UvA-DARE (Digital Academic Repository) Reproductive ... · ChapterChapter6 Abstractt TheemalespecificregionofthehumanYchromosome(MSY)containsmany amplicons—large,,nearlyidenticalrepeats—thatharbornearlyallofthechromosome's

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Reproductive fitness of the human Y chromosome

Repping, S.

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Citation for published version (APA):Repping, S. (2003). Reproductive fitness of the human Y chromosome. Amsterdam.

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Chapter Chapter

Conservedd large-scale organization of the human Y chromosomee despite its inherent tendency

forr genomic rearrangements

SjoerdSjoerd Repping, Saskia K.M. van Daalen, Cindy M. Korver, Lauraa Brown, Jan W.A. de Vries, Nico J. Leschot,

Fulcoo van der Veen, David C. Page, and Steve Rozen

ToTo be submitted

ChapterChapter 6

Abstrac t t

Thee male-specific region of the human Y chromosome (MSY) contains many

amplicons—large,, nearly identical repeats—that harbor nearly all of the chromosome's

genes.. The large-scale organization of the MSY, i.e, its genomic architecture, shows

ampliconss in direct and inverted orientation and palindromes. Ectopic (non-allelic)

homologouss recombination between amplicons has been shown to alter the genomic

architecturee of the MSY through deletions, duplications and/or inversions, and the rate

off such rearrangements appears to be relatively high. Here, we report the results of a

systematicc survey of variation in MSY genomic architecture in a sample of 47 human Y

chromosomess representing worldwide diversity. We found no rearrangements in the

proximall long arm, indicating that these are rare or non-existent. In contrast, in the

distall long arm encompassing the AZFc region, we found 18 large-scale

rearrangements.. Further analysis indicates that a single homologous recombination

eventt or two, or three successive events have caused nearly all these rearrangements.

Despitee these rearrangements, the genomic architecture of AZFc and, even more so, the

copyy number of AZFc gene families is conserved in the majority of modern day Y

chromosomes.. Such relative uniformity is unexpected if one assumes that

rearrangementss via homologous recombination are selectively neutral, and suggests

thatt selection has constrained the genomic architecture of the MSY.

100 0

VariationVariation in genomic architecture of the MSY

Thee segment of the human Y chromosome that does not undergo meiotic

recombinationn with the X chromosome is referred to as the male-specific region of the Y

chromosome,, or MSY 1. The MSY comprises 95% of the entire chromosome and harbors

fourr discrete sequence classes. One of these is the ampliconic sequence class which is

constructedd of large nearly identical repeats, or amplicons. In total, the ampliconic

sequencess represent 45% (or ~10 Mb) of the euchromatic MSY sequence. The large-

scalee organization of the MSY, i.e. its genomic architecture, shows amplicons in direct

andd inverted orientation, including palindromes—inverted amplicons almost without

interveningg sequence (Fig. 1).

Ampliconss on the MSY have previously been shown to be mediators in generating

deletions,, duplications and inversions that change the MSY genomic architecture 2"9.

Homologouss recombination between inverted repeats causes inversion of the

interveningg sequence, while recombination between direct repeats leads to either a

duplicationn or a deletion. In fact, the vast majority of deletions on the MSY result from

suchh ectopic (non-allelic) homologous recombination between amplicons 5_7'9'10. The rate

att which these deletions occur seems to be relatively high: over 1/4000 male births 5.

Thuss far, seven distinct deletions on the MSY have been shown to arise through

homologouss recombination: azoospermia factor a {AZFa), P5/proximal-Pl, P5/distal-Pl,

AZFcAZFc (b2/b4), bl /b3, gr/gr, and b2/b3 deletions (Fig. 1) 5-7<9<10. Molecular

characterizationn of these deletions has shown precisely which amplicons are responsible

forr each rearrangement. In fact, the deletion nomenclature of the majority of these

deletionss is based on the amplicons involved in the homologous recombination event

thatt causes the deletion. For instance, the bl/b3 deletion is caused by homologous

recombinationn between amplicons b l and b3 (Fig. 1).

