-Arozena2,‡

    ABSTRACTA wide range of genetic mouse models is available to helpresearchers dissect human disease mechanisms. Each type ofmodel has its own distinctive characteristics arising from the nature ofthe introduced mutation, as well as from the specific changes to thegene of interest. Here, we review the current range of mouse modelswith mutations in genes causative for the human neurodegenerativedisease amyotrophic lateral sclerosis. We focus on the two maintypes of available mutants: transgenic mice and those that expressmutant genes at physiological levels from gene targeting or fromchemical mutagenesis. We compare the phenotypes for genes inwhich the two classes of model exist, to illustrate what they can teachus about different aspects of the disease, noting that informativemodels may not necessarily mimic the full trajectory of the humancondition. Transgenic models can greatly overexpress mutant or wild-type proteins, giving us insight into protein deposition mechanisms,whereas models expressing mutant genes at physiological levelsmay develop slowly progressing phenotypes but illustrate early-stagedisease processes. Although no mouse models fully recapitulate thehuman condition, almost all help researchers to understand normaland abnormal biological processes, providing that the individualcharacteristics of each model type, and how these may affect theinterpretation of the data generated from each model, are consideredand appreciated.

    KEY WORDS: Amyotrophic lateral sclerosis, ALS, Transgenic,Knock-in, ENU, Gene targeted

    IntroductionAmyotrophic lateral sclerosis (ALS) is a progressiveneurodegenerative disorder first described in 1869 by Jean-MartinCharcot (Charcot and Joffroy, 1869). It has a mean incidence of ∼2/100,000 worldwide and a prevalence of ∼6/100,000 in Europe(Costa and de Carvalho, 2016; Marin et al., 2016), with a lifetimerisk of ∼1 in 300 in Western populations (Brown and Al-Chalabi,2017). ALS patients typically present a focal onset, starting asunilateral limb weakness or bulbar impairment. Clinical symptomsusually start in mid-life and are a consequence of the dysfunctionand death of motor neurons (MNs) in the primary motor cortex,

    brainstem and spinal cord, which causes spasticity, weakness andmuscle wasting, gradually leading to paralysis and death fromrespiratory failure, typically less than 5 years from diagnosis(Huynh et al., 2016; Van Damme et al., 2017).

    There are no effective treatments for ALS apart from daily careand support to counteract the symptoms. Currently, there are onlytwo US Food and Drug Administration (FDA)- and EuropeanMedicines Agency (EMA)-approved neuroprotective drugs thatincrease the lifespan of some patients by a few months: Riluzole,which blocks excessive glutamatergic neurotransmission, andEdaravone, which prevents oxidative stress damage.

    Although 90% of ALS patients have sporadic (sALS) diseasewithout apparent family history, ∼5-10% of cases are familial(fALS), usually showing monogenic autosomal dominantinheritance (Brown and Al-Chalabi, 2017). In 1993, the firstcausative gene for ALS was discovered, encoding the enzymeCu/Zn superoxide dismutase 1 (SOD1) (Rosen et al., 1993).Research shows that SOD1-ALS accounts for ∼20% of fALS and∼2% of sALS, with >150 mutations identified throughout thecoding region and causing an unknown toxic gain of function(GOF) (Saccon et al., 2013; Kaur et al., 2016). SOD1 isubiquitously expressed and important for the removal of freeradicals, although it likely has other non-canonical roles; forexample, as a transcriptional regulator under oxidative stress,possibly as an RNA-binding protein and a signalling molecule(Bunton-Stasyshyn et al., 2015).

    Since the discovery of SOD1’s association with ALS, mutationsin more than 20 genes were found to be causative, most withan autosomal-dominant pattern of transmission, together with >30potential disease-modifying genes (Li and Wu, 2016). Causativegenes include the chromosome 9 open reading frame 72 (C9ORF72),in which an intronic hexanucleotide repeat expansion gives rise toALS. This mutation is the most common cause of fALS, and is foundin up to 40% of fALS and ∼9% of sALS in Caucasians (DeJesus-Hernandez et al., 2011; Renton et al., 2011; Goldstein et al., 2018).Other well known ‘ALS genes’ include TAR DNA-binding protein(TARDBP; encoding TDP-43), found in ∼5% of fALS and ∼2% ofsALS, and fused in sarcoma (also known as FUS RNA-bindingprotein; FUS), found in∼6% of fALS and∼1% of sALS (Ingre et al.,2015; Tarlarini et al., 2015). TDP-43 and FUS are RNA-bindingheterogeneous nuclear ribonucleoproteins (hnRNPs) mainlylocalised in the nucleus, and are involved in mRNA splicing, genetranscription and microRNA maturation, mRNA shuttling from thenucleus to the cytoplasm and stress granule formation. Cytoplasmicmislocalisation and nuclear depletion of TDP-43 is a key feature ofmost ALS cases and may contribute to disease pathogenesis(Guerrero et al., 2016). Protein aggregates containing truncatedhyperphosphorylated and/or ubiquitinated TDP-43 are found withinMNs in >95% of ALS-affected brains and spinal cords (Chou et al.,2018), and can occur in other neurological disorders, includingAlzheimer’s, Parkinson’s and Huntington’s diseases, highlighting

    1Department of Neuromuscular Diseases, UCL Institute of Neurology, and MRCCentre for Neuromuscular Disease, University College London, Queen Square,London WC1N 3BG, UK. 2Unidad de Investigación Hospital Universitario deCanarias, Fundación Canaria de Investigación Sanitaria and Instituto deTecnologıás Biomédicas (ITB), La Laguna, 38320 Tenerife, Spain.*These authors contributed equally to this work

    ‡Authors for correspondence ([email protected]; [email protected])

    E.M.C.F., 0000-0003-2850-9936; A.A.-A., 0000-0001-6127-7116

    This is an Open Access article distributed under the terms of the Creative Commons AttributionLicense (https://creativecommons.org/licenses/by/4.0), which permits unrestricted use,distribution and reproduction in any medium provided that the original work is properly attributed.

    1

    © 2019. Published by The Company of Biologists Ltd | Disease Models & Mechanisms (2019) 12, dmm037424. doi:10.1242/dmm.037424

    Disea

    seModels&Mechan

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    mailto:[email protected]:[email protected]://orcid.org/0000-0003-2850-9936http://orcid.org/0000-0001-6127-7116

  • the importance of TDP-43 in neurodegeneration (Liu et al., 2017;St-Amour et al., 2018).Other genes less frequently mutated in ALS include coiled-

    coil-helix-coiled-coil-helix domain-containing 10 (CHCHD10)(Bannwarth et al., 2014), kinesin family member 5A (KIF5A)(Brenner et al., 2018), matrin 3 (MATR3) (Johnson et al., 2014),optineurin (OPTN) (Maruyama et al., 2010), profilin 1 (PFN1) (Wuet al., 2012), senataxin (SETX) (Chen et al., 2004), sequestosome 1(SQSTM1/p62) (Fecto, 2011), TANK-binding kinase 1 (TBK1)(Cirulli et al., 2015; Freischmidt et al., 2015), ubiquilin 2(UBQLN2) (Deng et al., 2011), valosin-containing protein (VCP)(Johnson et al., 2010) and VAMP-associated protein B and C(VAPB) (Nishimura et al., 2004). As each new gene is identified, thenext step is to make a mouse model. There are different types ofmutant mice, which yield different insights and should be used toaddress different research questions.

    Mouse models of ALSWe know little of early-stage ALS pathomechanisms, and we stillhave a lot to learn about the disease trajectories for fALS and sALS.Here, we discuss the main features of the different types of mousemodels that are helping us to elucidate the molecular pathology ofALS and its phenotypic implications: transgenic mice, and targeted

    and ENU mutant mice (Fig. 1). We then focus on comparing thephenotypes of mice with ALS gene mutations for which at least twoof these types of model have been published; namely, FUS, SOD1,TARDBP, VAPB, VCP and UBQLN2.

    Transgenic mouse modelsALS is mostly an autosomal-dominant disorder and therefore themajority of mouse models have been transgenic lines, made byrandomly inserting human (in most cases) mutant ALS genes intothe mouse genome (Table 1). This is a fast method of producing newstrains and, because the disease is dominant, the phenotype usuallymanifests, despite the presence of intact orthologous mouse genes.Indeed, the first model of ALS, the SOD1G93A transgenic strain[Tg(SOD1*G93A)1Gur], was published a year after the discoveryof SOD1-ALS mutations in humans (Gurney et al., 1994)(Table 1A) and remains the most commonly used ALS mousemodel. Owing to the early onset, fast disease progression towards anearly humane endpoint, progressive MN loss and low variability ofthe phenotype on defined genetic backgrounds, the SOD1G93A

    transgenic strain has become the workhorse for testing therapeuticsaimed at ameliorating ALS.

    Around 30 FUS and TARDBP mutant transgenic lines have alsobeen created, with variable levels ofMN degeneration (Table 1B,C).

    Pre-ALSPost-transcriptional

    and molecular changes

    Gene-targeted Vcp R155H (Yin et al., 2012)

    Early stageMuscle weakness or atrophy

    localised in hind/forelimb

    Middle stage

  • Table1.

    Mou

    semod

    elsof

    ALS

    forwhich

    both

    tran

    sgen

    ican

    dkn

    ock-in

    strainsarepu

    blishe

    d

    Strainna

    me

    Trans

    genic/ge

    ne-

    targeted

    knoc

    k-in/ENU

    Gen

    eticba

    ckgrou

    ndProtein

    Inclus

    ions

    /agg

    rega

    tes

    Gliosis

    MA/M

    DNMJ

    loss

    Final-stage

    dise

    ase

    (terminal

    MNloss)

    Surviva

    l(wee

    ks)

    Beh

    avioural

    analysis

    Other

    phen

    otyp

    esReferen

    ce

    (A)SOD1mou

    semod

    els

    WThS

    OD1

    (JAX00

    2297

    )Trans

    genic

    Promoter:h

    SOD1

    C57

    BL/6×

    SJL

    HighhS

    OD1ex

    pres

    sion

    with

    increa

    seden

    zymeac

    tivity

    Ubiqu

    itin:

    PP

    Mild

    ND

    Earliersign

    s:30

    -50wee

    ks;M

    Nloss:

    20-30%

    at∼10

    4wee

    ks

    75-104

    Mild

    motor

    coordina

    tion

    impa

    irmen

    tArgyrop

    hilic

    fibre

    dege

    neratio

    nin

    SC.

    (Gurne

    yet

    al.,19

    94;

    Jaarsm

    aet

    al.,

    2000

    )WThS

    OD1

    Trans

    genic

    Promoter:h

    SOD1

    C57

    BL/6×

    CBA

    hSOD1ex

    pres

    sed50

    ×high

    erthan

    endo

    geno

    usmSOD1bu

    twith

    low

    enzymeac

    tivity

    Inclus

    ions

    :PSOD1:

    PP

    ND

    ND

    Earliersign

    s:∼36

    wee

    ks;

    MNloss:∼

    41%

    atlate

    stag

    e

    ∼52

    Mild

    motor

    coordina

    tion

    impa

    irmen

    tHea

    vyva

    cuolisationin

    subicu

    lum,w

    eigh

    tlos

    s,loss

    ofPurkinjece

    lls.

