Summary Fragile-X syndrome is a trinucleotide-repeat–expansion disorder in which the clinical phenotype is believed to result from transcriptional silencing of the fragile-X mental retardation 1 ( FMR1 ) gene as the number of CGG repeats exceeds ∼200. For premutation alleles (∼55–200 repeats), no abnormalities in FMR1 -gene expression have been described, despite growing evidence of clinical involvement in premutation carriers. To address this (apparent) paradox, we have determined, for 16 carrier males (55–192 repeats), the relative levels of leukocyte FMR1 mRNA, by use of automated fluorescence-detection reverse transcriptase–PCR, and the percent of lymphocytes that are immunoreactive for FMR1 protein (FMRP). For some alleles with >100 repeats, there was a reduction in the number of FMRP-positive cells. Unexpectedly, FMR1 mRNA levels were elevated at least fivefold within this same range. No significant increase in FMR1 mRNA stability was observed in a lymphoblastoid cell line (160 repeats) derived from one of the carrier males, suggesting that the increased message levels are due to an increased rate of transcription. Current results support a mechanism of involvement in premutation carriers, in which reduced translational efficiency is at least partially compensated through increased transcriptional activity. Thus, diminished translational efficiency may be important throughout much of the premutation range, with a mechanistic switch occurring in the full-mutation range as the FMR1 gene is silenced.

Elevated levels of FMR1 mRNA in carrier males : A new mechanism of involvement in the fragile-X syndrome

FMR1 and the fragile X syndrome: Human genome epidemiology review

The 5' untranslated CGG repeat in the fragile X mental retardation-1 (FMR1) gene is expanded in families with fragile X syndrome, with more than 200 CGGs resulting in mental retardation due to the absence of the encoded fragile X mental retardation protein (FMRP). Intermediate and premutation alleles, containing between approximately 40 and 200 repeats, express grossly normal FMRP levels and such carriers are widely believed to be non-penetrant, despite continued reports of subtle cognitive/psychosocial impairment and other phenotypes. Using a highly sensitive quantification assay, we demonstrate significantly diminished FMRP levels in carriers, negatively correlated with repeat number. Despite reduced FMRP, these carrier alleles overexpress FMR1, resulting in a positive correlation between repeat number and FMR1 message level. These biochemical deviations associated with intermediate and premutation FMR1 alleles, found in approximately 4% of the population, suggest that the phenotypic spectrum of fragile X syndrome may need to be revisited.

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Reduced FMRP and increased FMR1 transcription is proportionally associated with CGG repeat number in intermediate-length and premutation carriers

A Consensus Conference utilizing available literature and expert opinion sponsored by the American College of Medical Genetics in October 1995 evaluated the rational approach to the individual with mental retardation. Although no uniform protocol replaces individual clinician judgement, the consensus recommendations were as follows: 1. The individual with mental retardation, the family, and medical care providers benefit from a focused clinical and laboratory evaluation aimed at establishing causation and in providing counseling, prognosis, recurrence risks, and guidelines for management. 2. Essential elements of the evaluation include a three-generation pedigree: pre-, peri-, and post-natal history, complete physical examination focused on the presence of minor anomalies, neurologic examination, and assessment of the behavioral phenotype. 3. Selective laboratory testing should, in most patients, include a banded karyotype. Fragile X testing should be strongly considered in both males and females with unexplained mental retardation, especially in the presence of a positive family history, a consistent physical and behavioral phenotype and absence of major structural abnormalities. Metabolic testing should be initialed in the presence of suggestive clinical and physical findings. Neuroimaging should be considered in patients without a known diagnosis especially in the presence of neurologic symptoms, cranial contour abnormalities, microcephaly, or macrocephaly. In most situations MRI is the testing modality of choice. 4. Sequential evaluation of the patient, occasionally over several years, is often necessary for diagnosis, allowing for delineation of the physical and behavioral phenotype, a logical approach to ancillary testing and appropriate prognostic and reproductive counseling.

Evaluation of mental retardation: Recommendations of a consensus conference

The present study extends our previous work on social behavior impairment in young males with fragile X syndrome (FraX). Specifically, we evaluated whether the autistic phenomenon in FraX is expressed as a range of behavioral impairments as in idiopathic autism (Aut). We also examined whether there are behaviors, identified as items of the Autism Diagnostic Interview-Revised (ADIR), that in FraX predispose to or differentiate subjects with autism spectrum disorder (ASD) diagnosis. Finally, regression models were utilized to test the relative contribution of reduced communication and socialization skills to ADI-R scores and diagnoses. A cohort of 56 boys (3– 8 years) with FraX was examined in terms of scores on measures of cognition (IQ was a co-variate in most analyses.), autistic behavior, problem/aberrant behavior, adaptive behavior, and language development. We found that, indeed, in terms of problem behavior and adaptive skills, there is a range of severity from FraX þ Aut to FraX þ PDD (Pervasive Developmental Disorder) to FraX þ none. ADI-R items representing ‘‘Play’’ types of interaction appear to be ‘‘susceptibility’’ factors since they were abnormal across the FraX cohort. Integrated regression models demonstrated that items reflecting complex social interaction differentiated the FraX þ ASD (Aut þ PDD) subgroup from the rest of the FraX cohort, while abnormalities in basic verbal and non-verbal communication distinguished the most severely affected boys with FraX þ Aut from the milder FraX þ PDD cohort. Models incorporating language, adaptive communication, and adaptive socialization skills revealed that socialization was not only the main influence on scores but also a predictor of ASD diagnosis. Altogether, our findings demonstrate

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Autism Spectrum Disorder in Fragile X Syndrome: Communication, Social Interaction, and Specific Behaviors

We report the results of a 14-center collaborative study of genotype-phenotype correlations in 318 fragile X families; these families comprised 2,253 individuals, 1,344 of whom carried a fragile X mutation and 693 of whom had a typical full fragile X mutation This study demonstrates that direct DNA diagnosis establishes the genotype at the FRAXA-FMR-1 locus There was a significantly higher prevalence of “mosaic” cases among males who carry a full mutation (12%) than among females who carry a full mutation (6%); the mosaic males had a larger expansion than did the mosaic females Mental status of premutated individuals did not differ from that of those with a normal genotype Both the abnormal methylation of the FMR-1–EagI site and the size of the expansion were highly correlated with cytogenetics, facial dysmorphism, macroorchidism, and mental retardation (MR) Among female carriers of a full mutation, those with MR had significantly larger expansion than did those without MR Among 164 independent couples, 3 unrelated husbands carried a premutation that suggests that the prevalence of fragile X premutations in the general population is ~09% of the X chromosomes Our data validate the use of direct DNA testing for fragile X diagnosis as well as for carrier identification and support and complete the established relationships among the DNA results and the cytogenetic, physical, and psychological aspects of the disease

E Mornet

Papers

A Multicenter Study on Genotype-Phenotype Correlations in the Fragile X Syndrome, Using Direct Diagnosis with Probe StB12.3: The First 2,253 Cases