Thesee deletions have been studied in great detail, partly because they can easily

bee ascertained via plus/minus STS deletion screening. Detection of other large-scale

genomicc rearrangements on the MSY requires more laborious methodologies such as

FISHH and consequently has not been addressed thoroughly. Duplications can arise on

thee MSY via homologous recombination between the same amplicons that are involved

inn deletions 3,7(9. Inversions in the AZFc region due to homologous recombination have

alsoo been described as intermediate steps in the generation of b2/b3 deletions 7.

101 1

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Wee sought to determine the spectrum of variation in genomic architecture on the

MSYY by examining a worldwide diverse sample of human Y chromosomes. To make our

searchh systematic, we tested representative samples from 47 major branches of the Y-

chromosomee genealogical tree in our analysis (Fig. 2) n"13. We used one-color

Fluorescencee in-situ hybridization (FISH) that would detect duplications and deletions of

thee AZFa region and the region between P5 and PI (Fig. 1). Similarly, we used one- or

two-colorr FISH that would detect deletions, duplications and inversions in AZFc caused

byy homologous recombination between amplicons (Fig. 1).

Wee found no deletions or duplications of the AZFa and P5/P1 region (Table 1).

Thus,, these rearrangements must be relatively uncommon or non-existent (0/47, 95%

CII 0.0 to 0.01). In contrast, we detected genomic rearrangements in the AZFc region in

188 out of the 47 samples tested. Four chromosomes showed deletions, nine showed

inversions,, and five showed duplications (Table 1 and Fig. 2).

Inn our initial screen, we included only one sample per branch of the tree. To

determinee whether the detected rearrangements are representative for other samples

fromm these branches, we tested additional samples when available. Out of 12 branches

tested,, five contained exclusively chromosomes with a specific AZFc rearrangement

(Fig.. 2). Thus, in these five branches, the rearrangement appears to be common or

universal.. In the remaining seven branches, we found additional samples with

architecturess distinct from those identified in the primary samples (Table 1 and Fig. 2).

Wee then performed an in-silico analysis to systematically enumerate all

rearrangementss that could be produced from the AZFc reference sequence by

homologouss recombination between amplicons. We found that a single homologous

recombinationn event can generate nine different architectures besides the reference

sequencee (Fig. 3A). Similarly, our analysis shows that sequences of two and three

homologouss recombination events can produce 69 (Suppl. Fig. 1) and 969 distinct

genomicc architectures, respectively.

Analysiss of our new data together with previously published data on AZFc

rearrangements,, showed that six of the nine predicted rearrangements that can be

generatedd by a single homologous recombination event exist (Table 1 and Fig. 3A) 5'7,9.

Inn addition, we detected five rearrangements that could be produced by two successive

homologouss recombination events and one instance of a three-step rearrangement

(Tablee 1, Fig. 3B,C and Suppl. Fig. 2). Two duplications detected in the survey could not

bee explained by homologous recombination between amplicons (Table 1). These may

bee due to homologous recombination between short repeats or to non-homologous

recombination;; such rearrangements are much less common on the Y chromosome than

homologouss recombination between amplicons, but have been reported 5'6.

103 3

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Tablee 1. Results from screening for genomic rearrangements on the MSY.

## of signals for FISH-probes c

-- ^ *~; Organization '' XÏ Organization of of green (G)

jyy iS" r* N] r N green (G) and and red (R) '' yellow (Y) dots dots

I-a a I-b b I-c c I-d d I-d d I-d d I l -a a I l -b b I l -b b I l l - a a I l l - b b I I I -c c I I I -d d I l l - e e I l l - f f I l l - g g I l l - h h IV-a a IV-a a IV-a a IV-a a IV-a a IV-a a IX-a a IX-b b IX-c c IX-d d V-a a V-a a Vl -a a Vl -a a V l -b b VI-c c VI-c c VI-c c V l -d d v i - e e V l - f f V l - f f V l - f f V l - f f V l - f f V l - f f V l - f f

W H T 4 1 6 3 3 G M 0 6 3 4 2 2 YCC038 8 N A 0 3 0 4 3 3 YCC022 2 YCC034 4 P D 0 6 1 1 N A 1 0 4 7 0 0 WHT3882 2 PD339 9 YCC037 7 N A 0 2 0 9 0 0 PD399 9 PD123 3 W H T 3 0 2 7 7 P D 1 1 1 1 W H T 3 1 5 9 9 PD178 8 NA04535 5 WHT3341 1

