18.03.09 Fragile X

Question 1

Where is FMR1 and what disorders are associated with mutations of this genes?

Answer 1

FMR1 Xq27.3

Fragile X syndrome
FXTAS
FXPOI

Question 2

What is the underlying disease mechanism of FMR1-associated disorders?

Answer 2

CGG triplet repeat expansion in 5’UTR

X-linked dominant with variable penetrance.

Question 3

What is the function of FMRP?

Answer 3

FMRP regulates the production of other proteins and plays a role in the development of synapses.

FMRP is thought to act as a shuttle within cells by transporting messenger RNA (mRNA) from the nucleus to areas of the cell where proteins are assembled.

FMRP also helps to control when the instructions in these mRNA molecules are used to build proteins

Question 4

Where is FMRP expressed?

Answer 4

FMRP is present in many tissues including the brain, testes and ovaries.

Question 5

What is the effect of a repeat expansion in the 5’UTR of FMR1 OF >200 repeats?

Answer 5

Expansion of an unstable 5’UTR CGG repeat in FMR1 to over 200 repeats (full mutations) causes gene silencing.

Aberrant methylation of a CpG island 250 base pairs downstream of the CGG repeat causes Fragile X syndrome in (male) patients when the CGG tract expands to over 200 repeats.

Full mutations (>200 repeats) result in hypermethylation of the DNA in and around the CGG tract, curtailed gene expression and no FMRP production.

Question 6

What is the underlying molecular pathology of FXTAS and FXPOI?

Answer 6

Expansion of the CGG repeat tract into the premutation range.

Premutation carriers have been found to have higher levels of FMR1 mRNA and normal levels of FMRP.

FXTAS and POI are unlikely to be due to a lack of FMRP as these symptoms are not present in Fragile X affected patients caused by a full mutation. It is possible that the excess of FMR1 mRNA could be responsible for a toxic gain of function phenotype resulting in FXTAS and POI.

Question 7

What is the incidence of Fragile X syndrome?

Answer 7

Fragile X syndrome is probably the most common single-gene cause of learning disability in humans.

Estimated prevalence of approximately 1 in 4000 males and 1 in 5000-8000 females.

Question 8

What is the phenotype of Fragile X syndrome in males?

Answer 8

In males, it is characterised by moderate to severe intellectual and social impairment, characteristic appearance (large head, long face, large ears, prominent forehead and chin, protruding ears), joint laxity and macro-orchidism.

Question 9

What is the phenotype of Fragile X syndrome in females?

Answer 9

Females with a full mutation have variable phenotypes ranging from apparently normal (about 50%) to mild to moderate mental and social impairment, with or without fragile site expression.

Question 10

How can Fragile X syndrome be detected cytogenetically?

Answer 10

A fragile site (FRAXA) is expressible at the gene locus at Xq27.3 in around 2-40% of blood cells in affected males.

Question 11

What is the size of the smallest repeat known to expand to a full mutation in one generation?

Answer 11

56 (no AGG interruptions)

Question 12

What is thought to be the cause of the variable expression of Fragile X syndrome in females?

Answer 12

Variable phenotype in females are due to skewed X-inactivation

Question 13

What % of Fragile X syndrome cases do not have a pathogenic CGG expansion?

Answer 13

1% caused by point mutations or partial/whole deletions

These mutations do not exhibit fragile site expression of hypomethylation

Question 14

What proportion of females with a premutation will experience FXPOI? What is the phenotype of FXPOI?

Answer 14

20%

POI in females is characterised by cessation of menses before age 40 years. In addition, age of menopause is on average five years earlier in premutation carriers compared to non-carriers.

Question 15

What % of POI does FXPOI account for?

Answer 15

FMR1-related POI accounts for about 4% of all POI and around 12% of familial POI.

Question 16

What are the features of FXTAS?

Answer 16

FXTAS occurs in males (and some females) who have an FMR1 premutation and is characterised by late-onset progressive cerebellar ataxia and intention tremor.

Question 17

Is the penetrance of FXTAS higher in males or females?

Answer 17

Males (45.5% v 16.5%) - no ‘normal’ X to compensate

Question 18

Describe the FRAXE locus.

Answer 18

FRAXE (Xq28)

FRAXE is a distinct condition caused by mutations in the FMR2 gene

It is associated with a fragile site (FRAXE).

Full FRAXE mutations are large expansions of a GCC tract in the 5’ UTR of FMR2, deriving from expansions of smaller premutation alleles.