    (Graffm

    oet

    al.,20

    13)

    SOD1A4V

    Trans

    genic

    Promoter:h

    SOD1

    C57

    BL/6×

    SJL

    hSOD1;

    lowex

    pres

    sion

    leve

    lswith

    noen

    zyme

    activity

    Ab

    Ab

    Ab

    Ab

    Ab

    Normal

    ND

    A4V

    /hSOD1do

    uble

    tran

    sgen

    icssh

    owph

    enotyp

    eat

    ∼35

    wee

    ksan

    ddieat

    ∼45

    wee

    ksof

    age.

    (Gurne

    yet

    al.,19

    94;

    Den

    get

    al.,20

    06)

    SOD1G37

    R(JAX

    0083

    42)

    Trans

    genic

    Promoter:h

    SOD1

    C57

    BL/6J

    ×C3H

    /HeJ

    hSOD1;

    lowex

    pres

    sion

    leve

    lswith

    noen

    zyme

    activity

    Ubiqu

    itin:

    PP

    PND

    Earliersign

    s:∼11

    -17wee

    ks;e

    nd-

    stag

    epa

    ralysis:

    P

    25-29

    Motor

    coordina

    tion

    impa

    irmen

    tand

    cogn

    itive

    impa

    irmen

    t

    Dev

    elop

    rigid

    thorac

    olum

    bar

    kyph

    osis.

    (Won

    get

    al.,19

    95;

    Filaliet

    al.,20

    11)

    SOD1G37

    RTrans

    genic

    Promoter:h

    NFL

    C57

    BL/6J

    ×C3H

    /HeJ

    hSOD14×

    high

    erex

    pres

    sion

    than

    endo

    geno

    usSOD1

    ND

    ND

    ND

    ND

    Ab/no

    rmal

    Normal

    Normal

    Ab

    (Pramatarov

    aet

    al.,

    2001

    )SOD1H46

    RTrans

    genic

    Promoter:h

    SOD1

    BDF1(C

    57BL/

    6×DBA/2)

    hSOD1

    Inclus

    ions

    :PSOD1:

    PUbiqu

    itin:

    P

    PND

    ND

    Earliersign

    s:20

    wee

    ks;

    MNloss:%

    ND

    24Motor

    coordina

    tion

    impa

    irmen

    tBod

    yweigh

    tlos

    s.(C

    hang

    -Hon

    get

    al.,

    2005

    )

    SOD1H46

    R/H48

    QTrans

    genic

    Promoter:h

    SOD1

    C3H

    /HeJ

    ×C57

    BL/6J

    hSOD1;

    high

    expres

    sion

    but

    inac

    tiveprotein

    Inclus

    ions

    :PUbiqu

    itin:

    PP

    ND

    ND

    Earliersign

    s:17

    -26wee

    ks;M

    Nloss:

    %ND;e

    nd-stage

    paralysis:

    ∼21

    wee

    ks

    ND

    Motor

    coordina

    tion

    impa

    irmen

    tHya

    line,

    thiofla

    vin-S-pos

    itive

    inclus

    ions

    .(W

    anget

    al.,20

    02)

    SOD1H46

    R/H48

    Q/

    H63

    G/H12

    0GTrans

    genic

    Promoter:h

    SOD1

    C3H

    /HeJ

    ×C57

    BL/6J

    hSOD1;

    high

    expres

    sion

    but

    inac

    tiveprotein

    Inclus

    ions

    :PSOD1:

    PUbiqu

    itin:

    P

    PND

    ND

    Earliersign

    s:35

    -52wee

    ks;M

    Nloss:

    %ND;e

    nd-stage

    paralysis:

    34-52wee

    ks

    44-52

    Motor

    coordina

    tion

    impa

    irmen

    tThioflavin-S-pos

    itive

    inclus

    ions

    .(W

    anget

    al.,20

    03)

    SOD1L8

    4VTrans

    genic

    Promoter:h

    SOD1

    ND

    hSOD1

    Ab

    Ab

    Ab

    Ab

    Earliersign

    s:21

    -26wee

    ks;n

    ofin

    alMNloss

    26-30

    ND

    ND

    (Tob

    isaw

    aet

    al.,

    2003

    )

    SOD1G85

    R(JAX

    0082

    48)

    Trans

    genic(G

    85R/W

    ThS

    OD1)

    Promoter:h

    SOD1

    ND

    hSOD1;

    lowex

    pres

    sion

    Inclus

    ions

    :PSOD1:

    PUbiqu

    itin:

    P

    PP

    ND

    Earliersign

    s:26

    -35wee

    ks;M

    Nloss:

    %ND;e

    nd-stage

    paralysis:2wee

    ksafter

    initial

    sign

    s

    28-37

    Progres

    sive

    motor

    coordina

    tionim

    pairm

    ent

    Late

    onse

    t(>34

    wee

    ks),bu

    trapidprog

    ressionto

    paralysis.

    (Bruijn

    etal.,19

    97)

    SOD1Thy

    1.2-G85

    R(EGFP)

    bicistronic

    Trans

    genic

    Promoter:T

    hy1.2

    C57

    BL/6

    hSOD1;

    high

    expres

    sion

    but

    lowmetab

    olicstab

    ility

    Ab

    Ab

    ND

    Ab

    Ab

    Normal

    Ab

    Ab

    (Linoet

    al.,20

    02)

    SOD1G85

    RTrans

    genic(also

    gene

    ratedG85

    R/

    WThS

    OD1)

    Promoter:h

    SOD1

    C57

    BL/6J

    hSOD1;

    high

    expres

    sion

    .Lo

    ssof

    endo

    geno

    usSOD1ac

    tivity

    andmutan

    tSOD1

    Inclus

    ions

    :PSOD1:

    PND

    ND

    PEarliersigns

    :44-48

    wee

    ks;

    MNloss:s

    ignifican

    t(%

    ND);dise

    ase

    duratio

    n∼5-6wee

    ks

    50-54

    Progres

    sive

    motor

    coordina

    tionim

    pairm

    ent

    ND

    (Wan

    get

    al.,20

    09)

    SOD1G86

    R(JAX

    0051

    10)

    Trans

    genic

    Promoter:m

    Sod

    1FVB/N

    mSOD1;

    increa

    sed

    endo

    geno

    usex

    pres

    sion

    ND

    ND

    ND

    ND

    Earliersign

    s:13

    -17wee

    ks;M

    Nloss:

    %ND

    17Motor

    coordina

    tion

    impa

    irmen

    tPykno

    sisan

    dka

    rorrhe

    xisin

    MNs.

    Profoun

    dmus

    clewas

    ting

    andun

    able

    totake

    food

    andwater.

    (Ripps

    etal.,19

    95)

    SOD1GFAP-G

    86R

    Trans

    genic

    Promoter:h

    GFAP

    C57

    BL/6/CBA

    Increa

    sedmSOD1

    expres

    sion

    ,correlates

    with

    copy

    numbe

    r

    Ab

    ND

    Ab

    Ab

    Earliersign

    s:>70

    wee

    ks;

    nofin

    alMNloss

    >70

    Normal

    Astrocytic

    morph

    olog

    ych

    ange

    s.(W

    onget

    al.,19

    95)

    SOD1D90

    ATrans

    genic

    Promoter:h

    SOD1

    C57

    BL/6SJL

    ×C5B

    L/6J

    Bom

    hSOD1;

    high

    expres

    sion

    and

    activity

    (6-8×high

    erthan

    that

    ofNtg

    inCNS)

    Inclus

    ions

    :PSOD1:

    PP

    ND

    ND

    Earliersign

    s:52

    wee

    ks;

    40%

    loss

    ofve

    ntral

    horn

    neuron

    s;dise

    ase

    duratio

    n∼50

    days

    61Motor

    coordina

    tion

    impa

    irmen

    tHOMs:

    disten

    dedblad

    der

    (res

    emblingHOM

    D90

    Apa

    tients)

    andva

    cuoles

    throug

    hout

    ventral

    neurop

    hil.

    Age

    -dep

    ende

    ntph

    enotyp

    ein

    HEM.

    (Jon

    sson

    etal.,

    2006

    )

    SOD1G93

    A(JAX

    0027

    26)

    Trans

    genic

    Promoter:h

    SOD1

    C57

    BL/6

    SJL

    ×C57

    BL/6

    hSOD1;

    high

    expres

    sion

    Inclus

    ions

    :PP

    PP

    Earliersign

    s:13

    -17wee

    ks;M

    Nloss:

    50%

    at∼17

    wee

    ks;

    end-stag

    epa

    ralysis:

    ∼19

    wee

    ks

    17-26

    Motor

    coordina

    tion

    impa

    irmen

    tFem

    ales

    survivelong

    erthan

    males

    .Anx

    iety-like

    beha

    viou

    r,alteratio

    nsin

    spatialnav

    igationlearning

    andmem

    ory.

    (Gurne

    yet

    al.,19

    94;

    Tuet

    al.,19

    96;

    Den

    get

    al.,20

    06;

    Qua

    rtaet

    al.,

    2015

    )

    Con

    tinue

    d

    3

    REVIEW Disease Models & Mechanisms (2019) 12, dmm037424. doi:10.1242/dmm.037424

    Disea

    seModels&Mechan

    isms

  • Tab

    le1.

    Continued

    Strainna

    me

    Trans

    genic/ge

    ne-

    targeted

    knoc

    k-in/ENU

    Gen

    eticba

    ckgrou

    ndProtein

    Inclus

    ions

    /agg

    rega

    tes

    Gliosis

    MA/M

    DNMJ

    loss

    Final-stage

    dise

    ase

    (terminal

    MNloss)

    Surviva

    l(wee

    ks)

    Beh

    avioural

    analysis

    Other

    phen

    otyp

    esReferen

    ce

    SOD1G93

    A(JAX

    0023

    00)

    Trans

    genic

    Promoter:h

    SOD1

    C57

    BL/6

    SJL

    ×C57

    BL/6

    hSOD1;

    lower

    expres

    sion

    Inclus

    ions

    :PP

    PP

    Earliersign

    s:∼24

    -34wee

    ks;M

    Nloss:>

    50%;e

    nd-stage

    paralysis:

    at29

    -39wee

    ks

    29-39

    Motor

    coordina

    tion

    impa

    irmen

    t.Lo

    wco

    pyGurne

    yG93

    Aline

    Fem

    ales

    survivelong

    erthan

    males

    .(G

    urne

    yet

    al.,19

    94;

    Alexa

    nder

    etal.,

    2004

    ;Jaa

    rsma

    etal.,20

    00)

    SOD1Thy

    1.2-G93

    A(EGFP)

    bicistronic

    Trans

    genic

    Promoter:T

    hy1.2

    C57

    BL/6

    hSOD1;

    high

    expres

    sion

    Ab

    Ab

    ND

    Ab

    Ab

    Normal

    Ab

    Ab

    (Linoet

    al.,20

    02)

    SOD1Thy

    1.2-G93

    A(JAX00

    8230

    )Trans

    genic

    Promoter:T

    hy1.2

    FVBxB

    CBA

    hSOD1;

    lowex

    pres

    sion

    Inclus

    ions

    :PSOD1:

    PUbiqu

    itin:

    P

    PND

    PEarliersign

    s:54

    -104

    wee

    ks;M

    Nloss:∼

    40%

    62->10

    4Motor

    coordina

    tion

    impa

    irmen

    tNon

    -foc

    alon

    set.