PD0436 6 NA00743 3 NA10810 0 W H T 3 8 7 4 4 PD217 7 W H T 2 6 3 0 0 W H T 3 2 4 2 2 W H T 3 5 5 2 2 WHT3474 4 W H T 2 6 1 1 1 WHT2709 9 W H T 3 4 4 9 9 W H T 2 4 2 6 6 WHT3560 0 WHT2783 3 PD437 7 W H T 3 2 5 5 5 PD388 8 PD335 5 WHT3167 7 WHT3298 8 WHT3431 1 WHT3442 2 WHT3689 9

1 1 1 1 1 1 1 1 1 1

1 1

1 1 1 1

1 1 1 1 1 1 1 1

2 2 LL 3

2 2 LL 3

3 3 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2

2 2 2 2 2 2 2 2 2 2 2 2 2 2 1 1 2 2

LL 3 LL 6

1 1 2 2 2 2

LL 4

2 2 2 2 2 2 2 2 2 2 2 2

3 3 5 5 4 4 4 4 4 4 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 2 2 2 2 2 2 2 2 2 2 2 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 2 2 3 3 5 5

9-12 2 2 2 3 3 3 3 6 6 3 3 3 3 3 3 3 3 3 3 3 3

2 2 3 3 2 2 3 3 3 3 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2

2 2 2 2 2 2 2 2 2 2 2 2 2 2 1 1 2 2 3 3

6-8 8 1 1 2 2 2 2 4 4 2 2 2 2 2 2 2 2 2 2 2 2

G-Y-G-G-Y Y G-G-Y-G-G-Y-G-Y Y

G-Y-G-G-Y-G G G-G-Y-G-Y-G-Y Y G-G-Y-G-Y-G-Y Y

G-Y-G-G-Y Y G-G-Y-G-Y Y G-G-Y-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-G-Y-G-Y Y

G-G-Y Y G-G-Y Y G-G-Y Y G-G-Y Y G-G-Y Y G-G-Y Y

G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-G-Y-G-Y Y G-G-Y-G-Y Y G-G-Y-G-Y Y

G-Y-G-G-Y Y j j

.. f

G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y

__ r

G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y

G-R-G-R-G G G-R-G-G-R-G-R-G G

G-R-G-R-G-G G G-R-G-R-G-R-G G G-R-G-R-G-R-G G

G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G

G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G

G-R-G G G-R-G G G-R-G G G-R-G G G-R-G G G-R-G G

G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G

G-R-G-R-G G G-R-G G

G-R-G-R-G G G-R-G-G-R-G-R-G G

__ f

G-R-G G G-R-G-R-G G G-R-G-R-G G

__ f

G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G

G-R-G-R-G G

# 3 6 6 ? ?

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105 5

ChapterChapter 6

Tablee 1 (cont'd from previous page). ## of signals for FISH-probes c

to to

Q. .

O O O. . ra ra X X VI-g g VI-h h VI- i i V I - j j VI I -a a vil-b b V l l - b b VI I -c c V l l -d d VH-e e VH-f f VH-g g VH-h h

VI I I -a a VI I I -a a V I I I -a a

I D b b

W H T 3 6 3 5 5 P D 2 7 6 6 PD073 3 P D 1 4 6 6 P D 0 1 6 6 P D 2 6 4 4 PD098 8 P D 1 3 1 1 P D 1 7 0 0 P D 1 8 9 9 P D 2 7 4 4 PD143 3 PD197 7 P D 3 2 1 1 PD403 3 WHT3645 5

V I I I - bb PD427 VI I I -bb WHT716 VI I I -bb WHT3420 VI I I -bb WHT4704 VI I I -bb WHT4830 VI I I -c c VHI-d d V I I I -e e V I I I -e e VI I I -e e VI I I -e e V I I I - f f X-a a X-b b X-c c X-c c X-c c X-c c

