Genetic Testing- TNR Expansion Flashcards

Question

What is the molecular mechanism for allele expansion?

Answer 1

In spite of much research, unanswered questions remain about the molecular mechanism for allele expansion. There is evidence of "looping" out of one strand of the repeat segment during DNA repair or replication that is thought to account for increased copy number.  In alleles that are large enough to be unstable, expansion occurs in germ cells, and an expanded mutant allele may be passed to the next generation. 

Answer 2

Those where the triplet repeat is in coding sequence, and those where the triplet repeat is in a noncoding portion of the gene. 

Answer 3

CAG, coding for glutamine

Answer 4

When runs of glutamine get too big, the protein has a toxic effect on neurons. In affected individuals, fragments of the abnormal protein are visible as aggregates within neurons

Answer 5

a gain of function abnormality, since presence of the abnormal protein exerts a dominant deleterious effect. Although loss of function (absence) of huntington protein (for example) may have serious developmental consequences, it does NOT cause huntington disease. It’s the TNR expansion that accounts for the observed pathogenicity. 

Answer 6

variable, could be CGG, GAA, CTG, and locations vary within the genes

Answer 7

no protein produced and RNA toxicity

Answer 8

methylation of the promoter region and silencing of the gene

Answer 9

since it is downstream of the normal stop codon, it's deleterious effects of repeat expansion are exerted at the RNA level

Answer 10

expanded polyglutamine causes toxic aggregates to form

Answer 11

a condition where disease severity worsens in successive generations. Anticipation is explained by the fact that repeat copy number tends to expand each time it passes through meiosis (gametogenesis), and disease severity does correlate with repeat copy number. leads to earlier progressive onset (not always) and increased severity in subsequent generations

Answer 12

Considered a targeted approach to testing since assays look directly at the TNR and provide an estimate of repeat number. Tests use patient DNA, and the result is reported as the number of repeats in each allele that the patient has.  Main approach is PCR

Answer 13

T. The goal of TNR testing is to generate a PCR product that spans the repeat segment and has defined end points in conserved flanking sequences. Thus, the size of the PCR product reflects the number of repeats present. The forward and reverse PCR primers would be specific for the relevant gene sequences on either side of the TNR repeat. Thus, for example, a PCR product from a mutant allele with 40 repeats would be 90bp larger than the product from a normal allele that had 10 repeats. Since a person has two copies of the HTT gene, both alleles would be sized. T

Answer 14

For all indications, testing has the capacity to provide an accurate estimate of allele size. While testing can determine allele size in a parent and, also in a fetus or child, it is not possible to make precise predictions about how much a parent's unstable allele will expand in the next generation

Answer 15

NO, others include: - Spinocerebellar ataxia - Kennedy disease - Machado-Joseph disease Note that all these disorders are autosomal dominant, except for Kennedy disease, which is caused by expansion within a gene on the X chromosome

Answer 16

autosomal dominant

Answer 17

40 y/o; 15 years; progressive disease

Answer 18

~5;100,000

Answer 19

TNR expansion leading to toxic aggregates of neurons involved in protein-protein interactions. The polyglutamine tract resulting from the expansion is near the N-terminus

Answer 20

normal- 10-35 (most common is 18) unstable- 36-40 (reduced penetrance and may expand during meiosis) mutated- >40 (symptomatic and will continue to expand in further generations)

Answer 21

T. However, while we think of HD as an adult onset disorder, those with very large expansions (100+) can express symptoms in the first or second decade of life. 

Answer 22

autosomal recessive

Answer 23

autosomal dominant

Answer 24

progressive muscle weakness and wasting, cardiac conduction defects, cataracts, prolonged muscle contraction, variable expressivity correlated to allele size A common description of people with this disorder is that when they engage in a handshake or grab a doorknob, they often have difficulty letting go. 

Answer 25

Yes, there is earlier onset and increased severity with successive generations In comparison, in HD, the primary evidence of anticipation is age of onset. Anticipation may vary in the specifics, but it is a general feature of TNR disorders. 