    Terminal

    micesh

    owse

    vere

    weigh

    tlos

    s>30

    %.

    Age

    -dep

    ende

    ntclinical/

    patholog

    ical

    motor

    abno

    rmalities

    inHEM.

    (Jaa

    rsmaet

    al.,

    2008

    )

    SOD1G12

    7XTrans

    genic

    Promoter:h

    SOD1

    C57

    BL6

    /CBA

    xC57

    BL/6J

    hSOD1;

    lowex

    pres

    sion

    and

    noac

    tivity

    Inclus

    ions

    :PSOD1:

    P(in

    term

    inal

    mice)

    Ubiqu

    itin:

    P

    ND

    ND

    ND

    Earliersign

    s:35

    wee

    ks;

    MNloss

    %:N

    D;1

    /3of

    micesh

    owforelim

    bpa

    ralysis

    36ND

    Only1/3of

    G12

    7Xmice

    show

    edforelegon

    set,bu

    trapiddise

    aseco

    urse

    (7-

    10da

    ysafterfirst

    sign

    ).

    (Jon

    sson

    etal.,

    2004

    )

    SOD1L1

    26Z

    Trans

    genic

    Promoter:h

    SOD1

    C3H

    /HeJ

    ×C57

    BL/6J

    F2

    hSOD1;

    lowex

    pres

    sion

    Inclus

    ions

    :PSOD1:

    Ab

    Ubiqu

    itin:

    P

    PND

    ND

    Earliersign

    s:28

    -36wee

    ks;M

    Nloss

    %:N

    D;e

    nd-stage

    paralysis:

    ∼39

    wee

    ks

    ND

    ND

    Nothiofla

    vin-S-pos

    itive

    inclus

    ions

    inBSor

    SC

    unlikeothe

    rfALS

    mice.

    (Wan

    get

    al.,20

    05)

    SOD1L1

    26Zde

    lTT

    Trans

    genic

    Promoter:h

    SOD1

    C57

    BL/6

    hSOD1;

    lowex

    pres

    sion

    Inclus

    ions

    :PSOD1:

    PPSC

    PND

    Earliers

    igns

    :25-43

    wee

    ks;

    MNloss

    %:N

    D;e

    nd-

    stag

    epa

    ralysis:

    P

    26-45

    Motor

    coordina

    tion

    impa

    irmen

    tND

    (Watan

    abeet

    al.,

    2005

    )

    SOD1L1

    26Z

    Trans

    genic

    Promoter:h

    SOD1

    ND

    hSOD1

    Inclus

    ions

    :PSOD1:

    PND

    ND

    ND

    Earliersign

    s:48

    wee

    ks;

    MNloss

    %:N

    D51

    -57

    ND

    ND

    (Den

    get

    al.,20

    06)

    SOD1T11

    6XTrans

    genic

    Promoter:h

    SOD1

    C57

    BL/6×

    SJL

    hSOD1,

    lowex

    pres

    sion

    Inclus

    ions

    :PUbiqu

    itin:

    PP

    PND

    Earliersign

    s:41

    wee

    ks;

    MNloss

    %:N

    D;e

    nd-

    stag

    epa

    ralysis:

    P

    ∼43

    Motor

    coordina

    tion

    impa

    irmen

    tHEMs:no

    phen

    otyp

    e;HOMs:

    repo

    rted

    tode

    velopALS

    .Dou

    bletran

    sgen

    ics

    (T11

    6x/W

    ThS

    OD1)

    deve

    lopALS

    phen

    otyp

    e;weigh

    tlos

    sby

    endstag

    e.

    (Han

    -Xiang

    etal.,

    2008

    )

    Sod

    1D83

    G(JAX

    0204

    40)

    ENUpo

    intm

    utan

    tPromoter:m

    Sod

    1C57

    BL/6J

    -C3H

    backcros

    sedto

    C57

    BL/6J

    mSOD1;

    redu

    cedprotein

    leve

    lsan

    dloss

    ofac

    tivity

    Ab

    PP

    PHOMs:

    Earliersign

    s:15

    wee

    ks;U

    MNloss:

    ∼20

    %,b

    y29

    wee

    ks∼23

    %;L

    MNloss:

    ∼23

    %by

    wee

    k15

    -52

    ∼88

    Motor

    coordina

    tion

    impa

    irmen

    tLive

    rtumou

    rs,k

    ypho

    sis,

    redu

    cedbo

    dyweigh

    t(∼4wee

    ks).

    HOMs:

    males

    have

    sign

    ifica

    ntlyredu

    ced

    lifes

    panco

    mpa

    redwith

    that

    offemales

    .

    (Joy

    ceet

    al.,20

    15)

    (B)TDP-43mou

    semod

    els

    hTDP-43WT(JAX

    0166

    08)

    Trans

    genic

    Promoter:m

    Prp

    C57

    BL/6

    >hT

    DP-43an

    d<WTmTDP-

    43,c

    ytotox

    ic∼25

    kDa

    Inclus

    ions

    :PUbiqu

    itin:

    PP

    Ab

    ND

    Earlysign

    sat

    3wee

    ks;n

    ofin

    alMNloss

    ∼4-8

    Mild

    motor

    coordina

    tion

    impa

    irmen

    tHOM

    show

    seve

    remotor

    deficits,b

    rain

    atroph

    yan

    dweigh

    tlos

    s.

    (Xuet

    al.,20

    10)

    hTDP-43WTlines

    3,4,

    21(JAX

    0162

    01)

    Trans

    genic

    Promoter:m

    Prp

    Initiallyon

    mixed

    backgrou

    nd(B6

    SJL

    Fan

    dCD1),

    nowB6SJL

    F

    Red

    uced

    hTDP-43+

    25-

    35kD

    afra

    gmen

    tsInclus

    ions

    :PUbiqu

    itin:

    PTDP-43:

    co-lo

    calises

    with

    Ub

    ND

    PND

    Ab

    ∼15

    ND

    Som

    efoun

    ders

    show

    aggres

    sive

    motor

    phen

    otyp

    e(surviva

    lof

    1-5wee

    ks).

    Later-on

    setlines

    show

    wea

    knes

    san

    dhy

    potonia.

    Weigh

    tlos

    s.

    (Stalling

    set

    al.,

    2010

    )

    hTDP-43WT(JAX

    0179

    07)

    Trans

    genic

    Promoter:m

    Prp

    C57

    BL/6/C3H

    backcros

    sedwith

    C57

    BL/6J

    Nonu

    clea

    rloss

    ofhT

    DP-43.

    Increa

    seof

    hTDP-43

    expres

    sion

    decrea

    seof

    endo

    geno

    usprotein

    Ab

    Ab

    Ab

    Ab

    Nofin

    alMNloss

    ∼10

    4Noor

    mild

    motor

    coordina

    tion

    impa

    irmen

    tPhe

    notypica

    llyno

    rmal.

    Noweigh

    tlos

    s.(Arnoldet

    al.,20

    13;

    Mitche

    llet

    al.,

    2015

    )

    hTDP-43WTlineW3

    Trans

    genic

    Promoter:m

    Thy

    1.2

    C57

    BL/6

    hTDP-43nu

    clea

    rinclus

    ions

    containing

    FUS,S

    C35

    (alsokn

    ownas

    SRSF2)

    andproteins

    invo

    lved

    inRNAmetab

    olism

    Inclus

    ions

    :PUbiqu

    itin:

    PTDP-43:

    Ab

    Ab

    PP

    Earlysign

    sat

    ∼2-

    12wee

    ks;n

    ofin

    alMN

    loss

    Normal

    Motor

    coordina

    tion

    impa

    irmen

    tPhe

    notype

    morese

    vere

    inmales

    than

    females

    .Significan

    tinc

    reas

    ein

    GEM

    bodies

    inMNs.

    (Sha

    net

    al.,20

    10)

    hTDP-43WT(JAX

    0128

    36)

    Trans

    genic

    Promoter:m

    Thy

    1.2

    C57

    BL/6/

    SJL

    ×C57

    BL/6J

    hTDP-43

    HOM Inclus

    ions

    :PUbiqu

    itin:

    PTDP-43:

    rare

    but

    co-lo

    calises

    with

    Ub

    PND

    ND

    Earlysign

    sat

    ∼2-

    56wee

    ks;s

    ignifican

    tMNloss

    inlineTA

    R6/6

    and25

    %MNloss

    inlineTA

    R4/4;

    endstag

    epa

    ralysis:

    P

    HOM

    ∼3HEM

    ∼10

    4Mild

    tose

    vere

    motor

    coordina

    tionim

    pairm

    ent

    Progres

    sive

    motor

    impa

    irmen

    tswith

    fasciculationan

    dsp

    asms

    offacial

    mus

    cle.

    Differen

    cesin

    the

    phen

    otyp

    ebe

    twee

    nTg

    lines

    .

    (Wils

    etal.,20

    10)

    4

    REVIEW Disease Models & Mechanisms (2019) 12, dmm037424. doi:10.1242/dmm.037424

    Disea

    seModels&Mechan

    isms

  • hTDP-43WTline

    W12

    (JAX

    0168

    41)

    Trans

    genic

    Promoter:C

    aMKII

    tTa×

    tet-off

    C57

    BL/6J

    ×C3H

    /HeJ

    hTDP-43;

    loss

    ofnu

    clea

    rmTDP-43an

    dcytoplas

    michT

    DP-43

    Inclus

    ions

    :PUbiqu

    itin:

    PTDP-43:

    co-lo

    calises

    with

    Ub

    PAb

    ND

    UMN(%

    MNloss:N

    D)

    ∼24

    Mild

    tono

    motor

    coordina

    tion

    impa

    irmen

    tBrain

    atroph

    y.SingleTgs

    expres

    slittle

    tono

    hTDP-

    43.D

    egen

    erationof

    neuron

    sin

    DG.

    (Iga

    zet

    al.,20

    11)

    hTDP-43WT

    Trans

    genic

    Promoter:C

    aMKII

    tTa×

    tet-off

    FVB/N

    hTDP-43ov

    erex

    pres

    sin

    forebrain

    Inclus

    ions

    :PUbiqu

    itin:

    PTDP-43:

    P

    PND

    ND

    Earlysign

    sat

    8wee

    ks;n

    ofin

    alMNloss

    ∼70

    Motor

    coordina

    tion

    impa

    irmen

    tLe

    arning

    andmem

    ory

    deficits.

    DisruptionERK

    phos

    phorylation,

    inhibitio

    nGABA

    neurotrans

    mitter.

    (Tsa

    ieta

    l.,20

    10)

    iTDP-43WT/diTDP-

    43WT

    Trans

    genic

    Promoter:C

    aMKII

    tTa×

    tet-off

    FVB/NCr×12

    9S6

    hTDP-43;

    mTDP-43

    downreg

    ulated

    inresp

    onse

    toov

    erex

    pres

    sion

    ofhT

    DP-43

    Inclus

    ions

    :PUbiqu

    itin:

    POccas

    iona

    llyp6

    2/TDP-

    43:P

    (iTDP-43W

    T)

    pTDP-43:

    P(diTDP-

    43WT)

    PND

    ND

    Earlysign

    sat

    ∼4wee

    ksNofin

    alMNloss

    <8-52

    ND

    Twomou

    selines

    :iTDP-43

    WT(exp

    ressingiTDP-43)

    show

    aggres

    sive

    phen

    otyp

    ein

    early

    deve

    lopm

    ents

    tage

    s.diTDP-43WTon

    Dox

    upto

    21da

    yssh

    owlong

    erlifes

    panan

    dslow

    erprog

    ressive

    neurod

    egen

    eration.