Total l

W H T 3 2 5 7 7 W H T 3 2 9 9 9 PD116 6 PD378 8 WHT3427 7

WHT3730 0 N A 1 0 5 4 1 A A P D 4 2 1 1 PD306 6 PD024 4 PD066 6 PD296 6

f»» rvj

CÉÉ O CU it-

1 1

1 1 1 1 1 1

1 1 1 1 1 1 1 1 1 1 1 1

GM112000 1

Initiall samples 4 7

( total)) 78

s s c c (0 0

000 Sr

000 Q.

2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 1 1 2 2 3 3

2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 1 1 1 1 2 2 2 2

LL. .

3 3 3 3 3 3 3 3 3 3 3 3 2 2 3 3 3 3 3 3 3 3 3 3 3 3 1 1 2 2 2 2

3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 1 1 1 1 4 4 3 3

o o

5 5 . 11 iV

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2 2 2 2 2 2 2 2 2 2 2 2 3 3 2 2 2 2 2 2 2 2 2 2 2 2 1 1 2 2 2 2 1 1 1 1 1 1 1 1 1 1 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 1 1 1 1 2 2 2 2

Organizationn of greenn (G) and yelloww (Y) dots

G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y

G-Y-G-Y Y G-Y-G-Y Y

G-Y Y G-Y Y G-Y Y

G-Y-G-G-Y Y G-Y-G-G-Y Y G-G-Y-G-Y Y G-G-Y-G-Y Y G-G-Y-G-Y Y G-G-Y-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y G-Y-G-G-Y Y

G-Y Y G-Y Y

G-Y-G-G-Y Y

OrganizationOrganization of greenn (G) and redd (R) dots G-R-G-R-G G G-R-G-R-G G R-G-G-R-G G G-R-G-R-G G R-G-G-R-G G R-G-G-R-G G G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G G-R-G-R-G G

G-R R G-R-G-R R

R-G-R-R-G G G-R R G-R R G-R R G-R R G-R R



G-R R G-R R

G-R-G-G-R-G G G-R-G-R-G G

c c <L> > o> > C. C.

10 0 QJ J

it it N N

# 7 7

# 7 7 # 7 7

#35 5 #449 9

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#35 5 #35 5 #35 5 #35 5 #35 5

#10 0 #10 0 #10 0 #10 0

#35 5 #35 5 #56 6

Rearrangementss 18

( total)) 39

33 Samples indicated in bold were part of the initial screen. Other samples were tested as additional sampless in branches with rearrangements.

bb See figure 2. cc See figure 1. dd Samples were only tested with RP11-1325K3 if probe 18E8 showed more than two signals since

aa duplication of the P5/P1 region would produce either three or four 18E8 signals (see Fig. 1). ee Assuming homologous recombination with a minimum number of steps as the mechanism

underlyingg the rearrangements (Fig. 3 and Suppl. Fig. 1). Question marks indicate that the rearrangementt cannot be explained by homologous recombination between amplicons.

ff For these samples it was impossible to determine the organization of amplicons because too manyy signals were present.

106 6


Althoughh deletions, inversions and duplications seem to occur fairly frequent in

thee AZFc region, the majority of Y chromosomes (29/47) has the same genomic

architecturee as the reference sequence, excluding the possibility of unassayed bl/b4

inversionss (see Methods). Even more striking is the conserved gene content: 38 out of

thee 47 chromosomes examined (~81%) have the same copy number of AZFc gene

familiess as the reference sequence (Table 2). Preliminary results from computer

simulationss suggest that all Y chromosomes would be deleted for (part of) AZFc if there

weree no selection against these architectures. This is consistent with previous reports

thatt have established that these deletions are selected against because they negatively

affectt spermatogenesis and hence fitness 5-9. If we incorporate selection against these

deletionss in our simulation, we observe a highly variable frequency of duplications from

simulationn to simulation.

Tablee 2. Gene content of chromosomes with rearranged AZFc regions.

