scholarly journals Combined Molecular and Cytogenetic Analysis for the Rapid Diagnosis of Fragile X Syndrome

1996 ◽  
Vol 45 (1-2) ◽  
pp. 165-168
Author(s):  
C. Bussani Mastellone ◽  
M.L. Giovannucci Uzielli ◽  
M. Grasso ◽  
P. Chiurazzi ◽  
G. Neri ◽  
...  
Keyword(s):  
Southern Blotting ◽  
Screening Programme ◽  
Fragile X ◽  
Repeat Sequence ◽  
Molecular Techniques ◽  
Cgg Repeat ◽  
Normal Carrier ◽  
Prevention Studies ◽  
Cytogenetic Testing

AbstractThe fragile X mutation is the result of an abnormal expansion of a CGG repeat sequence in the FMR-1 gene.Molecular techniques enable the detection of the mutation and also of the exact length of this DNA sequence, allowing the classification of the tested subjects as normal, carrier or affected.We propose a protocol of analysis that combines a method of non-radioactive PCR, Southern blotting and cytogenetic testing.This protocol can be used for screening programme of selected groups of mentally retarded individuals and for prevention studies in families at risk.

Diagnosis and Prevention of Fragile-X Syndrome. From the Family Study to the Population Screening Programme: Eighteen Years of Activity

1996 ◽  
Vol 45 (1-2) ◽  
pp. 303-308
Author(s):  
M.L. Giovannucci Uzielli ◽  
S. Guarducci ◽  
A. Cecconi ◽  
S. Lenzi ◽  
U. Ricci ◽  
...  
Keyword(s):  
Fragile X Syndrome ◽  
Family Study ◽  
Screening Programme ◽  
Fragile Site ◽  
Fragile X ◽  
Cgg Repeat ◽  
Transcriptional Suppression ◽  
Evolutionarily Conserved ◽  
The Family ◽  
Conserved Gene

Fragile-X syndrome, which derives its name from the expression of a fragile site (FRAXA) at Xq27.3 associated with the phenotype, has achieved distinction as the most common inherited cause of mental retardation. It is the first disorder shown to be due to dynamic mutation in heritable instable DNA.In 1991 the mutation responsible for Fragile-X syndrome was delineated as an expansion of the trinucleotide (CGG) sequence within an evolutionarily conserved gene, at the position of the fragile-X site.The DNA of the promoter in the 5' UTR region of FMR-1 gene becomes abnormally methylated when the CGG sequence exceeds approximately 230 repeats, resulting in the transcriptional suppression of FMR-1. Based on the length of CGG repeat in the FMR-1 gene, the alleles are usually classified as normal, premutation or full mutation. CGG instability correlates with the length of repeats and number of AGGs within the FMR-1 CGG tract. In a minority of cases the Fragile-X syndrome may be due to deletion, or to point mutation in the FMR-1 gene.

scholarly journals A Novel FMR1 PCR Method for the Routine Detection of Low Abundance Expanded Alleles and Full Mutations in Fragile X Syndrome

2010 ◽  
Vol 56 (3) ◽  
pp. 399-408 ◽  
Author(s):  
Stela Filipovic-Sadic ◽  
Sachin Sah ◽  
Liangjing Chen ◽  
Julie Krosting ◽  
Edward Sekinger ◽  
...  
Keyword(s):  
Fragile X Syndrome ◽  
Southern Blotting ◽  
Trinucleotide Repeat ◽  
Fragile X ◽  
Clinical Samples ◽  
Full Mutation ◽  
Cgg Repeat ◽  
Specific Pcr ◽  
Cgg Repeats ◽  
Pcr Technology