    (Can

    nonet

    al.,20

    12)

    hTDP-43WT

    BACtran

    sgen

    icPromoter:m

    Tardb

    pC3H

    ×C57

    BL/6

    hTDP-43mainlyin

    nucleu

    s.Increa

    sedlevelofcytotox

    ic∼25

    kDaC-te

    rminal

    fragm

    ento

    fTDP-43

    Ab

    PND

    PEarlysign

    at∼42

    wee

    ks;

    nofin

    alMNloss

    ND

    Motor

    coordina

    tion

    impa

    irmen

    tPeriphe

    rinag

    greg

    ates

    inthe

    hipp

    ocam

    pus.

    Age

    -as

    sociated

    cogn

    itive

    and

    motor

    deficits.

    (Swarup

    etal.,20

    11)

    TDP-43A31

    5T/

    G34

    8CBACtran

    sgen

    icPromoter:m

    Tardb

    pC3H

    ×C57

    BL/6

    hTDP-43;

    increa

    sedleve

    lof

    cytotoxic∼25

    kDa

    C-terminal

    frag

    men

    tof

    TDP-43

    Inclus

    ions

    :PUbiqu

    itin:

    PTDP-43:

    P

    PND

    PEarlysign

    sat

    ∼42

    wee

    ks;

    nofin

    alMNloss.

    ND

    Motor

    coordina

    tion

    impa

    irmen

    tPeriphe

    rinag

    greg

    ates

    inthe

    hipp

    ocam

    pus.

    Age

    -as

    sociated

    cogn

    itive

    and

    motor

    deficits.

    (Swarup

    etal.,20

    11)

    TARDBPA31

    5T(JAX01

    0700

    )Trans

    genic

    Promoter:m

    Prp

    C57

    BL/6J

    ×CBAthen

    cros

    sedwith

    C57

    BL/6J

    byHatzipe

    tros

    etal.

    (201

    4)

    hTDP-43;

    frag

    men

    ts25

    -35

    kDabu

    tnot

    inde

    tergen

    t-inso

    luble

    phas

    e

    Ubiqu

    itin:

    PTDP-43:

    rare

    PP

    PEarlysign

    sat

    8-12

    wee

    ks;

    MNloss:2

    0%at

    ∼26

    wee

    ks

    13-26

    Motor

    coordina

    tion

    impa

    irmen

    tOrig

    inal

    lineon

    ano

    n-co

    ngen

    icge

    netic

    backgrou

    ndsh

    owed

    cons

    iderab

    lene

    urom

    uscu

    larde

    ficits.

    Hatzipe

    tros

    etal.(20

    14)

    backcros

    sedthelinewith

    C57

    BL/6J

    before

    startin

    gthestud

    y.Motor

    phen

    otyp

    eco

    nfus

    edby

    gutp

    heno

    type

    .Significan

    tse

    xdiffe

    renc

    es.

    (Weg

    orze

    wskaet

    al.,

    2009

    ;Hatzipe

    tros

    etal.,20

    14)

    TDP-43A31

    5Tlines

    23,2

    7,35

    ,61

    (JAX01

    6143

    )

    Trans

    genic

    Promoter:m

    Prp

    C57

    BL/Ntac+

    Ptprc

    andPep

    3(also

    know

    nas

    Pep

    c)allelesfrom

    SJL

    /Jstrain×CD1then

    colony

    rand

    omly

    bred

    toCD1an

    dmixed

    backgrou

    ndmice

    HighhT

    DP-43+

    25-35kD

    afra

    gmen

    tsInclus

    ions

    :PUbiqu

    itin:

    PTDP-43:

    rare

    pTDP-43

    PP

    ND

    Earlysign

    sat

    2-8wee

    ks;

    nofin

    alMNloss

    ∼10

    Mild

    motor

    coordina

    tion

    impa

    irmen

    tLa

    ter-on

    setp

    rogres

    sive

    motor

    phen

    otyp

    e.(Stalling

    set

    al.,

    2010

    )

    TDP-43M33

    7VTrans

    genic

    Promoter:m

    Prp

    C57

    BL/Ntac+

    Ptprc

    andPep

    3alleles

    from

    SJL

    /Jstrain×CD1then

    rand

    omlybred

    toCD1an

    dmixed

    backgrou

    nd

    HighhT

    DP-43+

    25-35kD

    afra

    gmen

    tsInclus

    ions

    :PUbiqu

    itin:

    PTDP-43:

    P

    PND

    ND

    ND

    ∼2-6

    ND

    Trans

    genicfoun

    ders

    show

    aggres

    sive

    motor

    phen

    otyp

    e.

    (Stalling

    set

    al.,

    2010

    )

    TDP-43M33

    7V(JAX01

    7604

    )Trans

    genic

    Promoter:m

    Prp

    C57

    BL/6

    hTDP-43mainlynu

    clea

    r.Dos

    ede

    pend

    ent>hT

    DP-

    43an

    d<WTmTDP-43

    Inclus

    ions

    :PUbiqu

    itin:

    PP

    ND

    ND

    ND

    ∼4

    Motor

    coordina

    tion

    impa

    irmen

    tHEMs:no

    phen

    otyp

    e.HOMs:

    early

    sign

    sat

    40wee

    kswith

    brainatroph

    yan

    dhy

    perpho

    spho

    rylatedTau

    incytoplas

    m.W

    eigh

    tlos

    s.

    (Xuet

    al.,20

    11)

    hTDP-43M33

    7VTrans

    genic

    Promoter:m

    Prp

    C57

    BL/6/C3H

    backcros

    sedwith

    C57

    BL/6J

    hTDP-43;

    nonu

    clea

    rloss

    ofTDP-43

    Ab

    Ab

    Ab

    Ab

    Earlysign

    sat

    40wee

    ks;

    age-de

    pend

    entM

    Nloss

    (%ND)

    ND

    Motor

    coordina

    tion

    impa

    irmen

    tAb

    (Arnoldet

    al.,20

    13)

    TDP-43M33

    7VTrans

    genic

    Promoter:m

    Thy

    1.2

    BL/6/SJL

    ×C57

    BL/6J

    hTDP-43;

    diffe

    rent

    leve

    lof

    expres

    sion

    depe

    ndingon

    thefoun

    der.Fragm

    ents

    25-35kD

    a.Noco

    -loca

    lisationbe

    twee

    nTDP-

    43an

    dstress

    gran

    ules

    Inclus

    ions

    :PUbiqu

    itin:

    PTDP-43:

    Abbu

    tsom

    ep6

    2/TDP-43

    Som

    efoun

    ders

    show

    UBQLN

    2inclus

    ions

    PAb

    Ab

    Earlysign

    sat

    1.5-56

    wee

    ks;M

    Nloss

    %:N

    D;e

    nd-stage

    paralysis:

    ∼4wee

    ks

    HOM

    ∼2.5

    HEM

    ∼71

    Motor

    coordina

    tion

    impa

    irmen

    tSev

    eral

    foun

    ders

    with

    aggres

    sive

    phen

    otyp

    e.Fou

    nderswith

    milder

    phen

    otyp

    esh

    owlate

    onse

    tand

    long

    ersu

    rvival.

    Weigh

    tlos

    s.

    (Jan

    ssen

    set

    al.,

    2013

    )

    Con

    tinue

    d

    5

    REVIEW Disease Models & Mechanisms (2019) 12, dmm037424. doi:10.1242/dmm.037424

    Disea

    seModels&Mechan

    isms

  • Tab

    le1.

    Continued

    Strainna

    me

    Trans

    genic/ge

    ne-

    targeted

    knoc

    k-in/ENU

    Gen

    eticba

    ckgrou

    ndProtein

    Inclus

    ions

    /agg

    rega

    tes

    Gliosis

    MA/M

    DNMJ

    loss

    Final-stage

    dise

    ase

    (terminal

    MNloss)

    Surviva

    l(wee

    ks)

    Beh

    avioural

    analysis

    Other

    phen

    otyp

    esReferen

    ce

    hTDP-43Q33

    1K(JAX01

    7933

    )Trans

    genic

    Promoter:m

    Prp

    C57

    BL6

    /C3H

    backcros

    sedwith

    C57

    BL/6J

    Nonu

    clea

    rloss

    ofTDP-43

    Ab

    Ab

    PP

    Earlysign

    sat

    12wee

    ks;

    LMNloss:∼

    35-40%

    at52

    wee

    ks

    Normal

    Motor

    coordina

    tion

    impa

    irmen

    tMotor

    deficits

    andmus

    cle

    fibrillatio

    n.Misregu

    lated

    cassette

    exon

    sdu

    eto

    mutated

    TDP-43.

    (Arnoldet

    al.,20

    13)

    hTDP-43Q33

    1K-lo

    w(JAX01

    7930

    )Trans

    genic

    Promoter:m

    Prp

    C57

    BL6

    /C3H

    backcros

    sedwith

    C57

    BL/6J

    hTDP-43;

    nonu

    clea

    rloss

    ofTDP-43

    Ab

    Ab

    PP

    Earlysign

    sat

    40wee

    ks;

    LMNloss:∼

    35-40%

    at52

    wee

    ks

    ND

    Motor

    coordina

    tion

    impa

    irmen

    tMotor

    deficits

    andmus

    cle

    fibrillatio

    n.Misregu

    lated

    cassette

    exon

    sdu

    eto

    mutated

    TDP-43.

    (Arnoldet

    al.,20

    13)

    TDP-43Q33

    1K(JAX03

    0157

    )Trans

    genic

    Promoter:m

    Prp

    C57

    BL/6/C3H

    maintaine

    dC57

    BL/6J

    backgrou

    nd

    Nuc

    lear

    loss

    ofhT

    DP-43.

    Q33

    1K-TDP-43mutation

    appe

    arsto

    have

    more

    tend

    ency

    toag

    greg

    atein

    thecytoplas

    m

    Inclus

    ions

    :PUbiqu

    itin:

    PTDP-43:

    endo

    geno

    us

    PPHOM

    PEarlysign

    sat

    3wee

    ks;

    MNloss:7

    0%in

    WT

    xQ33

    1K;e

    nd-stage

    paralysis:

    by8-10

    wee

    ks

    8-10

    Motor

    coordina

    tion

    impa

    irmen

    tSam

    emou

    seas

    Arnoldet

    al.,

    2013

    ,but

    thepa

    per

    focu

    sedon

    doub

    le-m

    utan

    thT

    DP-43WT×hT

    DP-43-

    Q33

    1Kov

    erex

    pres

    sor.No

    gutp

    heno

    type

    .Rap

    iddise

    aseprog

    ression.

    HEM

    Q33

    1Klinesh

    ows

    late-ons

    etag

    e-de

    pend

    ent

    motor

    deficit.