Inversions s Rearrangements s

Deletions s Duplications s

Genee or transcript t familyy a

RBMY Y BPY2 BPY2 DAZ DAZ CDY1 CDY1 PRY PRY CSPG4LY CSPG4LY GOLGA2LY GOLGA2LY TTTY3 TTTY3 TTTY4 TTTY4 TTTY5 TTTY5 TTTY6 TTTY6 TTTY17 TTTY17 Total l

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ChapterChapter 6

i i >> M I I I I (referencee sequence)

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Figuree 3 (this and opposite page) . AZFc rearrangements. A, Rearrangements that can be producedd by a single homologous recombination event. Genomic architectures are shown as a sequencee of colored arrows representing the different amplicons (color coding is identical to that inn f igure 1). The nomenclature of each rearrangement is based on the amplicons involved in the homologouss recombination event. For example, homologous recombination between the inverted ampliconss b2 and b3 leads to rearrangement #7 and is referred to as a b2/b3 inversion. Next to thee genomic architectures are shown FISH images of nuclei hybridized with amplicon-specific probess shown in figure 1. The color of each FISH signal corresponds to the color of the amplicon itt detects. B, Genomic architectures detected in our survey that result f rom a sequence of two homologouss recombination events. C, Rearrangement #449 that can only be explained as the resultt of a sequence of at least three homologous recombination events.

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Severall factors could explain the relatively low frequency (~10%) of duplications

inn our survey. One possible explanation could be that duplications simply occur less

frequentt than deletions. This would not be surprising given the different mechanisms

thatt could cause deletions and duplications on the Y chromosome. Deletions on the Y

chromosomee can arise through both homologous recombination within a chromatid and

homologouss recombination between two sister chromatids of a single Y chromosome 14.

Inn contrast, duplications can only arise through homologous recombination between two

sisterr chromatids, and thus may be generated less frequently than deletions.

Alternatively,, our observation of conserved gene content in the AZFc region could

indicatee selection against duplications in a similar fashion as selection against deletions.

Indeed,, similar phenotypic consequences for deletions and duplications have been

describedd for other regions in the genome. For example, hereditary neuropathy with

liabilityy to pressure palsies (HNPP) is caused by a deletion on chromosome 17pl2 while

duplicationn of the same region causes a different neuropathy, Charcot-Marie-Tooth

syndromee type IA (CMT1A) 15,16. Thus, duplications in the AZFc region, like deletions,

couldd have a negative impact on spermatogenesis and fitness which leads to their

inabilityy to persist in the population.

Ourr data present one of the first glimpses of the spectrum of variation in large-

scalee genomic organization in the human population. Cataloging genetic variation and

determiningg its role in differences in disease susceptibility is one of the grand challenges

off the Human Genome Project 17. With regard to variation in genomic architecture on

thee MSY, certain architectures in addition to deletions might confer risk for

spermatogenicc failure. The haplotype-based investigation of variation together with the

FISHH methodology described here should provide the foundation for future research

aimedd at determining the phenotypic effects of other genomic rearrangements on the

MSY. .

110 0


Methods s

Samples Samples

Thee majority of samples used was purchased from the NHGRI/NIGMS DNA

Polymorphismm Discovery Resource (Coriell Cell Repositories) 18. In some cases, we

identifiedd additional samples from our own collection of Y chromosomes to gain as much

diversityy as possible and to have at least one representative of each branch of the

genealogicall tree.

HaplotypeHaplotype analysis

Individualss were haplotyped using the Y-linked polymorphisms listed in

Supplementaryy Table 1.

FluorescenceFluorescence in-situ Hybridization

One-- or two-color FISH was performed as described previously 9. We used

interphasee nuclei as targets and BAC RP11-217J19 (AZFa) 19f BAC RPU-1325K03

(P5/P1)) 19 and cosmid 18E8 (AZFc) 20 as probes to search for duplications or deletions

(seee Fig. 1). For each sample, at least 200 nuclei were counted. To determine the

genomicc architecture in the AZFc region we used BAC RP11-336F2 (green amplicon) and

BACC RP11-79J10 (yellow amplicon) 19. Our FISH analysis did not allow the detection of

b l / b 44 inversions since this would require three-color FISH.

EnumerationEnumeration of AZFc rearrangements

Wee wrote a computer program to enumerate AZFc rearrangements that could be

producedd by homologous recombination events between amplicons. We used it to

exhaustivelyy enumerated all rearrangements of the AZFc region that could be generated

byy one, two or three successive homologous recombination events.

