Genetic Mutation

Week 1 NURS 2003H Winter 2015Professor Andrea Mowry

Dr. Jane Mackie

Learning Outcomes

• Differentiate between mitosis and meiosis• Describe mutations affecting DNA• Describe mutations affecting protein• Understand the the pathophysiology of

Osteogenesis Imperfecta (OI)• Relate the clinical manifestations of OI (Types 1-IV)

to the pathophysiology• Analyze the appropriateness of diagnostic tests

and treatment for OI

Mutation Rates

• Approximately 23,000 genes in the human genome– Chance of a new harmful mutation in any one person is ~20%

• Only 5000 single-gene disorders recognized to cause human disease

• 1% frequency of disease caused by defects in single gene

• Table 2-2 p.8

• Many conditions are multifactorial– Atheroscelosis– Breast cancer

Clinical Issues

• Most persons with genetic disease present during early childhood with symptoms– Assessment and history leads to a diagnosis & treatment

• i.e. Down syndrome

• Key issues– Correct diagnosis– Counseling for child and parents regarding…

• Natural history• Prognosis• Genetics

Self Review

• Mitosis

• Meiosis

Mitosis & Cytokinesis

https://biolabs.wikispaces.com/chomosomes+and+mitosis

Meiosis

Overview

Genetic information is stored in DNA, transcribed to from RNA and translated (codons read in triplets) to generate protein

DNA RNA Protein

CTC GAG Glu

Types of Genetic Mutations

Mistakes in cell DNA

1. Mutations that affect DNA

2. Mutations that affect protein

All mutations result in a biochemical or molecular biologic phenotype but only some result in a clinically abnormal phenotype.

Two individuals with the same mutated gene may have different phenotypes**

Mutations Affecting DNA

1. Point Mutation– One DNA nucleotide is replaced with another

DNA CTC CTC CTC CTC CTC CACRNA GAG GAG GAG GAG GAG GUGProtein Glu Glu Glu Glu Glu Val

Mutations Affecting DNA

2. Frame Shift Mutation

DNA CTC CTC CTC CTC CCTC CTCRNA GAG GAG GAG GAG GGA GGAGProtein Glu Glu Glu Glu Gly Gly

Mutations Affecting Protein

1. Non-Sense Mutation

DNA ACA ACT

RNA UGU UGA

Protein Cys STOP!

Mutations Affecting Protein

2. Mis-Sense Mutation

DNA ACA ACC

RNA UGU UGG

Protein Cys Trp

More Mis-Sense Mutations• Silent Mutations

– Resulting protein is not affected by the mutation– Many RNA codons can code for the same amino acid

• CCA & CCG & CCT & CCC all code for Gly

• Conservative Mutations– The new amino acid is of the same type as the original– Glu Asp are both acidic amino acids

• Swapping one for the other would be conservative

Non-Conservative Mutations– One amino acid is a different type from the original– Ser Phe

Genetic Diseases

• Most genetic diseases are caused by alterations in DNA sequence

• Every cell in an affected individual carries the mutated gene(s) inherited from the gamete

• A “New Mutation” occurs when the mutation arises during the development of the gamete – Somatic cells of the parent do not carry the mutation

Genetic Diseases

• Mosaic of mutant and non-mutant cells– Mutations may occur during embryogenesis

• Time and cell type dependent– Tissues of the affected individual contain a mixture of cells

• Germline Mosaicism– Some germs cells carry the mutation

• Somatic Mosaicism– Some somatic cells carry the mutation

• Combination of germline and somatic mosaicism

Terms to KnowSelf Review• Gene• Locus• Alleles• Mutation• Polymorphism• Homozygous• Heterozygous• Hemizygosity• Phenotype• Recessive• Dominant

Penetrance• Penetrance– Recognition of the condition in individuals known to carry the

mutated gene– Varies according to…

1. Age– 7 of 10 individuals older than 40 years with Type 1 osteogenesis imperfecta have an

abnormal bone density scan– 70% penetrance

2. Set of criteria– Type 1 osteogenesis imperfecta may be 90% penetrance at age 40 years when bone

scan results are considered in conjunction with lab results for abnormal collagen synthesis

Penetrance

• Reduced or age dependent Penetrance is common in conditions that are dominantly inherited & have a high level of fitness

• Fitness– The extent to which Individuals carrying the

mutant allele produce offspring relative to individuals who do not carry the mutant• i.e. Huntington disease

Expressivity

• The extent to which a mutant genotype affects the phenotype including…– Affected tissues– Severity of effects

• Variable Expressivity– The same mutated gene is penetrant is two individuals but a different spectrum of

phenotypes is expressedi.e. Type 1 Osteogenesis imperfecta

– One sibling has blue sclera and short stature in one persons– Other sibling is confined to a wheelchair

– Differences are related to…• Modifier genes• Environment• Chance

Recessive Inheritance• Recessive Inheritance & Loss of Function Mutations

– Failure of gene to be transcribed or translated– failure of the translated gene product to function correctly

– Expression from non-mutant allele usually does not change• No dosage compensation

– Gene expression in a heterozygous carrier is reduced to 50%

– A 50% reduction in enzyme concentration is not sufficient to produce a disease state

Dominant Inheritance• Dominant Inheritance & Loss of Function Mutations

– 50% of a genetic product is inadequate for the cell or tissue to function normally

– Often occurs in structural proteins• Type 1 osteogenesis imperfecta

– Most dominantly inherited phenotypes are Semi-dominant• Individuals who carry 2 copies of the mutant allele is affected more severely than some who

carries one mutant and one normal copy

– Homozygous mutant individuals are rarely observed

– Huntington’s disease is the only known human condition in which the homozygous and heterozygous mutant phenotype are identical (True Dominant)