    (Mitche

    llet

    al.,20

    15)

    hTDP-43ΔNLS

    (JAX01

    4650

    )Trans

    genic

    Promoter:C

    aMKII

    tTa×

    tet-off

    C57

    BL/6J

    ×C3H

    /HeJ

    hTDP-43;

    dram

    aticloss

    nuclea

    rWTTDP-43an

    dcytoplas

    michT

    DP-43

    Inclus

    ions

    :PUbiqu

    itin:

    PTDP-43:

    lowbu

    tco

    -loca

    lises

    with

    Ub

    Ab

    Ab

    ND

    Earlysign

    sat

    1-3wee

    ksoffD

    ox;n

    ofin

    alMN

    loss

    ∼24

    Motor

    coordina

    tion

    impa

    irmen

    tTim

    e-de

    pend

    ent

    neurod

    egen

    erationoff

    Dox

    .SingleTgs

    expres

    slittle

    tono

    hTD-43.

    Alte

    red

    expres

    sion

    ofge

    nese.g.

    Tardb

    p,Hnrnp

    a3,e

    tc.

    (Iga

    zet

    al.,20

    11)

    NEFH-tTA

    (JAX02

    8412

    )Trans

    genic

    Crossed

    with

    hTDP-

    43ΔNLS

    from

    Igaz

    etal.,20

    11Promoter:h

    NEFH/

    CaM

    KIItTA×tet-off

    C57

    BL/6J

    ×C3H

    eJF1

    Prese

    nceof

    cytoplas

    mic

    hTDP-43ΔNLS

    isDox

    depe

    nden

    t.Not

    foun

    dwhe

    ncros

    sedwith

    hTDP-

    43WT

    Inclus

    ions

    :PUbiqu

    itin:

    PTDP-43:

    co-lo

    calises

    with

    Ub

    PP

    PEarlysign

    sat

    2wee

    ksoff

    Dox

    ;MNloss:5

    0%by

    6wee

    ksoffD

    ox

    Surviva

    l8-18wee

    ksoff

    Dox

    andup

    to20

    -32

    wee

    kswhe

    nDox

    reintrod

    uced

    Motor

    coordina

    tion

    impa

    irmen

    tDox

    stop

    pedat

    5wee

    ks.

    Line

    also

    cros

    sedwith

    hTDP-43WTfro

    mIgaz

    etal.,20

    11to

    confirm

    that

    cytoplas

    micinclus

    ions

    wererelatedto

    ΔNLS

    line.

    Sup

    pres

    sion

    ofhT

    DP-

    43ΔNLS

    byre-in

    trod

    ucing

    Dox

    partially

    rescue

    sph

    enotyp

    e.Weigh

    tlos

    s.

    (Walke

    ret

    al.,20

    15;

    Spilleret

    al.,

    2018

    )

    TDP-43WT

    Hum

    ange

    nomic

    sequ

    ence

    targeted

    into

    Ros

    a26

    [Gt(ROSA26

    )Sor]

    locu

    sin

    asing

    leco

    pywith

    aC-

    term

    inal

    Ype

    tflu

    ores

    cent

    tag

    C57

    BL/6×

    C57

    BL/6J

    hTDP-43ex

    pres

    sion

    lower

    than

    endo

    geno

    usmTDP-

    43

    Ab

    Ab

    Ab

    Ab

    WTco

    mpa

    rablewith

    Ntg

    Normal

    Indistingu

    isha

    blefrom

    Ntg

    ND

    (Gordo

    net

    al.,20

    19)

    hTDP-43A31

    5TGen

    e-targeted

    knoc

    k-in

    End

    ogen

    ous

    Promoter:m

    Tardb

    p

    129S

    2/12

    9P2/

    Ola×C57

    BL/

    6NTac

    Hum

    ancD

    NAge

    netargeted

    into

    mou

    seTardb

    plocu

    s;alteratio

    nof

    3′UTR

    aligning

    impo

    rtan

    cefor

    autoregu

    latio

    n

    Inclus

    ions

    :PUbiqu

    itin:

    PTDP:c

    o-loca

    lises

    with

    Ub

    p62:

    Ab

    ND

    Ab

    ND

    Earlysign

    sat

    ∼12

    -20wee

    ks.M

    Nloss:1

    0%at

    65wee

    ks

    Normal

    Mild

    motor

    coordina

    tion

    impa

    irmen

    tPre-sym

    ptom

    aticmod

    el.

    Red

    uctio

    nof

    CD36

    .Alte

    redex

    pres

    sion

    ofge

    nesinvo

    lved

    ince

    llde

    athan

    dlipid

    metab

    olism.

    (Strible

    tal.,

    2014

    )

    TDP-43M33

    7V(JAX02

    9266

    )Hum

    ange

    nomic

    sequ

    ence

    targeted

    into

    Ros

    a26

    [Gt(ROSA26

    )Sor]

    locu

    sin

    asing

    leco

    pywith

    aC-

    term

    inal

    Ype

    tflu

    ores

    cent

    tag

    C57

    BL/6×

    C57

    BL/6J

    hTDP-43ex

    pres

    sion

    lower

    than

    endo

    geno

    usmTDP-

    43

    Ab

    Ab

    Ab

    PHOMs:

    early

    sign

    sat

    ∼26

    wee

    ks;

    symptom

    aticmiceat

    ∼39

    wee

    ks;nofin

    alMN

    loss

    Normal

    Motor

    coordina

    tion

    impa

    irmen

    tWeigh

    tred

    uctio

    nin

    male

    HOM.C

    ytop

    lasm

    icmisloca

    lisationof

    hTDP-

    43ob

    served

    inem

    bryo

    nic

    stem

    cell-de

    rived

    MNs.

    (Gordo

    net

    al.,20

    19)

    Tardb

    pQ33

    1K(JAX03

    1345

    )Gen

    e-targeted

    knoc

    k-in

    Promoter:m

    Tardb

    pC57

    BL/6J

    mTDP-43;

    TDP-43GOF.

    45%

    increa

    sein

    nuclea

    rTDP-43in

    mutan

    t,im

    paire

    dau

    toregu

    latio

    n

    Ab

    Ab

    Ab

    Ab

    Earlysign

    sat

    ∼20

    wee

    ks;

    nofin

    alMNloss

    ∼80

    Mild

    tono

    motor

    coordina

    tion

    impa

    irmen

    tat2

    0an

    d24

    wee

    ks.C

    ognitivean

    dmem

    oryim

    pairm

    ent

    Abe

    rran

    tbeh

    aviour,

    hype

    rpha

    gia,

    high

    weigh

    t.Phe

    notypic

    heteroge

    neity.

    Significan

    texp

    ression/

    splicingdiffe

    renc

    esin

    brain,

    SCby

    20wee

    ks.

    (White

    etal.,20

    18)

    Tardb

    pQ33

    1KGen

    e-targeted

    knoc

    k-in

    Promoter:m

    Tardb

    pC57

    BL/6J

    mTDP-43;

    TDP-43ga

    in-of-

    splicingfunc

    tion

    ND

    ND

    ND

    ND

    ND

    ND

    ND

    Skipp

    ingof

    cons

    erve

    dex

    ons.

    (Fratta

    etal.,20

    18)

    6

    REVIEW Disease Models & Mechanisms (2019) 12, dmm037424. doi:10.1242/dmm.037424

    Disea

    seModels&Mechan

    isms

  • Tardb

    pQ10

    1X(JAX

    0198

    99)

    ENUpo

    intm

    utan

    tPromoter:mTardb

    pC57

    BL/6J

    ×C3H

    /HeH

    mTDP-43;

    nodiffe

    renc

    ein

    proteinleve

    lbetwee

    nWT/

    mutan

    tTDP-43

    Ab

    Ab

    Ab

    Ab

    Earlysign

    sat

    32-61wee

    ks;n

    ofin

    alMNloss

    ND

    Mild/nomotor

    coordina

    tion

    impa

    irmen

    tLo

    ssof

    body

    tone

    .Noweigh

    tloss.A

    berran

    texo

    ninclus

    ion.

    (Ricke

    ttset

    al.,20

    14)

    Tardb

    pF21

    0I(BRC#

    GD00

    0108

    )ENUpo

    intm

    utan

    tPromoter:m

    Tardb

    pOnC57

    BL/6J

    embryo

    nicda

    y18

    .5;v

    iableHOM

    onC57

    BL/6J

    -DBA/2J

    mTDP-43;

    TDP-43LO

    F(shift

    towards

    exon

    inclus

    ion),

    cryp

    ticex

    on.R

    educ

    edRNAbind

    ing

    Ab

    Ab

    Ab

    Ab

    Ab

    ND

    Ab

    HOMs:

    embryo

    niclethal.

    TDP43

    LOFeffectson

    expres

    sion

    andsp

    licing.

    (Fratta

    etal.,20

    18)

    Tardb

    pM32

    3K(BRC#

    GD00

    0110

    )

    ENUpo

    intm

    utan

    tPromoter:m

    Tardb

    pOnC57

    BL/6J

    embryo

    niclethal;

    viab

    leHOM

    onC57

    BL/6J

    -DBA/

    2J

    mTDP-43;

    TDP-43GOF

    (increa

    sedex

    onex

    clus

    ion),s

    kipticex

    on.

    Nonu

    clea

    rde

    pletion.

    Increa

    sedTardb

    pintron

    7retention

    Inclus

    ions

    :PUbiqu

    itin:

    PTDP-43:

    Ab

    ND

    ND

    Ab

    Earlysign

    sat

    ∼52

    wee

    ks;

    MNloss:2

    8%at

    104wee

    ks

    Normal

    Motor

    coordina

    tion

    impa

    irmen

    tTDP43

    GOFeffectson

    expres

    sion

    andsp

    licing.

    (Fratta

    etal.,20

    18)

    (C)FUSmou

    semod

    els

    hFUS(+/+)

    hFUS(+/−)

    (JAX01

    7916

    )

    Trans

    genic

    Promoter:m

    Prp

    C57

    BL/6/SJL

    Diffus

    ecytoplas

    michF

    US

    staining

    ,noloss

    ofnu

    clea

    rFUS.h

    FUS

    decrea

    sesWTmFUS.

    Tox

    icGOFan

    ddo

    se-

    depe

    nden

    ttox

    icity

    Inclus

    ions

    :PUbiqu

    itin:

    PFUS:P

    PP

    PHOMs:

    MNloss:6

    0%;

    end-stag

    epa

    ralysis:

    at∼11

    wee

    ks

    10-104

    Motor

    coordina

    tion

    impa

    irmen

    tHEMs:

    nomotor

    phen

    otyp

    e,mild

    MNloss

    andgliosisat

    104wee

    ks.H

    OMs:

    aggres

    sive

    phen

    otyp

    e,hind

    limbpa

    ralysisan

    drapiddise

    aseprog

    ression

    at10

    -13wee

    ks.

    (Mitche

    llet

    al.,20

    13)

    hgFUS-W

    Tline88

    BACtran

    sgen

    icPromoter:h

    FUS

    C57

    BL/6

    Leve

    lsof

    hFUSsimilarto

    norm

    alen

    doge

    nous

    mou

    seleve

    ls.m

    FUS

    endo

    geno

    usleve

    lsdo

    wn

    dueto

    auto-reg

    ulation.

    Ab

    Ab

    ND

    PAb

    Normal

    Non

    -significan

    tmotor

    orco

    gnitive

    abno

    rmalities

    Mod

    estred

    uctio

    nin

    α-m

    otor

    axon

    san

    dNMJs

    by24

    mon

    ths.

