L9, Repeat Expansion Disorders II Flashcards

1
Q

DM Characteristics: (onset, inheritance, clinical features, types)

A
  • Myotonic dystrophy -> Often adult onset; age of onset largely determined by number of repeats
  • Second most common type of muscular dystrophy
  • Dominant inheritance pattern
  • Affects 1 in 8500 individuals worldwide
  • Multisystemic disorder with varied clinical features (myotonia, muscle wasting, insulin resistance, cardiac conduction defects, cataracts, with cognitive dysfunction, ID in more sever form)
  • Two forms: DM1, DM2
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2
Q

Broad pathophysiology of DM1 and DM2:

A
  • Different genes (non-coding regions)
  • Disease caused by expansion resulting in gain of RNA function
  • DM1: CTG repeats in 3’ UTR of DMPK gene
  • DM2: CCTG repeats in intron 1 of ZNF9/CNBP gene
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3
Q

Thresholds for repeats in DM1, additional features:

A
  • DM1: CTG repeats in 3’ UTR of DMPK gene
  • Normal: 5-37
  • Premutation: 37-50
  • Pathogenic: 50 -1000
  • Congenital form: >1000
  • Maternal expansion bias
  • Extensive somatic instability in proliferative tissue particularly muscle
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4
Q

Thresholds for repeats in DM2:

A
  • DM2: CCTG repeats in intron 1 of ZNF9/CNBP gene
  • Normal: <30
  • Premutation: 31-74
  • Pathogenic: 75-11000
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5
Q

How does the RNA GOF mechanism work in DM?

A
  • Transcribed RNA forms unusual structures -> imperfect hairpins in repeat containing sections
  • DM1: CUG
  • DM2: CCUG
  • In both cases, GC pairing occurs -> partial base-pairing along hairpin
  • These imperfect hairpins bind and sequester certain proteins
  • e.g. DM1 hairpin sequesters MBNL1 -> loss of function (alternative splicing as a result), additionally results in PKC activation -> CUGBP1 stabilised and level thus increased -> alternative splicing, mRNA translation, mRNA decay
  • See FC
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6
Q

Give four examples of aberrant splicing as a result of DM:

A
  • Insulin receptor -> insulin resistance
  • Chloride channel -> Myotonia (Link to B302!)
  • Cardiac troponin T -> Cardiac abnormalities
  • Unidentified genes -> cataracts, testicular failure
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7
Q

Discuss the different diseases arising from abnormal amounts of repeats in FMR1:

A
  • Fragile X syndrome (200-4000 repeats)
  • Fragile X-associated tremor/ataxia syndrome/FXTAS (40% males, 8% females with premutation repeats of 55-200) -> Late onset neuropsychiatric degenerative disorder
  • Fragile X associated premature ovarian insufficiency/FXPOI (25% of females carrying premutation repeat 55-200)
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8
Q

Describe the disease mechanisms in premutation repeat length potentially leading to FXTAS ad FXPOI:

A
  • 5-55 repeats -> transcription, translation -> FMRP protein
  • 55-200 repeats -> increased transcription, up to 8x normal RNA level (repeat stretch thought to sequester RNA binding proteins, resulting in dysregulation of protein expression)
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9
Q

RAN: Effect

A
  • Repeat-associated Non-ATG translation of repeat containing RNA
  • Leads to production of toxic peptides
  • Repeat expansion can be transcribed in two directions leading to sense and antisense transcripts -> protein toxicity in both cases (not fully understood; lack of ATG initiation codon)
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10
Q

Amyotrophic lateral sclerosis: How does RAN translation cause this?

A
  • Expanded GGGGCC repeats in first intron of C9orf72 cause autosomal dominant ALS
  • 3 different contributory mechanisms…
  • Loss of function -> haploinsufficiency
  • RNA toxicity
  • Toxic RAN peptides
  • There is selective vulnerability of motor neurons in this condition, possibly as a result of differential expression of VEGF (higher expression may have a protective role)
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11
Q

Describe the mechanism for repeat instability:

A
  • Stable inheritance below threshold
  • Above repeat threshold, intergenerational instability -> dynamic mutation (both expansion and contraction) -> somatic instability in both dividing and non-dividing cells
  • Can depend on maternal vs paternal line
  • Scale of expansion can vary hugely; from a few repeats to many hundreds of them
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12
Q

Types of unusual non B-DNA structures formed by expanded repeats:

A
  • H-DNA (triplex)
  • G-quadruplex
  • Hairpin (perfect or imperfect)
  • DNA unwinding elements (must be AT rich region)
  • Inverted repeats can form perfect cruciforms
  • Direct repeats can result in imperfect cruciform structures (both stable and slipped strand from reannealing)
  • These structures in DNA are hard to replicate and hard to transcribe
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13
Q

How do stabilising interruptions affect RED alleles?

A
  • Longer repetitive runs with stabilising interruptions (‘long-normal’ alleles) e.g. AGG insertions can sill have normal phenotype
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14
Q

How may DNA replication lead to expanded/contracted repeats?

A
  • Replication of dsDNA is asymmetric -> leading and lagging strands
  • Structure prone sequence on lagging strand promotes instability
  • Skipping hairpin on one -> contraction whereas stalling, reversal, restart -> expansion
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15
Q

What processes are involved in transition from normal to unstable repeat sequence:

A
  • Change of replication context of structure prone strand
  • Two origins; first one would have unstable side on leading strand
  • If the first origin becomes inactivated, the unstable region ends up on lagging strand -> much more likely to form hairpin structures -> instability
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16
Q

How does mismatch repair promote repeat instability? (Toxic oxidation cycle)

A
  • Proposed mechanisms of instability have a role for MutSB in stabilising DNA structures
  • Oxidative damage is usually repaired by BER (glycosylase removes base, AP endonuclease nicks DNA strand)
  • DNA pol sigma initiates repair synthesis, displacing repeat sequence
  • Instead of being cleaved by Fen1, the flap of a repeat sequence forms a hairpin (stabilised by MutSB), preventing cleavage by Fen1 endonuclease
  • Incorporation of hairpin leads to repeat expansion
17
Q

Transcription coupled repair-based instability:

A
  • Transcription promotes formation of hairpins or slipped strand structures (stabilised by MutSB binding)
  • RNA pol blocked at hairpin -> promotes TC-NER
  • NER may excise hairpin, leading to contraction, or nick on one side of hairpin only, leading to expansion
18
Q

Repeat instability during intergenerational transmission: Influence of parental age

A
  • Influenced by parental age; indicates that repeat instability precedes fertilisation; longer lifetime provides more opportunity for instability
19
Q

Repeat instability during intergenerational transmission: Influence of parental sex

A
  • Influenced by parental sex; differences in spermatogenesis, oogenesis; counterselection for expansions differ in sperm and oocytes; differential DNA repair during spermatogenesis and oogenesis; differential pattern of origin firing; chromatin status in oocytes vs sperm