Answer 26

The pathogenic mechanism in DM1 is not well understood. The expanded TNR in the 3’ UTR of the mRNA causes the mRNA to accumulate in cells and to sequester certain RNA-binding proteins. This sequestration seems to exert a negative trans effect on the ability of other mRNAs to undergo normal translation (i.e. mutant alleles are transcribed but not translated). Thus, the mutation is thought to exert a dominant toxic effect on the DMPK gene and other genes

Answer 27

the normal protein is a protein kinase involved in intercellular conduction and impulse transmission in heart and skeletal muscle

Answer 28

slow progressive ataxia with onset at puberty. Cardiomyopathy is common

Answer 29

FXN encoded Frataxin, a nuclear-encoded gene involved in mitochondrial function

Answer 30

TNR is located in an intron of the FXN gene, where its expansion disrupts normal transcription (loss of function mutation)

Answer 31

Fragile X syndrome

Answer 32

mode: X linked prevalence: 1:5000 men Females heterozygous for a mutation may be affected, but phenotypes are more variable than in males (1/2-2/3 of full mutation females are affected). This variability presumably stems from effects of X inactivation.. All full mutation males are affected

Answer 33

characteristic appearance of long face, large ears, a prominent jaw, and macro-orchidist (enlarged testes) typical IQ in the 30-50 range, delayed developmental milestones, abnormal temperament with tantrums, hyperactivity, autism (Autistic features are common, and for this reason, testing for fragile X syndrome is fairly routine in the work up of children with suspected autism)

Answer 34

The syndrome’s name comes from early observations that during chromosome analysis of affected individuals, a physical break could be elicited at the end of the X chromosome long arm. This is the site of the FMR1 gene, and long trinucleotide expansions cause the chromosome fragility.

Answer 35

abundant in neurons and plays a role in binding mRNA and shuttling it from the nucleus to the cytoplasm

Answer 36

excessive DNA methylation in the 5' UTR of the gene which turns off the gene (loss of function)

Answer 37

6-54. These alleles are stable from one generation to the next, with no risk of fragile X syndrome. 

Answer 38

~30 repeats, although it is variable like other TNRs

Answer 39

females. The larger the premutation, the more likely it is to expand in the next generation. NOTE: Premutation carriers (male and female) do not have fragile X syndrome; their intellect is normal. 

Answer 40

Male premutation carriers are called transmitting males because they will transmit their premutation to all their daughters, but it will not expand in size. Thus, ***In fragile X, expansion from premutation to full mutation occurs only in female meiosis and never in male meiosis.***

Answer 41

Similarly, in myotonic dystrophy, expansion risk is concentrated in females. In particular, children affected with the severe, congenital form almost always have inherited a greatly expanded maternal allele. 

Answer 42

In contrast, although Huntington alleles can expand in both males and females, the most dramatic expansions are transmitted from fathers.  Juvenile onset also more likely to come from expansion of paternal allele

Answer 43

There are interesting phenotypic abnormalities associated with premutations, which I’ll take a minute to discuss. First, whereas full mutations are heavily methylated and not expressed (loss of function), premutations are overexpressed, and once again, we see evidence of ill-defined “RNA toxicity”.  This is associated with minor phenotypic abnormalities. In both male and female premutation carriers, intellect is normal, but there is increased risk for adult onset cerebellar ataxia and intention tremor (FXTAS-fragile X-associated tremor/ataxia syndrome). As expected for X-linked inheritance, this problem is milder in females. However, female premutation carriers have an additional risk; they are at risk for premature ovarian failure (menopause prior to age 40). The broader significance of these observations is that they add another example to a growing list of examples where carriers, once thought to be phenotypically normal, are now known to have mild phenotypic risks. 

Answer 44

- Fragile X syndrome - Huntington disease - Friedrich ataxia - Myotonic dystrophy

Answer 45

CGG repeat on the 5' untranslated end of FMR1 gene on the X chromosome

Answer 46

developmental delay and cognitive impairment

Answer 47

5-54 alleles

Answer 48

In fragile X syndrome, there are intermediate alleles (gray, 55-200 repeats) that are not large enough to cause disease in a person who carries such an allele, but are large enough to be unstable during meiosis (aka the 'premutation' range). Thus, a full mutation and disease may arise in the next generation.