    (Lóp

    ez-Eraus

    kin

    etal.,20

    18)

    CAG-FUSWT(JAX

    0278

    98)

    Trans

    genic

    Promoter:C

    AG

    CAG-Z-FUS-IRES-

    EGFPWTmice

    areC57

    BL6

    /ICR

    CAG-Z-FUS-

    IRES-EGFPWT

    cros

    sedwith

    Meo

    x2-C

    remice

    (129

    S4/

    SvJae

    ×C57

    BL/6)

    tocrea

    teCAG-

    FUSWT

    hFUSno

    tove

    rtly

    misloca

    lised

    Ab

    PP

    PAb

    2-4

    Motor

    coordina

    tion

    impa

    irmen

    t10

    0%of

    theCAG-FUS_W

    Tdieat

    less

    than

    postna

    tal

    day30

    .Significan

    tbod

    yweigh

    tlos

    s.Alte

    redge

    neex

    pres

    sion

    .

    (Sep

    hton

    etal.,

    2014

    )

    hFUS-W

    T(JAX

    0207

    83)

    Trans

    genic

    Promoter:m

    PrP

    C57

    BL/6×

    SJL

    F2

    then

    C57

    BL/6

    hFUS

    Ab

    ND

    Ab

    ND

    ND

    ∼30

    ND

    ND

    (Tibsh

    irani

    etal.,

    2015

    )hF

    US-R

    495X

    (JAX

    0197

    28)

    Trans

    genic

    Promoter:m

    PrP

    C57

    BL/6×

    SJL

    F2

    then

    C57

    BL/6

    Highleve

    lsof

    cytoplas

    mic

    hFUS

    Ab

    Ab

    ND

    PEarlysign

    sat

    ∼34

    wee

    ks;

    nofin

    alMNloss

    Som

    eHEM

    ∼17

    /HOM

    ∼8.

    Ave

    rage

    show

    sno

    rmal

    survival

    Ab

    Som

    eHEM

    died

    prem

    aturely

    dueto

    intestinal

    swelling.

    Dec

    reas

    eintra

    nscriptio

    nal

    activity

    inside

    neuron

    s.

    (Tibsh

    irani

    etal.,

    2015

    )

    hgFUS-R

    521C

    line

    10BACtran

    sgen

    icPromoter:h

    FUS

    C57

    BL/6

    Leve

    lsof

    hFUSsimilarto

    norm

    alen

    doge

    nous

    mou

    seleve

    ls.m

    FUS

    endo

    geno

    usleve

    lsdo

    wn

    dueto

    auto-reg

    ulation.

    Ab

    PND

    PMild

    MNde

    gene

    ratio

    nby

    24mon

    ths

    Normal

    Motor

    coordina

    tion

    impa

    irmen

    t.Lo

    ssof

    grip

    streng

    thstartin

    gby

    8mon

    ths.

    Progres

    sive

    cogn

    itive

    impa

    irmen

    t.

    Age

    -dep

    ende

    ntloss

    ofα-

    motor

    axon

    s.Syn

    apsis

    alteratio

    ns.A

    lteredge

    neex

    pres

    sion

    .

    (Lóp

    ez-Eraus

    kin

    etal.,20

    18)

    FUS-R

    521C

    (Flag

    tagg

    ed)(JAX

    0264

    06)

    Trans

    genic

    Promoter:S

    yrian

    hamster

    prion

    (Sha

    PrP)

    C57

    BL/6

    SJL

    ×C57

    BL/6

    Leve

    lofh

    FUS-R

    521C

    inbrainan

    dSCsimilarto

    endo

    geno

    usFUS.L

    evels

    ofen

    doge

    nous

    FUSse

    emto

    beincrea

    sed

    Inclus

    ions

    :PUbiqu

    itin:

    ND

    FUS:P

    PP

    PEarlysign

    sat

    5-14

    wee

    ks;

    MNloss:H

    EM

    >50

    %at

    4-12

    wee

    ks

    HEM

    23-46

    Motor

    coordina

    tion

    impa

    irmen

    tIncrea

    sedγH

    2AXleve

    lsin

    cortex

    andSC(C

    hat+);

    increa

    sedATF3indica

    ting

    increa

    sedDNAda

    mag

    e.Sev

    ereim

    pairm

    entin

    BDNF-TrkBsign

    alling.

    (Qiu

    etal.,20

    14)

    hgFUS-R

    521H

    line9

    BACtran

    sgen

    icPromoter:h

    FUS

    C57

    BL/6

    Leve

    lsof

    hFUSsimilarto

    norm

    alen

    doge

    nous

    mou

    seleve

    ls.m

    FUS

    endo

    geno

    usleve

    lsdo

    wn

    dueto

    auto-reg

    ulation.

    Ab

    PND

    PMild

    MNde

    gene

    ratio

    nby

    24mon

    ths

    Normal

    Motor

    coordina

    tion

    impa

    irmen

    t.Lo

    ssof

    grip

    streng

    thstartin

    gby

    8mon

    ths.

    Progres

    sive

    cogn

    itive

    impa

    irmen

    t

    Age

    -dep

    ende

    ntloss

    ofα-

    motor

    axon

    s.Syn

    apsis

    alteratio

    ns.A

    lteredge

    neex

    pres

    sion

    .

    (Lóp

    ez-Eraus

    kin

    etal.,20

    18)

    CAG-FUS-R

    521G

    (JAX02

    8021

    )Trans

    genic

    Promoter:C

    AG

    CAG-Z-FUS-IRES-

    EGFPR52

    1GmiceareC57

    BL6

    /ICRCAG-Z-FUS-

    IRES-EGFP-

    R52

    1Gcros

    sed

    with

    Meo

    x2-C

    remice(129

    S4/

    SvJae

    ×C57

    BL6

    )

    hFUSR52

    1Gno

    tove

    rtly

    misloca

    lised

    Ab

    PP

    PAbbu

    talte

    ratio

    nin

    dend

    riticbran

    ches

    obse

    rved

    inCAG-

    FUSR52

    1GUMNan

    dLM

    N

    50-70%

    dieat

    <4wee

    ks;

    remaining

    30-50%

    reac

    had

    ulthoo

    d

    Inthe50

    -70%

    that

    dieea

    rly:

    seve

    remotor,c

    o-ordina

    tionim

    pairm

    ent.In

    theremaining

    30-50%

    :mod

    eratemotor,c

    o-ordina

    tionim

    pairm

    ent

    Inthe50

    -70%

    that

    dieea

    rly:

    similarto

    CAG-FUSWT

    butn

    oalteredge

    neex

    pres

    sion

    .In

    theremaining

    30-50%

    :redu

    cedbo

    dyweigh

    t,im

    pairm

    ents

    inso

    ciab

    ility,

    impa

    iredforelim

    bs,

    redu

    ctionin

    loco

    motion

    (Sep

    hton

    etal.,

    2014

    )

    Con

    tinue

    d

    7

    REVIEW Disease Models & Mechanisms (2019) 12, dmm037424. doi:10.1242/dmm.037424

    Disea

    seModels&Mechan

    isms

  • Tab

    le1.

    Continued

    Strainna

    me

    Trans

    genic/ge

    ne-

    targeted

    knoc

    k-in/ENU

    Gen

    eticba

    ckgrou

    ndProtein

    Inclus

    ions

    /agg

    rega

    tes

    Gliosis

    MA/M

    DNMJ

    loss

    Final-stage

    dise

    ase

    (terminal

    MNloss)

    Surviva

    l(wee

    ks)

    Beh

    avioural

    analysis

    Other

    phen

    otyp

    esReferen

    ce

    tocrea

    teCAG-

    FUSR52

    1Gac

    tivities

    butn

    oaltered

    gene

    expres

    sion

    .ΔNLS

    -hFUS(m

    yc-

    tagg

    ed)

    Trans

    genic

    Promoter:m

    Thy

    1.2

    BDF1×

    C57

    BL/6

    Cytop

    lasm

    ichF

    US

    Inclus

    ions

    :PUbiqu

    itin:

    PG3B

    Pprotein:

    P

    P(FC)

    ND

    ND

    Earlysign

    sat

    12wee

    ks;

    nofin

    alMNloss

    ∼60

    Motor

    coordina

    tion

    impa

    irmen

    tRed

    uced

    GEM

    numbe

    rsin

    cortex

    ,but

    noMNloss.

    Age

    -dep

    ende

    ntph

    enotyp

    e.

    (Shiihas

    hiet

    al.,

    2016

    )

    Fus

    ΔNLS

    /+/Fus

    ΔNLS

    /ΔNLS

    Gen

    etargeted

    Promoter:m

    FUS

    C57

    BL/6.

    Also

    cros

    sedwith

    Cha

    t-Cre

    line

    (129

    S6/

    SVEvT

    ac)

    mFUS;inc

    reas

    eof

    ADMA-

    FUSin

    cytoplas

    man

    dnu

    cleu

    s

    Inclus

    ions

    :nolarge

    Ubiqu

    itin:

    Pp6

    2:Ab

    PND

    PEarlysign

    sat

    ∼40

    wee

    ks;

    HETs:

    MN

    loss

    ∼30

    %;

    HOMs:

    MNloss

    50%

    (new

    born)

    HET:∼

    88;

    HOM:n

    eona

    tal

    lethality

    Motor

    coordina

    tion

    impa

    irmen

    tMicecros

    sedwith

    Cha

    t-Cre

    micesh

    owde

    laye

    dMN

    dege

    neratio

    n.Alte

    ratio

    nof

    gene

    sinvo

    lved

    inmye

    linationan

    din

    seve

    ral

    FUSbind

    ingpa

    rtne

    rs.

    Defec

    tsin

    Sch

    wan

    nce

    lls.

    (Sce

    kic-Zah

    irovic

    etal.,20

    16,2

    017)

    τOFF/O

    NhF

    US-

    P52

    5LGen

    etargeted

    into

    the

    Map

    tloc

    us.

    Promoter:m

    Map

    t(silent

    untilac

    tivated

    byCre-m

    ediated

    reco

    mbina

    tion)

    Ola/129

    /C57

    BL/6J

    hFUS;e

    ndog

    enou

    smFUSin

    nucleu

    s,no

    sign

    ifica

    ntch

    ange

    inex

    pres

    sion

    ofmFUSan

    dmTau

    inHET.

    Nointeractionbe

    twee

    nmFUSan

    dhF

    US,

    possible

    toxicGOFof

    hFUS

    Inclus

    ions

    :nolarge

    Ubiqu

    itin:

    Ab

    FUS:c

    ytop

    lasm

    icloca

    lisation

    PP

    PEarlysign

    sat

    ∼4wee

    ks;

    MNloss:p

    rogres

    sive

    ,23

    .6%

    at∼52

    wee

    ks

    ND

    ND

    hFUSWTlinecrea

    tedas

    control(no

    phen

    otyp

    e).

    Crossed

    τ-hF

    US/τOFF×Prm

    1-Cre

    tocrea

    teτO

    Nlines

    .Cross

    τOFF×Cha

    t-Cre

    show

    edthat

    expres

    sion

    ofmutan

    thF

    USin

    MNsissu

    fficien

    tforcell-a

    uton

    omou

    smotor

    dege

    neratio

    n.