Abstract Background: Fragile X syndrome (FXS) is a trinucleotide-repeat disease caused by the expansion of CGG sequences in the 5′ untranslated region of the FMR1 (fragile X mental retardation 1) gene. Molecular diagnoses of FXS and other emerging FMR1 disorders typically rely on 2 tests, PCR and Southern blotting; however, performance or throughput limitations of these methods currently constrain routine testing. Methods: We evaluated a novel FMR1 gene–specific PCR technology with DNA templates from 20 cell lines and 146 blinded clinical samples. The CGG repeat number was determined by fragment sizing of PCR amplicons with capillary electrophoresis, and results were compared with those for FMR1 Southern blotting analyses with the same samples. Results: The FMR1 PCR accurately detected full-mutation alleles up to at least 1300 CGG repeats and consisting of >99% GC character. All categories of alleles detected by Southern blotting, including 66 samples with full mutations, were also identified by the FMR1 PCR for each of the 146 clinical samples. Because all full mutation alleles in samples from heterozygous females were detected by the PCR, allele zygosity was reconciled in every case. The PCR reagents also detected a 1% mass fraction of a 940-CGG allele in a background of 99% 23-CGG allele—a roughly 5- fold greater sensitivity than obtained with Southern blotting. Conclusions: The novel PCR technology can accurately categorize the spectrum of FMR1 alleles, including alleles previously considered too large to amplify; reproducibly detect low abundance full mutation alleles; and correctly infer homozygosity in female samples, thus greatly reducing the need for sample reflexing to Southern blotting.

Evidence for an FMR1 mRNA Gain-Of-Function Toxicity Mechanism in the Pathogenesis of Fragile-X Associated Premature Ovarian Insufficiency

2021 ◽  
Author(s):  
Roseanne Rosario ◽  
Hazel Stewart ◽  
Nila Roy Choudhury ◽  
Gracjan Michlewski ◽  
Nicolas Charlet-Berguerand ◽  
...  
Keyword(s):  
Granulosa Cell ◽  
Cell Model ◽  
Fragile X ◽  
Repeat Sequence ◽  
Premature Ovarian Insufficiency ◽  
Gain Of Function ◽  
Ovarian Insufficiency ◽  
Cgg Repeat ◽  
Function Mechanism

Abstract Fragile X-associated premature ovarian insufficiency (FXPOI) is caused by expansion of a CGG repeat sequence located in the 5’ untranslated region of the FMR1 gene. Women with FXPOI have a depleted ovarian reserve, resulting in amenorrhea, hypoestrogenism, and loss of fertility before the age of 40. FXPOI is caused by CGG sequence expansions to lengths between 55 and 200 repeats, known as a FMRI premutation, however the mechanism by which the premutation drives disease pathogenesis remains unclear. Two main hypotheses exist, which describe an mRNA toxic gain-of-function mechanism or that repeat-associated non-AUG (RAN) translation results in the production of an abnormal protein, called FMRpolyG. We have developed an in vitro granulosa cell model of the FMR1 premutation by ectopically expressing CGG-repeat RNA and FMRpolyG protein. We show that expanded CGG-repeat RNA accumulated in intranuclear RNA structures, and these aggregates were able to cause significant granulosa cell death independent of FMRpolyG expression. Furthermore, using an innovative RNA pulldown, mass spectrometry-based approach we have identified proteins that bind CGG-repeat RNA in granulosa cells in vitro, and thus may be deregulated as consequence of this interaction. Collectively, these data provide evidence for the contribution of an mRNA gain-of-function mechanism to FXPOI disease biology.

scholarly journals Secondary structural choice of DNA and RNA associated with CGG/CCG trinucleotide repeat expansion rationalizes the RNA misprocessing in FXTAS

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yogeeshwar Ajjugal ◽  
Narendar Kolimi ◽  
Thenmalarchelvi Rathinavelan
Keyword(s):  
Rna Binding ◽  
Trinucleotide Repeat ◽  
Rna Binding Proteins ◽  
Fragile X ◽  
Repeat Expansion ◽  
Mobility Shift ◽  
Cgg Repeat ◽  
Dna And Rna ◽  
Structural Choice ◽  
Sense Strand

AbstractCGG tandem repeat expansion in the 5′-untranslated region of the fragile X mental retardation-1 (FMR1) gene leads to unusual nucleic acid conformations, hence causing genetic instabilities. We show that the number of G…G (in CGG repeat) or C…C (in CCG repeat) mismatches (other than A…T, T…A, C…G and G…C canonical base pairs) dictates the secondary structural choice of the sense and antisense strands of the FMR1 gene and their corresponding transcripts in fragile X-associated tremor/ataxia syndrome (FXTAS). The circular dichroism (CD) spectra and electrophoretic mobility shift assay (EMSA) reveal that CGG DNA (sense strand of the FMR1 gene) and its transcript favor a quadruplex structure. CD, EMSA and molecular dynamics (MD) simulations also show that more than four C…C mismatches cannot be accommodated in the RNA duplex consisting of the CCG repeat (antisense transcript); instead, it favors an i-motif conformational intermediate. Such a preference for unusual secondary structures provides a convincing justification for the RNA foci formation due to the sequestration of RNA-binding proteins to the bidirectional transcripts and the repeat-associated non-AUG translation that are observed in FXTAS. The results presented here also suggest that small molecule modulators that can destabilize FMR1 CGG DNA and RNA quadruplex structures could be promising candidates for treating FXTAS.