SimulatedSimulated evolution of AZFc rearrangements

Wee investigated the possible evolutionary behavior of rearrangements of the AZFc

regionn using a computer simulation. This was a forward Monte-Carlo simulation of the

evolutionn of a fixed-size population of Y chromosomes incorporating Wright-Fisher

randomm genetic drift. Fitness of Y chromosomes could be parameterized as a function

off the number of "DAZ clusters" (closely spaced, 3 ' ^5 ' : :5 ' -»3 ' pairs of DAZ genes in the

" r ed "" amplicons, detected by FISH probe 18E8). The frequency of deletion

rearrangementss was a function of the size of the targets of homologous recombination.

Thiss frequency was estimated from the rate of b2/b4 AZFc deletions: 1/4000 per

generationn per 229 kb 5. Frequencies of duplications and inversions could be specified

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ChapterChapter 6

ass fractions of the rate of deletions. This allowed us to explore models in which

duplicationss and inversions are less frequent than deletions caused by recombination

betweenn amplicons of the same size. Effective population size was estimated at 1000,

whichh gives an expected time to most recent common ancestor (TMRCA) of 2000

generations;; at a 30-year male generation, this is 60,000 years, in approximate

agreementt with recent estimates of the actual TMRCA of human Y chromosomes.

References s

1.. Skaletsky, H., Kuroda-Kawaguchi, T., Minx, P.J., Cordum, H.S., Hillier, L, Brown, L.G., Repping,, S., et al. The male-specific region of the human Y chromosome is a mosaic of discretee sequence classes. Nature 423, 825-837 (2003).

2.. Blanco, P., Shlumukova, M., Sargent, CA., Jobling, M.A., Affara, N. & Hurles, M.E. Divergentt outcomes of intrachromosomal recombination on the human Y chromosome: malee infertility and recurrent polymorphism. J.Med.Genet. 37, 752-758 (2000).

3.. Bosch, E. & Jobling, M.A. Duplications of the AZFa region of the human Y chromosome are mediatedd by homologous recombination between HERVs and are compatible with male fertility.. Hum.Mol.Genet 12, 341-347 (2003).

4.. Kamp, C, Hirschmann, P., Voss, H., Huellen, K. & Vogt, P.H. Two long homologous retrovirall sequence blocks in proximal Y q l l cause AZFa microdeletions as a result of intrachromosomall recombination events. Hum.Mol.Genet. 9, 2563-2572 (2000).

5.. Kuroda-Kawaguchi, T., Skaletsky, H., Brown, L.G., Minx, P.J., Cordum, H.S., Waterston, R.H.,, Wilson, R.K., et al. The AZFc region of the Y chromosome features massive palindromess and uniform recurrent deletions in infertile men. Nat.Genet. 29, 279-286 (2001). .

6.. Repping, S., Skaletsky, H., Lange, J., Silber, S., van der Veen, F., Oates, R.D., Page, D.C., ett al. Recombination between palindromes P5 and PI on the human Y chromosome causes massivee deletions and spermatogenic failure. Am.J.Hum.Genet. 7 1 , 906-922 (2002).

7.. Repping, S., van Daalen, S.K., Korver, CM., Brown, L.G., Marszalek, J.D., Gianotten, J., Oates,, R.D., et al. A family of human Y chromosomes has persisted for millennia despite a deletionn of half of the Azoospermia Factor c region. Submitted (2003).

8.. Sun, C, Skaletsky, H., Rozen, S., Gromoll, J., Nieschlag, E., Oates, R. & Page, D.C. Deletionn of azoospermia factor a (AZFa) region of human Y chromosome caused by recombinationn between HERV15 proviruses. Hum.Mol.Genet.2000. 9, 2291-2296 (2000).

9.. Repping, S., Skaletsky, H., Brown, L.G., van Daalen, S.K., Korver, CM., Pyntikova, T., Kuroda-Kawaguchi,, T., et al. Polymorphism for a 1.6-Mb deletion of the human Y chromosomee persists through balance between mutation and haploid selection. Submitted (2003). .

10.. Vogt, P.H., Edelmann, A., Kirsch, S., Henegariu, O., Hirschmann, P., Kiesewetter, Kohn, F.M.,, et al. Human Y chromosome azoospermia factors (AZF) mapped to different subregionss in Y q l l , Hum.Mol.Genet. 5, 933-943 (1996),

11.. Underhill, P.A., Shen, P., Lin, A.A., Jin, L., Passarino, G., Yang, W.H., Kauffman, E., et al. YY chromosome sequence variation and the history of human populations. Nat.Genet. 26, 358-3611 (2000).