    (Sha

    rmaet

    al.,20

    16)

    τOFF/O

    NhF

    US-

    R52

    1CGen

    etargeted

    into

    the

    Map

    t loc

    us.

    Promoter:m

    Map

    t(silent

    untilac

    tivated

    byCre-m

    ediated

    reco

    mbina

    tion)

    Ola/129

    /C57

    BL/6J

    hFUS;e

    ndog

    enou

    smFUSin

    nucleu

    s,no

    sign

    ifica

    ntch

    ange

    inex

    pres

    sion

    ofmFUSan

    dmTau

    inHET.

    Nointeractionbe

    twee

    nmFUSan

    dhF

    US,

    possible

    toxicGOFof

    hFUS

    Inclus

    ions

    :nolarge

    Ubiqu

    itin:

    Ab

    FUS:c

    ytop

    lasm

    icloca

    lisation

    PP

    PEarlysign

    sat

    ∼8wee

    ks;

    MNloss:p

    rogres

    sive

    ,18

    .6%

    at∼52

    wee

    ks

    ND

    Mild

    motor

    coordina

    tion

    impa

    irmen

    thF

    USWTlinecrea

    tedas

    controlw

    ithno

    phen

    otyp

    e.Crossed

    τ-hF

    US/

    τOFF×Prm

    1-Cre

    tocrea

    teτO

    Nlines

    .Cross

    τOFF×Cha

    t-Cre

    show

    edthat

    expres

    sion

    ofmutan

    thF

    USin

    MNsissu

    fficien

    tforcell-a

    uton

    omou

    smotor

    dege

    neratio

    n.

    (Sha

    rmaet

    al.,20

    16)

    FUSDelta14

    (EMMA

    EM:111

    06)

    Gen

    etargeted

    and

    partialh

    uman

    isation

    Promoter:m

    FUS

    C57

    BL/6N

    /C57

    BL/

    6Jm/hFUS;m

    isloca

    lisationof

    FUSDelta14

    .Equ

    ivalen

    ten

    doge

    nous

    leve

    lofF

    US

    proteinin

    both

    WTan

    dFUSDelta14

    Ab

    ND

    ND

    PEarlysign

    sat

    48wee

    ks;

    MNloss:2

    0%at

    78wee

    ks

    <88

    Mild

    motor

    coordina

    tion

    impa

    irmen

    tTox

    icGOFof

    mutan

    tFUS.

    Rec

    ruitm

    ento

    fmutan

    tFUSinto

    SG

    obse

    rved

    infib

    roblas

    tsof

    both

    human

    /miceca

    rrying

    FUSDelta14

    mutation.

    Alte

    red

    expres

    sion

    ofge

    nes

    enco

    ding

    mito

    chon

    drial/

    ribos

    omal

    proteins

    .

    (Dev

    oyet

    al.,20

    17)

    (D)UBQLN

    2mou

    semod

    els

    UBQLN

    2WTline

    358

    Trans

    genic

    Promoter:m

    Thy

    1.2

    C57

    BL/6C

    3/C57

    BL/

    6J.T

    hencros

    sed

    with

    C57

    BL/6J

    hUBQLN

    2;low

    overex

    pres

    sion

    ofhu

    man

    UBQLN

    2

    Ab

    Variable

    Ab

    Ab

    Nofin

    alMNloss

    ND

    Nomotor

    coordina

    tion

    impa

    irmen

    tDiffus

    eTDP-43in

    nucleu

    srather

    than

    cytoplas

    m.

    Mild

    neuron

    loss

    inhipp

    ocam

    pus–toxicity.

    (Leet

    al.,20

    16)

    UBQLN

    2WTline

    356(JAX02

    9970

    )Trans

    genic

    Promoter:m

    Thy

    1.2

    C57

    BL/6C

    3/C57

    BL/

    6J.T

    hencros

    sed

    with

    C57

    BL/6J

    hUBQLN

    2;high

    overex

    pres

    sion

    ofhu

    man

    UBQLN

    2

    Ab

    Variable

    Ab

    Ab

    Nofin

    alMNloss

    ND

    Nomotor

    coordina

    tion

    impa

    irmen

    tDiffus

    eTDP-43in

    nucleu

    srather

    than

    cytoplas

    m.

    Mild

    neuron

    loss

    inhipp

    ocam

    pus–toxicity.

    (Leet

    al.,20

    16)

    UBQLN

    2P49

    7HTrans

    genic

    Promoter:

    hUBQLN

    2

    C57

    BL/6×

    SJL

    cros

    sSim

    ilarex

    pres

    sion

    leve

    lhU

    BQLN

    2P49

    7Han

    dmou

    seprotein.

    Mutated

    proteinbind

    sprotea

    some

    andsu

    bstrates

    ,affe

    cting

    theclea

    ranc

    eof

    ubiquitin

    ated

    proteins

    Inclus

    ions

    :PUbiqu

    itin:

    Pp6

    2:P

    UBQLN

    2:P

    OPTN:P

    TDP-43:

    Ab

    ND

    ND

    ND

    Earliersign

    s:∼4wee

    ks;

    nofin

    alMNloss

    ∼70

    Mild

    cogn

    itive

    andtempo

    ral

    mem

    orybu

    tnomotor

    coordina

    tionim

    pairm

    ent

    Obs

    erve

    d‘den

    dritic

    spinop

    athy

    ’with

    protein

    inclus

    ions

    (den

    dritic

    spines

    ).Trans

    gene

    onY

    chromos

    omeso

    onlymale

    mice.

    (Gorrie

    etal.,20

    14)

    UBQLN

    2P49

    7S_3

    /P50

    6T_6

    (JAX

    0299

    6802

    9969

    )

    Trans

    genic

    Promoter:m

    Thy

    1.2

    C57

    BL/6C

    3/C57

    BL/

    6J.T

    hencros

    sed

    with

    C57

    BL/6J

    hUBQLN

    2;inclus

    ions

    increa

    sewith

    age

    Increa

    sedmutan

    tand

    endo

    geno

    usUBQLN

    2at

    theen

    dstag

    e

    Inclus

    ions

    :PUbiqu

    itin:

    PThioflavinS:P

    TDP-43:

    P

    PP

    PEarliersign

    s:∼4wee

    ks;

    MNloss:2

    0%for

    P49

    7S_3

    and15

    %for

    P50

    6T_6

    at∼34

    wee

    ks

    ∼35

    /43

    Motor

    coordina

    tion

    impa

    irmen

    tand

    mild

    mem

    oryde

    ficits

    Red

    uctio

    nof

    nuclea

    rTDP-43

    andTDP-43+

    clus

    ters

    incytoplas

    m,som

    etim

    esco

    -loca

    lised

    with

    UBQLN

    2.Lo

    ssof

    neuron

    sin

    hipp

    ocam

    pus.

    (Leet

    al.,20

    16)

    8

    REVIEW Disease Models & Mechanisms (2019) 12, dmm037424. doi:10.1242/dmm.037424

    Disea

    seModels&Mechan

    isms

  • UBQLN

    2P52

    0TGen

    etargeted

    Promoter:m

    Ubq

    ln2

    C57

    BL/6J

    NTac

    mUBQLN

    2;ob

    served

    redu

    cedbind

    ingof

    mutan

    tUBQLN

    2to

    HSP70

    Inclus

    ions

    :PUbiqu

    itin:

    PND

    ND

    ND

    Earliersign

    s:∼39

    wee

    ks;

    nofin

    alMNloss

    ND

    Cog

    nitivebu

    tnomotor

    coordina

    tionim

    pairm

    ent.

    UBQLN

    2-P52

    0×R6/2

    show

    edincrea

    sedHTT,

    which

    co-lo

    calised

    with

    UBQNL2

    dueto

    itsLO

    F.

    (Hjerpeet

    al.,20

    16)

    (E)VAPBmou

    semod

    els

    hVAPBWT

    Trans

    genic(w

    ithIRES-

    EGFPrepo

    rter

    clon

    edafterSTOP

    codo

    nVAPB)

    Promoter:m

    Thy

    1.2

    C57

    BL/6×

    C57

    BL/6

    hVAPB;inc

    reas

    edex

    pres

    sion

    Ab

    ND

    ND

    ND

    Nofin

    alMNloss

    Normal

    Nosign

    ifica

    ntdiffe

    renc

    esbe

    twee

    nhV

    APBWTan

    dNtg

    ND

    (Aliaga

    etal.,20

    13)

    hVAPBP56

    STrans

    genic(w

    ithIRES-

    EGFPrepo

    rter

    clon

    edafterSTOP

    codo

    nVAPB)

    Promoter:m

    Thy

    1.2

    C57

    BL/6×

    C57

    BL/6

    hVAPB;inc

    reas

    edex

    pres

    sion

    but

    sign

    ifica

    ntlyless

    protein

    compa

    redwith

    hVAPB

    WT

    Inclus

    ions

    :PUbiqu

    itin:

    PVAPB:P

    p62:

    P

    ND

    ND

    ND

    Earliersign

    s:∼9-52

    wee

    ks;M

    Nloss:

    60%

    at78

    wee

    ks

    ND

    Mild

    motor

    coordina

    tion

    impa

    irmen

    tC-bou

    ton-med

    iated

    mus

    carin

    icrece

    ptor

    func

    tionsign

    ifica

    ntly

    comprom

    ised

    .

    (Aliaga

    etal.,20

    13)

    Vap

    bP56

    S(JAX

    0283

    60)

    Gen

    etargeted

    Promoter:m

    Vap

    bC57

    BL/6×

    C57

    BL/

    6NCrl

    mVap

    b;similarleve

    lsas

    WT

    (less

    proteinin

    HOM

    than

    HET).VAPBtra

    nsloca

    ted

    from

    ERto

    autoph

    agos

    ome

    Inclus

    ions

    :PUbiqu

    itin:

    PVAPB:P

    ND

    Mild

    Mild

    Earliersign

    s:∼48

    wee

    ks;

    nofin

    alMNloss

    ND

    Mild

    motor

    coordina

    tion

    impa

    irmen

    tAge

    -dep

    ende

    ntERstress.

    (Larroqu

    ette

    etal.,

    2015

    )

    (F)VCPmou

    semod

    els

    hVCP-R

    155H

    Trans

    genic(p97

    /VCP-

    WT)

    Promoter:m

    uscle

    crea

    tinekina

    se(m

    MCK)

    C57

    BL/6

    hVCP

    Inclus

    ions

    :PUbiqu

    itin:

    PVCP:A

    b

    ND

    PND

    Earliersign

    s:∼24

    wee

    ks;

    nofin

    alMNloss

    ND

    Mild

    motor

    coordina

    tion

    impa

    irmen

    tND

    (Weihl

    etal.,20

    07)

    hVCP-R

    155H

    Trans

    genic

    Promoter:C

    MV-

    enha

    nced

    chicke

    nβ-

    actin

    SJL

    xC57

    /C57

    BL/6

    hVCP

    Inclus

    ions

    :Ab

    Ubiqu

    itin:

    PTDP-43:

    P

    Mild

    PND

    Earliersign

    s:∼12

    wee

    ks;

    nofin

    alMNloss

    ∼66

    Motor

    coordina

    tion

    impa

    irmen

    tProgres

    sive

    mus

    cle

    wea

    knes

    s.Sev

    ere

    osteop

    enia(foc

    allytic

    and

    sclerotic

    lesion

    s).