scholarly journals Autism Profiles of Males With Fragile X Syndrome

2008 ◽  
Vol 113 (6) ◽  
pp. 427-438 ◽  
Author(s):  
Susan W. Harris ◽  
David Hessl ◽  
Beth Goodlin-Jones ◽  
Jessica Ferranti ◽  
Susan Bacalman ◽  
...  
Keyword(s):  
Fragile X Syndrome ◽  
Fragile X ◽  
Autism Diagnosis ◽  
Cgg Repeat ◽  
Molecular Features ◽  
Fmr1 Mrna ◽  
Two Measures ◽  
Dsm Iv ◽  
Relationship Of ◽  
The Relationship

Abstract Autism, which is common in individuals with fragile X syndrome, is often difficult to diagnose. We compared the diagnostic classifications of two measures for autism diagnosis, the ADOS and the ADI-R, in addition to the DSM-IV-TR in 63 males with this syndrome. Overall, 30% of the subjects met criteria for autistic disorder and 30% met criteria for PDD-NOS. The classifications on the ADOS and DSM-IV-TR were most similar, whereas the ADI-R classified subjects as autistic much more frequently. We further investigated the relationship of both FMRP and FMR1 mRNA to symptoms of autism in this cohort and found no significant relationship between the measures of autism and molecular features, including FMRP, FMR1 mRNA, and CGG repeat number.

CGG repeat length correlates with age of onset of motor signs of the fragile X-associated tremor/ataxia syndrome (FXTAS)

2007 ◽  
Vol 144B (4) ◽  
pp. 566-569 ◽  
Author(s):  
Flora Tassone ◽  
John Adams ◽  
Elizabeth M. Berry-Kravis ◽  
Susannah S. Cohen ◽  
Alfredo Brusco ◽  
...  
Keyword(s):  
Age Of Onset ◽  
Fragile X ◽  
Repeat Length ◽  
Cgg Repeat ◽  
Ataxia Syndrome

scholarly journals ASFMR1splice variant

2018 ◽  
Vol 4 (4) ◽  
pp. e246 ◽  
Author(s):  
Padmaja Vittal ◽  
Shrikant Pandya ◽  
Kevin Sharp ◽  
Elizabeth Berry-Kravis ◽  
Lili Zhou ◽  
...  
Keyword(s):  
Mental Retardation ◽  
Splice Variant ◽  
Messenger Rna ◽  
Clinical Symptoms ◽  
Fragile X ◽  
Cgg Repeat ◽  
Men And Women ◽  
Fragile X Mental Retardation ◽  
Repeat Size ◽  
Ataxia Syndrome

ObjectiveTo explore the association of a splice variant of theantisense fragile X mental retardation 1(ASFMR1) gene, loss offragile X mental retardation 1(FMR1) AGG interspersions andFMR1CGG repeat size with manifestation, and severity of clinical symptoms of fragile X-associated tremor/ataxia syndrome (FXTAS).MethodsPremutation carriers (PMCs) with FXTAS, without FXTAS, and normal controls (NCs) had a neurologic evaluation and collection of skin and blood samples. Expression ofASFMR1transcript/splice variant 2 (ASFMR1-TV2), nonsplicedASFMR1, totalASFMR1, andFMR1messenger RNA were quantified and compared using analysis of variance. Least absolute shrinkage and selection operator (LASSO) logistic regression and receiver operating characteristic analyses were performed.ResultsPremutation men and women both with and without FXTAS had higherASFMR1-TV2 levels compared with NC men and women (n = 135,135,p< 0.0001), andASFMR1-TV2 had good discriminating power for FXTAS compared with NCs but not for FXTAS from PMC. After adjusting for age, loss of AGG, larger CGG repeat size (in men), and elevatedASFMR1-TV2 level (in women) were strongly associated with FXTAS compared with NC and PMC (combined).ConclusionsThis study found elevated levels ofASFMR1-TV2and loss of AGG interruptions in both men and women with FXTAS. Future studies will be needed to determine whether these variables can provide useful diagnostic or predictive information.

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