Answer 49

>200 alleles. symptomatic

Answer 50

CAG repeat within an exon of the HTT gene on chromosome 4

Answer 51

progressive neuropsychiatric disorder

Answer 52

>40. Note that there is a small intermediate range, ranging from 36-40

Answer 53

GAA repeat within an intron of the FXN gene on chromosome 9

Answer 54

progressive ataxia

Answer 55

less than 30

Answer 56

CTG repeat in the 3' untranslated region of the DMPK gene on chromosome 19

Answer 57

progressive muscle wasting

Answer 58

less than 35

Answer 59

35-49 (premutation)

Answer 60

mild phenotype: ~50-150 classic phenotype: ~100-1000 congenital: >1000

Answer 61

neurologic and muscular

Answer 62

T. In general, if we examined multiple generations from a normal family, we would expect allele sizes to be perfectly maintained. However, at the same time, there is variation in allele sizes across the normal population. Using the HD gene in normal families as an example, if we looked at one family, we could track an allele with 6 repeats through multiple generations, and there would be no change in its size. A different normal family can have allele sizes anywhere within the normal range (up to ~35 repeats for HD), and these will segregate through that family in a stable manner. 

Answer 63

In spite of much research, unanswered questions remain about the molecular mechanism for allele expansion. There is evidence of "looping" out of one strand of the repeat segment during DNA repair or replication that is thought to account for increased copy number.  In alleles that are large enough to be unstable, expansion occurs in germ cells, and an expanded mutant allele may be passed to the next generation. 

Answer 64

Those where the triplet repeat is in coding sequence, and those where the triplet repeat is in a noncoding portion of the gene. 

Answer 65

CAG, coding for glutamine

Answer 66

When runs of glutamine get too big, the protein has a toxic effect on neurons. In affected individuals, fragments of the abnormal protein are visible as aggregates within neurons

Answer 67

a gain of function abnormality, since presence of the abnormal protein exerts a dominant deleterious effect. Although loss of function (absence) of huntington protein (for example) may have serious developmental consequences, it does NOT cause huntington disease. It’s the TNR expansion that accounts for the observed pathogenicity. 

Answer 68

variable, could be CGG, GAA, CTG, and locations vary within the genes

Answer 69

no protein produced and RNA toxicity

Answer 70

methylation of the promoter region and silencing of the gene

Answer 71

since it is downstream of the normal stop codon, it's deleterious effects of repeat expansion are exerted at the RNA level

Answer 72

expanded polyglutamine causes toxic aggregates to form

Answer 73

a condition where disease severity worsens in successive generations. Anticipation is explained by the fact that repeat copy number tends to expand each time it passes through meiosis (gametogenesis), and disease severity does correlate with repeat copy number. leads to earlier progressive onset (not always) and increased severity in subsequent generations

Answer 74

Considered a targeted approach to testing since assays look directly at the TNR and provide an estimate of repeat number. Tests use patient DNA, and the result is reported as the number of repeats in each allele that the patient has.  Main approach is PCR

Answer 75

T. The goal of TNR testing is to generate a PCR product that spans the repeat segment and has defined end points in conserved flanking sequences. Thus, the size of the PCR product reflects the number of repeats present. The forward and reverse PCR primers would be specific for the relevant gene sequences on either side of the TNR repeat. Thus, for example, a PCR product from a mutant allele with 40 repeats would be 90bp larger than the product from a normal allele that had 10 repeats. Since a person has two copies of the HTT gene, both alleles would be sized. T

Answer 76

For all indications, testing has the capacity to provide an accurate estimate of allele size. While testing can determine allele size in a parent and, also in a fetus or child, it is not possible to make precise predictions about how much a parent's unstable allele will expand in the next generation

Answer 77

NO, others include: - Spinocerebellar ataxia - Kennedy disease - Machado-Joseph disease Note that all these disorders are autosomal dominant, except for Kennedy disease, which is caused by expansion within a gene on the X chromosome

Answer 78

autosomal dominant

Answer 79

40 y/o; 15 years; progressive disease

Answer 80

~5;100,000

Answer 81

TNR expansion leading to toxic aggregates of neurons involved in protein-protein interactions. The polyglutamine tract resulting from the expansion is near the N-terminus

Answer 82

normal- 10-35 (most common is 18) unstable- 36-40 (reduced penetrance and may expand during meiosis) mutated- >40 (symptomatic and will continue to expand in further generations)

Answer 83

T. However, while we think of HD as an adult onset disorder, those with very large expansions (100+) can express symptoms in the first or second decade of life. 