    (Cus

    teret

    al.,20

    10)

    hVCP-A23

    2ETrans

    genic

    Promoter:C

    MV-

    enha

    nced

    chicke

    nβ-

    actin

    SJL

    xC57

    /C57

    BL/6

    hVCP

    Inclus

    ions

    :Ab

    Ubiqu

    itin:

    PTDP-43:

    P

    Mild

    PND

    Earliersign

    s:∼12

    wee

    ks;

    nofin

    alMNloss

    ∼66

    Motor

    coordina

    tion

    impa

    irmen

    t,em

    otiona

    lan

    dco

    gnitive

    impa

    irmen

    t

    Progres

    sive

    mus

    cle

    wea

    knes

    s.Sev

    ere

    osteop

    enia(foc

    allytic

    and

    sclerotic

    lesion

    s).

    (Cus

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    REVIEW Disease Models & Mechanisms (2019) 12, dmm037424. doi:10.1242/dmm.037424

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  • In contrast, only one of the published UBQLN2 transgenic lines,carrying the P497S mutation, which disturbs proteasomaldegradation, shows motor impairment, mild MN loss (20%) andcytoplasmic aggregates positive for ubiquitin and TDP-43 (Le et al.,2016) (Table 1D). The only mouse model expressing a mutatedVAPB protein has a progressive phenotype resulting in ∼60% MNloss by 78 weeks of age (Aliaga et al., 2013) (Table 1E), whereasresults are mixed for the VCP transgenic models (Table 1F). Despitethe variability in phenotype presentation, transgenic mice remain acritical resource for understanding neurodegeneration but, like allmouse models, they have generic characteristics we need to take intoaccount, as discussed below.

    Site of insertionTransgene DNA is usually microinjected into fertilised eggs andrandomly inserts into the host mouse genome. This can lead toinsertional mutagenesis from disrupting a host gene, producing anaberrant phenotype, which is why multiple founder lines fromindependent transgenic embryos are studied – to be confident thatthe common phenotypes arise from the transgene. Almost alltransgenic lines in Table 1 do not have information on the insertionsite, as is the case for the vast majority of transgenic models ofneurodegenerative disease (Tosh et al., 2017; Goodwin et al., 2017).Fortunately, in SOD1G93Amice, the transgene insertion site does notdisrupt a known gene (Srivastava et al., 2014; Achilli et al., 2005).

    Transgene copy number and gene expressionTransgenic DNA tends to concatemerise as it inserts into the genome,leading to multiple copies of the exogenous sequence. This results inthe overexpression of the protein of interest, often leading toaccelerated phenotypes. Furthermore, a caveat to studying transgenicmice arises from the development of aberrant phenotypes due tooverexpression. The SOD1G93A model used most commonly carries∼25 copies of the human transgene, resulting in overexpression of theprotein (Gurney et al., 1994; Shibata, 2001), with MN degenerationprogressing rapidly: disease onset occurs at∼90 days and the humaneendpoint occurs by ∼130 days of age, depending on the geneticbackground of themouse. However, transgenic mice expressingwild-type human SOD1 at a similar level to mice expressing the mutanttransgene have neurological phenotypes likely arising fromoverexpression and not from mutation, including spinal cordvacuolation with early signs of paresis in one or more limbs(Jaarsma et al., 2000) and evenMN loss (Graffmo et al., 2013). Thus,the ideal controls for mutant transgenic mice are transgenic animalsexpressing thewild-type transgene at similar levels to themutantmiceto control for the effects of overexpression per se. However, the wild-type human SOD1 transgenic lines are not without problems. Forexample, transgene insertion sites have not been assessed, andalthough they develop phenotypes relevant to MN disease, these aremore profound in some of the mutant SOD1 transgenic lines, such asthe SOD1G93A model. Nevertheless, a large proportion of ALSstudies in mutant transgenic mice do not use wild-type transgeniccontrols, and this is an option that should at least be considered forfuture work.Some genes are highly dosage sensitive and a subtle deviation

    from the physiological levels leads to aberrant phenotypes, evenwhen the protein product is wild type. Many of the RNA-bindingproteins that cause ALS when mutated belong to this category,including TDP-43 and FUS (Table 1B,C). For example, transgenicmice overexpressing wild-type human TARDBP (from a Thy1.2promoter) by 1.2× to 2× fold over the endogenous gene level have25% MN loss with rare cytoplasmic inclusions containing TDP-43

    (Wils et al., 2010). Overexpression of human wild-type FUS (underthe mouse prion promoter) results in aggregation of human FUSprotein and 60% loss of MNs in homozygous transgenic mice,leading to a more severe phenotype in homozygotes than inhemizygotes (Mitchell et al., 2013) (Table 1C). Indeed, RNA-binding proteins such as TDP-43 often control the expression levelsof their own transcript through autoregulation. Therefore, whentransgene expression levels of wild-type or mutant proteins riseabove a threshold, the expression levels of the mouse endogenoustranscripts are reduced, possibly contributing towards toxicity.

    Furthermore, transgenes are often engineered to have exogenouspromotors to ensure high levels of expression in the tissues of interest,but such ectopic expression can result in novel phenotypes. Forexample, two unrelated transgenic mouse lines overexpressing VCPwith the R155H mutation, under the control of a muscle creatinekinase (mMCK) or a cytomegalovirus (CMV) promotor, havedifferences in the survival and presence of cytoplasmic aggregatescontaining VCP, and variability in the levels of motor impairment(Table 1F) (Weihl et al., 2007; Custer et al., 2010). Similarly,transgenic mice overexpressing mutant human TARDBPA315T drivenby the mouse prion promoter (the activity of which is strong inneurons, although it is also widely expressed in other cell types)unexpectedly die early from neurodegeneration in the gut rather thanin MNs (Wegorzewska et al., 2009; Hatzipetros et al., 2014).

    Finally, the transgene arraymay alter copy number at meiosis; thus,colonies need to be monitored constantly because the transgene’scopy number usually determines phenotype severity. For example,the Tg(SOD1*G93Adl)1Gur (SOD1G93Adl; also known as G1del)mice appear to have arisen from a deletion in the transgene array of aSOD1G93A mouse (http://jaxmice.jax.org/strain/002300.html). Theresulting ‘low copy’ SOD1G93A transgenic mouse strain carries∼8-10copies of the human SOD1G93A transgene instead of the ∼25 in theprogenitor line, and these ‘low copy’ animals develop paralysisbetween 24 and 34 weeks of age, considerably later than the ‘highcopy’ progenitor line (Alexander et al., 2004; Acevedo-Arozenaet al., 2011).

    BAC transgenic miceMost – but not all transgenic animals – have been made with thelongest known complementary DNA (cDNA) sequence for the geneof interest; this is usually because of constraints on DNA insert sizein the plasmid vectors used to subclone the transgenic constructs. Toavoid this size limit and to generate mice carrying the full genomicarchitecture of a gene (which is particularly important in the case ofC9ORF72-ALS, for which the mutation is intronic), researchers cangenerate transgenic mice with bacterial artificial chromosome(BAC) vectors, which can carry inserts of up to ∼200 kb. Thisapproach was used to generate, for example, C9ORF72 (Balendraand Isaacs, 2018), TDP-43 (Swarup et al., 2011) and FUS (López-Erauskin et al., 2018) BAC transgenic mice. BACs randomly insertinto the mouse genome, but generally with very low copy numbers(one to three copies), limiting the effects of overexpression of thegene of interest, although even subtle overexpression can alter thephenotype. As with all transgenics, there is the undesired possibilityof insertion mutagenesis, in which integration of the transgene candisrupt an important gene.

    Generic transgenic mouse features for ALS researchUntil recently, transgenics were the fastest technology to obtaingenetically modified mice, but this is changing as CRISPR/Cas9-based technologies develop. As discussed above, phenotypes can berapid and severe in transgenic models because of expression of the

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    http://jaxmice.jax.org/strain/002300.htmlhttp://jaxmice.jax.org/strain/002300.html

  • transgene above endogenous levels. This is helpful for understandingthe advanced stages of disease, which in the natural history of ALS iscomparable to when most patients receive the diagnosis. Severaltransgenic models can have quantifiable, progressive loss of MNssevere enough to lead to profound locomotion defects and paralysisduring the mouse lifespan (Table 1). These features made them themodels of choice for pre-clinical studies and, until recently, almost allALS therapeutics were solely tested on SOD1 transgenic models.This provides some explanation for the past failures of translatingpromising therapeutics from SOD1 transgenics to ALS patients, 98%of whom do not suffer from SOD1-ALS (Urushitani et al., 2007;Turner and Talbot, 2008; Riboldi et al., 2011; Vallarola et al., 2018).

    Mouse models with mutations at physiological levels inendogenous genesGene-targeted and ENU mutant strainsMouse models of ALS can be generated by mutating mouse geneorthologues, to express the relevant protein at physiological levels.Here, we discuss the two key types of model with mutations inendogenous genes, produced from gene-targeting strategies or byrandom mutagenesis with the chemical N-ethyl-N-nitrosourea(ENU). We describe both as ‘physiological’ models in this article,as ‘knock-in’ (KI) is generally used for gene-targeted mice becauseit implies purposely engineering the mouse genome.

    Gene-targeted models of ALSGene targeting entails introducing specific changes to a DNAsequence of interest. In mice, perhaps its most common use has beento create knockout (KO) animals in which the gene no longerfunctions, usually to help us understand the biology of individualgenes. For example, the International Mouse Knockout programaims to functionally KO each mouse gene, providing phenotypicdata for each KO line under the International Mouse Phenotypingprogram (Muñoz-Fuentes et al., 2018).

    Gene KOsAlthough most forms of ALS appear to be caused by toxic dominantGOF, KO models are an important resource as they can reveal notonly critical gene function but also whether there is a loss-of-function (LOF) component to disease pathogenesis. For example,TDP-43 is usually depleted from the nucleus of MNs in TDP-43-ALS, presumably leading to a loss of nuclear TDP-43 function.Although homozygous TDP-43 KO mice are not viable, andheterozygous KO mice express a normal amount of TDP-43 proteindue to its autoregulation, conditional TDP-43 KO lines and atransgenic line expressing small interfering RNA against TDP-43develop MN degeneration (Kraemer et al., 2010), showing thatacute TDP-43 LOF can be a driver of neurodegeneration. In SOD1-ALS, LOF can play a role in disease pathogenesis, as Sod1KOmicedevelop a severe peripheral neuropathy, leading to denervation(Fischer et al., 2011) and SOD1-ALS patients generally havediminished SOD1 dismutase activity (Saccon et al., 2013).

    KI mutationsGene targeting has been used to insert specific mutations, usually(but not always; see Sharma et al., 2016; Gordon et al., 2019) intothe endogenous mouse gene, with the aim of maintainingphysiological expression levels of the (mutant) protein. Thisapproach has been used thus far for Fus, Tardbp, Vapb, Vcp andUbqln2 mutations.Classical gene targeting involves creating recombinant vectors

    for homologous recombination in mouse embryonic stem cells,

    which can be time-consuming and relatively expensive. However,CRISPR/Cas9 targeting in zygotes has made the production ofgene-targeted mice – for example, such as two recently describedstrains recapitulating the


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