The FRAXE disease phenotype is considerably less severe than the Fragile X of FRAXA and lacks the specific syndromic features.

The prevalence of FRAXE full mutations is much lower than that of FRAXA.

No disease phenotype has been attributed to FRAXE premutations.

Question 19

Describe FMR1 normal alleles and the associated repeat range.

Answer 19

6-45 repeats

Stably transmitted without any increase or decrease in size or repeat number.

Question 20

Describe the role of AGG interspersions in CGG repeat tracts.

Answer 20

The CGG repeats are interrupted every 9-10 repeats by an AGG and this helps to maintain integrity, typically there are 2 or more AGG interspersions.

Repeat tracts of >35 CGG repeats with AGG interruptions are more likely to be unstable.

Most normal and intermediate alleles consist of (CGG)¬9 or 10AGG(CGG)9AGG(CGG)n, the distal tract of CGG accounting for most of the length variation between alleles.

Since unstable premutations are usually pure CGG repeats or contain only a proximal AGG interspersion, it is assumed that the loss of one or both AGG motifs or their conversion to CGG in an intermediate allele would predispose it to instability.

Question 21

Describe FMR1 intermediate alleles and the associated repeat range.

Answer 21

46-58 repeats (under 50 likely to be stable, 50-58 may show instability).

These alleles represent the overlap zone between stable normal alleles and unstable premutations.

Often transmitted stably but are more likely to show unstable transmission with increasing size in this range, usually by only one or two repeats.

The stability of transmissions can vary within a family.

It is not known whether any alleles in the intermediate range show clinical involvement in abnormal phenotypes such as POI, FXTAS or developmental delay.

The stability of an intermediate allele appears to correlate with the presence of two or more interspersed AGG motifs within the CGG tract.

Question 22

Describe FMR1 premutation alleles and the associated repeat range.

Answer 22

59-200 unmethylated on active X chromosomes but are subject to X-inactivation in females.

They are unmethylated with normal transcript and protein levels, but are extremely unstable during transmission to the next generation. Expansion from premutation to full mutation only occurs in maternal transmission and this is dependent on the length of the maternal premutation.

The distinction between a premutation and a full mutation is more likely due to methylation status rather than the exact size of the allele and so a methylation-sensitive enzyme is combined with another enzyme to give a convenient size of fragment for resolution of the expansion and their methylation status.

Question 23

What is the prevalence of FMR1 premutation alleles in the general population?

Answer 23

1 in 800 males

1 in 250 females

Question 24

What is the probability that a premutation will expand to a full mutation upon maternal transmission?

Answer 24

The probability of conversion to a full mutation on maternal transmission in a single generation is low for premutations of 59-70 repeats but rises to >90% for premutations of more than 90 repeats.

Question 25

Describe FMR1 full mutation alleles and the associated repeat range.

Answer 25

> 200 - ~1000 repeats; methylation of the DNA in and around the expanded repeat tract, even on the active X chromosome.

Question 26

What proportion of FMR1 full mutation patients show mosaicism?

Answer 26

Approximately 20% of full mutation patients also show some mosaicism for a premutation which may be the same size (methylation mosaic) but is usually smaller than the full mutation. Some patients are methylation mosaic, in which full mutations have varying degrees of methylation.

Almost always, extensive somatic variation of repeat number is observed in a peripheral blood sample of a patient with a full mutation.

The southern blot double digest detects most expansions but may have limits to its sensitivity in cases of full mutations which appear as diffuse smears due to somatic mosaicism (particularly in females, where the presence of the normal allele may draw attention away from any faint expanded fragments).

Question 27

Describe the phenotype of an unmethylated full mutation.

Answer 27

Nearly all males have ID, but often have high-functioning MR to low-normal intellect.

Females are highly variable. Range form normal intellect to affected.

Question 28

Describe the phenotype of a full mutation with partial methylation (methylation mosaicism).

Answer 28

Males: Nearly 100% affected with ID; may be higher functioning than males with full mutation.

Females: Highly variable: ranges from normal intellect to affected

Question 29

Describe the phenotype of a premutation/ full mutation mosaic.

Answer 29

Methylation: Partial as unmethylated in the premutation cell line; methylated in the full mutation cell line

Males: Nearly 100% affected with ID; may be higher functioning than males with full mutation

Females: Highly variable: ranges from normal intellect to affected