12.. Underhill, P.A., Passarino, G., Lin, A.A., Shen, P., Mirazon, L.M., Foley, R.A., Oefner, P.J., et al.. The phylogeography of Y chromosome binary haplotypes and the origins of modern humann populations. Ann.Hum.Genet. 65, 43-62 (2001).

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13.. The Y chromosome Consortium. A nomenclature system for the tree of human Y-chromosomall binary haplogroups. Genome Res. 12, 339-348 (2001).

14.. Stankiewicz, P. & Lupski, J.R. Genome architecture, rearrangements and genomic disorders.. Trends.Genet 18, 74-82 (2002).

15.. Potocki, L , Chen, K.S., Park, S.S., Osterholm, D.E., Withers, M.A., Kimonis, V., Summers, A.M.,, et al. Molecular mechanism for duplication 17pl l .2- the homologous recombination reciprocall of the Smith-Magenis microdeletion. Nat.Genet. 24, 84-87 (2000).

16.. Inoue, K., Dewar, K., Katsanis, N., Reiter, L.T., Lander, E.S., Devon, K.L., Wyman, D.W., et al.. The 1.4-Mb CMT1A duplication/HNPP deletion genomic region reveals unique genome architecturall features and provides insights into the recent evolution of new genes. Genome Res.Res. 1 1 , 1018-1033

17.. Collins, F.S., Green, E.D., Guttmacher, A.E. & Guyer, M.S. A vision for the future of genomicss research. Nature 422, 835-847 (2003).

18.. Collins, F.S., Brooks, L.D. & Chakravarti, A. A DNA polymorphism discovery resource for researchh on human genetic variation. Genome Res. 8, 1229-1231 (1998).

19.. Tilford, C.A., Kuroda-Kawaguchi, T., Skaletsky, H., Rozen, S,, Brown, L.G., Rosenberg, M., McPherson,, J.D., et al. A physical map of the human Y chromosome. Nature 409, 943-9455 (2001).

20.. Saxena, R.f de Vries, J.W.A., Repping, S., Alagappan, R.K., Skaletsky, H., Brown, LG, et al. Fourr DAZ genes in two clusters found in the AZFc region on the human Y chromosome. GenomicsGenomics 67, 256-267 (2000).

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Supplementaryy information

Supplementaryy Table 1. References for polymorphisms defining the Y-chromosome haplotypes andd genealogy in figure 2.

Polymorphismm Reference DYS257 7 DYS271(M2) ) M109 9 M112 2 M116 6 M117 7 M118 8 M119 9 M12 2 M122 2 M123 3 M124 4 M13 3 M134 4 M14 4 M144 4 M168 8 M167 7 M170 0 M172 2 M173 3 M175 5 M20 0 M3 3 M32 2 M35 5 M4 4 M50 0

Polymorphism m M51 1 M52 2 M58 8 M60 0 M64 4 M67 7 M69 9 M75 5 M76 6 M78 8 M81 1 M82 2 M89 9 M9 9 M91 1 M92 2 M95 5 M96 6 pl2f f pre-Tat t RPS4Y711 1 Tat t SRY+465 5 SRY10831 1 USP9Y+3178 8 USP9Y+3636 6 YAP P

Reference e

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Supplementaryy Figure 1 ( this and t w o previous pages) . Genomic architectures that can resultt f rom two or fewer successive homologous recombination events in AZFc. The distinct genomicc architectures that can arise through homologous recombination are numbered 1 through 69.. The color coding of amplicons is identical to that in figure 1. Below the different architectures aree shown the name of the rearrangement, if available, and the intermediate step (parent) of the rearrangement.. For instance, rearrangement #12 can arise on either a b l / b 3 duplicated chromosomee (#3 ) or a b2/b4 duplicated chromosome ( # 6 ) .

