Download Fragile X Syndrome

Fragile X Syndrome
(Martin-Bell Syndrome)
Amber Boone
 www.fragilex.org.uk/ page6.htm
Characteristics
 Mild to Moderate
Mental Retardation
 Long, narrow face
 Large, protuberant
ears
 Macroorchidism
(enlarged testicles)
Background
 X-linked disease
 Mutation is located at Xq27.3
 FMR1 Gene
– Polymorphic (CCG)n repeat in the 5’
untranslated reagion of exon 1
– Hypermethylation of a CpG island upstream of
the mutation
Finding the Causitive Gene
 Cloned the X Chromosome from a normal
human into YACs
 Digested with EcoRI
– Found a 5 Kb region that was unstable in
pedigrees with Fragile X (pfxa1)
 Digested with PSTI
– Narrowed the instability down to a 1 Kb region
(pfxa2)
 Sequenced this region
Pfxa2 sequence
Finding the Causitive Gene (cont)
 Used several RE to cut normal and Fragile
X DNA isolated from human lymphnodes
– Normal fragile X site varied from 45-95 bp
– Infected individuals fragile X site was almost
900 bp longer
Finding the Causitive Gene (Cont)
 Physical map across
the Fragile X region
– Mostly done through
Resriction Enzyme
cleavage
Finding the Causitive Gene (cont)
 Isolated a YAC in somatic cell hyprids containing
part of the Fragile X site
 Bought a YAC library and used their clone as a
probe
– Obtained a YAC with the whole region
 Created a cosmid library from the YAC clone
 Cosmid subclones used to screen a cDNA library
of human fetal brain RNA
– The cosmids hybridized to a portion of a gene
designated as FMR1
Pedigree of a family with the Fragile
X mutation segregating
FMR1 Expression
 Northern Blot of FMR1 was
done on Human Tissue
– Expressed in highest levels in
the Brain and Testes
– Slightly lower level in the
Placenta, Lungs, Liver, and
Kidneys
 FMR1 expression was turned
on early in embroyonic
development
Mouse Knockout
More Findings
 Fragile X phenotype is caused from lack of
FMRP expression
 FMRP was expressed in some men with full
mutation, but no methylation
 Found alternatively spiced FMRP proteins in
different locations in the body
Treatment
 Study on in vitro
reactivation
– Rare cases of individuals
with the full mutation and
unmethylation have shown
that the problem is in the
methylation, which inhibits
the translation
– 5-azadeoxycytidine to
induce DNA demethylation in
vitro
Bibliography
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