Answer 84

autosomal recessive

Answer 85

autosomal dominant

Answer 86

progressive muscle weakness and wasting, cardiac conduction defects, cataracts, prolonged muscle contraction, variable expressivity correlated to allele size A common description of people with this disorder is that when they engage in a handshake or grab a doorknob, they often have difficulty letting go. 

Answer 87

Yes, there is earlier onset and increased severity with successive generations In comparison, in HD, the primary evidence of anticipation is age of onset. Anticipation may vary in the specifics, but it is a general feature of TNR disorders. 

Answer 88

The pathogenic mechanism in DM1 is not well understood. The expanded TNR in the 3’ UTR of the mRNA causes the mRNA to accumulate in cells and to sequester certain RNA-binding proteins. This sequestration seems to exert a negative trans effect on the ability of other mRNAs to undergo normal translation (i.e. mutant alleles are transcribed but not translated). Thus, the mutation is thought to exert a dominant toxic effect on the DMPK gene and other genes

Answer 89

the normal protein is a protein kinase involved in intercellular conduction and impulse transmission in heart and skeletal muscle

Answer 90

slow progressive ataxia with onset at puberty. Cardiomyopathy is common

Answer 91

FXN encoded Frataxin, a nuclear-encoded gene involved in mitochondrial function

Answer 92

TNR is located in an intron of the FXN gene, where its expansion disrupts normal transcription (loss of function mutation)

Answer 93

Fragile X syndrome

Answer 94

mode: X linked prevalence: 1:5000 men Females heterozygous for a mutation may be affected, but phenotypes are more variable than in males (1/2-2/3 of full mutation females are affected). This variability presumably stems from effects of X inactivation.. All full mutation males are affected

Answer 95

characteristic appearance of long face, large ears, a prominent jaw, and macro-orchidist (enlarged testes) typical IQ in the 30-50 range, delayed developmental milestones, abnormal temperament with tantrums, hyperactivity, autism (Autistic features are common, and for this reason, testing for fragile X syndrome is fairly routine in the work up of children with suspected autism)

Answer 96

The syndrome’s name comes from early observations that during chromosome analysis of affected individuals, a physical break could be elicited at the end of the X chromosome long arm. This is the site of the FMR1 gene, and long trinucleotide expansions cause the chromosome fragility.

Answer 97

abundant in neurons and plays a role in binding mRNA and shuttling it from the nucleus to the cytoplasm

Answer 98

excessive DNA methylation in the 5' UTR of the gene which turns off the gene (loss of function)

Answer 99

6-54. These alleles are stable from one generation to the next, with no risk of fragile X syndrome. 

Answer 100

~30 repeats, although it is variable like other TNRs

Answer 101

females. The larger the premutation, the more likely it is to expand in the next generation. NOTE: Premutation carriers (male and female) do not have fragile X syndrome; their intellect is normal. 

Answer 102

Male premutation carriers are called transmitting males because they will transmit their premutation to all their daughters, but it will not expand in size. Thus, ***In fragile X, expansion from premutation to full mutation occurs only in female meiosis and never in male meiosis.***

Answer 103

Similarly, in myotonic dystrophy, expansion risk is concentrated in females. In particular, children affected with the severe, congenital form almost always have inherited a greatly expanded maternal allele. 

Answer 104

In contrast, although Huntington alleles can expand in both males and females, the most dramatic expansions are transmitted from fathers.  Juvenile onset also more likely to come from expansion of paternal allele

Answer 105

There are interesting phenotypic abnormalities associated with premutations, which I’ll take a minute to discuss. First, whereas full mutations are heavily methylated and not expressed (loss of function), premutations are overexpressed, and once again, we see evidence of ill-defined “RNA toxicity”.  This is associated with minor phenotypic abnormalities. In both male and female premutation carriers, intellect is normal, but there is increased risk for adult onset cerebellar ataxia and intention tremor (FXTAS-fragile X-associated tremor/ataxia syndrome). As expected for X-linked inheritance, this problem is milder in females. However, female premutation carriers have an additional risk; they are at risk for premature ovarian failure (menopause prior to age 40). The broader significance of these observations is that they add another example to a growing list of examples where carriers, once thought to be phenotypically normal, are now known to have mild phenotypic risks. 