Dominant Negative Gene Action

• Occurs frequently in human genetic disorders– Only molecular lesions that produce a protein• Mis-sense, Non-sense or splicing mutation

• Mutant allele causes structurally abnormal proteins to interfere with the function of the normal allelei.e. Type 2 osteogenesis imperfecta

Osteogenesis Imperfecta

• Heterogenous (100 different alleles) and pleiotropic group of disorders characterized by a tendency towards fragility of bone

• 90% of cases are caused by mutation of COL1A1 or COL1A2 genes which encode for subunits of type 1 collagen proα1(I) and proα2(I) respectively

• New group, caused by loss of function mutations in proteins required for proper folding, processing and secretion of collagen

OI Pathophysiology• Disease of Type 1 collagen

– Defect in a major extracellular protein

• Type 1 collagen found in…– Dermis– Connective tissue capsules– Vascular & GI adventitia– ONLY collagen in bone

• Type 1 collagen molecule– Two α1 chains and one α2 chains (FIG 2-2 p.11) are large precursors with amino & carboxyl

terminal propeptide extensions• 3 chains wind around each other in the cell into triple helix• Increased levels of hydroxylation result in a more stable helix• Ultimately secreted as type 1 procollagen molecule• Most all amino acid sequence in the triple helical portion is Gly-X-Y

Type I OI Pathophysiology • Loss of function mutations in COL1A1 is the fundamental defect in

type 1 osteogenesis imperfecta

• Mutant COL1A1allele greatly reduced mRNA (partial loss of function)

• Mutant COL1A1allele no mRNA (complete loss of function)

• Non-mutant COL1A1allele continues to produce mRNA– No dosage compensation

• Heterzygosity for complete loss of function mutation 50% reduction of total proα1 mRNA synthesis

• Hetergygosity for a partial loss of function mutation is a less severe reduction

Type I OI Pathophysiology cont.

• Reduced concentration of proα1 chains limits production of type 1 procollagen leading to…

1. Reduced amount of structurally normal type 1 collagen

2. Excess unassembled proα2 chains

• FIG 2-3 p.12

Type I OI Molecular Defects• COL1A1 mutations are potentially a result of..

– Alterations in the regulatory region leading to reduced transcription

– Splicing abnormalities leading to reduced steady state levels of RNA

– Deletion of the entire COL1A1 gene

– Many cases, a defect in a single single bas pair changed to a premature STOP codon (Nonsense mutation) Truncated protein is more damaging to the cell than production of no protein

For protection… Nonsense mediated decay degrades partially synthesized mRNA precursors

Type II OI Pathophysiology• Caused by structurally abnormal forms of type I collagen• More severe than type I OI• Mutations caused by defects in COL1A1 or COL1A2

– Usually are missense alterations of glycine residue• Allows mutant the peptide chain to bind to normal chains in the initial steps of three chain assembly

Triple helix formation is ineffective Increased post-translational modification by propyl hydroxylase Activation of the unfolded protein stress response

• These are critical events as gylcine substitutions toward the carboxyl end are more severe than the amino terminal end

• Type II OI is more severe than type I OI– Dominant negative gene action

• FIG 2-3 p.12

Type III & IV OI Pathophysiology• Diverse

– Include glycine substitution in the amino terminal portion of the collagen triple helix

– Internal deletions of COL1A1 and CLO1A2– Unusual alterations in non-triple helix extensions at amino and carboxyl

terminal of proαchains

• All types of OI involve a combination of..– reduced production of type 1 collagen in the extra cellular matrix and/or – dysfunctional intracellular collagen processing and maturation

Comparative Molecular Pathophysiology

OI Type Molecular PathophysiologyType IMild

Loss of Function Mutation in proα1(I) chainResulting in decreased amount of mRNAQuality of collagen is normalQuantity of collagen is reduce twofold

Type IIPerinatal Lethal

Structural Mutation in proα1(I) or proα2(I) chainResulting in mild effect on triple helix assemblyQuality of collagen is severely abnormalQuantity of collagen often reduced

Type IIIProgressive Deforming

Structural Mutation in proα1 (I) or proα2(I) chainResulting in mild effect on triple helix assemblyQuality of collagen is severely abnormalQuantity of collagen can be normal

Type IVDeforming with Normal Sclera

Structural Mutation in proα2(I) or less frequently proαI(I) chain that has little or no effect on triple helix assemblyQuality of collagen is usually abnormalQuantity of collagen can be normal

Clinical ManifestationsCliClinical ManifestationsOI Type Initial # X-Ray Stature Hearing Loss Sclera

Type I Childhood 10-20 #sDecrease in pubertyOccasional dental problems

Long bone # r/t minimal or no traumaMild osteopeniaLittle or no deformity

Mild short adult Premature conductive hearing loss

Blue hue

Type II At or before birthMultiple #sDeath from respiratory difficulty

Prenatal multiple #sBony deformityIncreased fragile non-bone connective tissueIsolated islands of mineralizationof skull & ribs

Blue hue

Type III At birth or infancyProgressive bony deformityMultiple 3s

Blue hue

Type IV Childhood>20 #sLong bone & spinal deformityDental abnormality

Same as type IMild to moderate deformity

Significantly short adult

Normal or grey hue

Collaborative Care

• Counseling for child and parents– Natural history of the disease– Treatment– Genetics

• Type III OI Corrective surgery• OT and PT therapy for ambulation

References

Hammer, G.D. & McPhee, S.J. (2010). Pathophysiology of disease; An introduction to clinical medicine (7th ed.). Toronto ON: McGraw Hill Education.

Kahn Academy, Introduction to Genetic Mutationhttps://www.khanacademy.org/test-prep/mcat/biomolecules/genetic-mutations/v/the-different-types-of-mutations