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ChapterChapter 6

Supplementaryy Figure 2 (opposite page) . Generation of rearrangement #38 through two independentt pathways. A, The ampliconic complex embedding AZFc, shown to scale 5. The centrall bar depicts the organization of the constituent amplicons, which are color-coded; sequencess with the same color are > 9 9 . 9 % identical. The green and red "bows" indicate the regionss involved in the first homologous recombination event in both pathways. B, FISH probes hybridizedd to interphase nuclei f rom a man with the reference sequence in A. C, Pathway one: gr / rgg inversion ( g l , r l , r2 recombining with g2, r3 , r4 ; green-shaded box) followed by a red-gray/red-grayy duplication (green-shaded box), both via homologous recombination. The green "bow"" in the inverted organization indicates the sequences involved in the deletion. D, Pathway two:: g r /g r duplication (red-shaded box) followed by gr / rg inversion (red-shaded box) both via homologouss recombination. The red "bow" in the inverted organization indicates the sequences involvedd in the deletion. E, Final arrangement in both pathways. F, Interphase nuclei of a man (YCC022)) with rearrangement #38 hybridized with FISH probes as in 0. The patterns are as predicted:: green-red-green-red-green-red-green (left) and green-green-yel low-green-yel low-green-yelloww (r ight) .

Supplementaryy References

1.. Shinka, T. et al . Genetic variations on the Y chromosome in the Japanese population and implicationss for modern human Y chromosome lineage. J.Hum.Genet. 4 4 , 240-45 (1999)

2.. Repping, S. et al . A family of human Y chromosomes has persisted for millennia despite a deletionn of half of the Azoospermia Factor c region. Submitted (2003)

3.. Hammer, M.F. et al . Out of Africa and back again: nested cladistic analysis of human Y chromosomee variat ion. Mol. Biol. Evol. 15 , 427-41 (1998).

4.. Seielstad, M.T. et al . Construction of human Y-chromosomal haplotypes using a new polymorphicc A to G transit ion. Hum. Mol. Genet 3, 2159 -2161 (1994),

5.. Underhil l, P.A. et al . Y chromosome sequence variation and the history of human populations.. Nat Genet 2 6 , 358-361 (2000).

6.. Underhil l, P.A. et al . Detection of numerous Y chromosome biallelic polymorphisms by denaturingg high-performance liquid chromatography. Genome Res. 7 , 996-1005 (1997).

7.. Underhil l, P.A., Jin, L , Zemans, R., Oefner, P.J. & Cavalli-Sforza, L.L. A pre-Columbian Y chromosome-specificc transit ion and its implications for human evolutionary history. Proc. Natl.Natl. Acad. Sci. U. S. A. 9 3 , 196-200 (1996).

8.. Shen, P. et al. Population genetic implications f rom sequence variation in four Y chromosomee genes. Proc. Natl. Acad. Sci. U. S. A. 9 7 , 7354-7359 (2000).

9.. Casanova, M. et al . A human Y-linked DNA polymorphism and its potential for estimating geneticc and evolutionary distance. Science 2 3 0 , 1403-1406 (1985) .

10.. Bergen, A.W. et al. An Asian-Native American paternal lineage identified by RPS4Y resequencingg and by microsatell ite typing. Ann. Hum. Genet. 6 3 , 63-80 (1999).

1 1 .. Zerjal, T. et al. Genetic relationships of Asians and Northern Europeans, revealed by Y-chromosomall DNA analysis. Am. J. Hum. Genet. 6 0 , 1174-1183 (1997).

12.. Whitf ield, L.S., Lovell-Badge, R. & Goodfellow, P.N. Rapid sequence evolution of the mammaliann sex-determining gene SPY. Nature 3 6 4 , 713-715 (1993) .

13.. Sun, C, et al . An azoospermic man with a de novo point mutat ion in the Y-chromosomal genee USP9Y. Nat. Genet. 23 , 429-432 (1999).

14.. Hammer, M.F. A recent insertion of an Alu element on the Y chromosome is a useful markerr for human population studies. Mol. Biol. Evol. 1 1 , 749-761 (1994).

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UvA-DARE (Digital Academic Repository) Reproductive ... · ChapterChapter6 Abstractt TheemalespecificregionofthehumanYchromosome(MSY)containsmany amplicons—large,,nearlyidenticalrepeats—thatharbornearlyallofthechromosome's