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 1100 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.

Both microscopic and macroscopic models of DNA flexibility should lead to the same quantitative description of the elastic properties of the DNA helix. This belief is reinforced by the fact that essentially all experimental (solution) studies to date support the macroscopic, elastic model. The performance of microscopic models can therefore be checked by their ability to produce the correct macroscopic quantities (P and C). To most carefully address the influence of such factors as base sequence, DNA damage, and drug or protein interaction on the flexibility of DNA, methods are required that are most sensitive for DNA molecules of less than 500-1000 bp. The use of molecules in this size range will maximize the signal due to the structural alteration as well as facilitate the construction of DNA sequences of any desired arrangement. I have emphasized three such methods and summarized their strengths and weaknesses; however, their concurrent application to the determination of DNA flexibility provides an important check of self-consistency. These studies have indicated that the persistence length of DNA in buffers of moderate salt concentration is 450-500 A. Synthetic DNA is now readily available, and many procedures for the construction and cloning of DNA molecules of defined length and sequence (107-108a) are in common use. The availability of restriction fragments of precisely defined length has transformed the study of the physical (particularly hydrodynamic) properties of such molecules, since the hitherto pervasive problem of length polydispersity has been eliminated. Sheared, sonicated, or otherwise abused calf thymus (or other) DNAs should no longer be considered acceptable materials for physical studies. Many studies of bending and torsional fluctuations in DNA have been excluded from this discussion because the DNA samples used were not precisely defined. The torsional elastic constant of DNA has been fairly well established as approximately 3.0 x 10(-19) erg-cm, mainly through a combination of elegant theoretical and experimental studies of topoisomer distributions in circular DNA molecules. The other general approach to the determination of the torsional elastic constant, luminescence decay, is still burdened by the poor characterization of the DNA used in many of the experimental studies as well as by some continued theoretical uncertainties.(ABSTRACT TRUNCATED AT 400 WORDS)

We present a series of 26 patients, all >50 years of age, who are carriers of the fragile X premutation and are affected by a multisystem, progressive neurological disorder. The two main clinical features of this new syndrome are cerebellar ataxia and/or intention tremor, which were chosen as clinical inclusion criteria for this series. Other documented symptoms were short-term memory loss, executive function deficits, cognitive decline, parkinsonism, peripheral neuropathy, lower limb proximal muscle weakness, and autonomic dysfunction. Symmetrical regions of increased T2 signal intensity in the middle cerebellar peduncles and adjacent cerebellar white matter are thought to be highly sensitive for this neurologic condition, and their presence is the radiological inclusion criterion for this series. Molecular findings include elevated mRNA and low-normal or mildly decreased levels of fragile X mental retardation 1 protein. The clinical presentation of these patients, coupled with a specific lesion visible on magnetic resonance imaging and with neuropathological findings, affords a more complete delineation of this fragile X premutation-associated tremor/ataxia syndrome and distinguishes it from other movement disorders.

The authors report five elderly men with the fragile X premutation who had a progressive action tremor associated with executive function deficits and generalized brain atrophy. These individuals had elevated fragile X mental retardation 1 gene (FMR1) messenger RNA and normal or borderline levels of FMR1 protein. The authors propose that elevations of FMR1 messenger RNA may be causative for a neurodegenerative syndrome in a subgroup of elderly men with the FMR1 premutation.

Fragile X-associated tremor/ataxia syndrome (FXTAS) is an adult-onset neurodegenerative disorder that affects carriers, principally males, of premutation alleles (55-200 CGG repeats) of the fragile X mental retardation 1 (FMR1) gene. Clinical features of FXTAS include progressive intention tremor and gait ataxia, accompanied by characteristic white matter abnormalities on MRI. The neuropathological hallmark of FXTAS is an intranuclear inclusion, present in both neurons and astrocytes throughout the CNS. Prior to the current work, the nature of the associations between inclusion loads and molecular measures (e.g. CGG repeat) was not defined. Post-mortem brain and spinal cord tissue has been examined for gross and microscopic pathology in a series of 11 FXTAS cases (males, age 67-87 years at the time of death). Quantitative counts of inclusion numbers were performed in various brain regions in both neurons and astrocytes. Inclusion counts were compared with specific molecular (CGG repeat, FMR1 mRNA level) and clinical (age of onset, age of death) parameters. In the current series, the three most prominent neuropathological characteristics are (i) significant cerebral and cerebellar white matter disease, (ii) associated astrocytic pathology with dramatically enlarged inclusion-bearing astrocytes prominent in cerebral white matter and (iii) the presence of intranuclear inclusions in both brain and spinal cord. The pattern of white matter pathology is distinct from that associated with hypertensive vascular disease and other diseases of white matter. Spongiosis was present in the middle cerebellar peduncles in seven of the eight cases in which those tissues were available for study. There is inclusion formation in cranial nerve nucleus XII and in autonomic neurons of the spinal cord. The most striking finding is the highly significant association between the number of CGG repeats and the numbers of intranuclear inclusions in both neurons and astrocytes, indicating that the CGG repeat is a powerful predictor of neurological involvement in males, both clinically (age of death) and neuropathologically (number of inclusions).

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Fragile X-associated Tremor/Ataxia Syndrome (FXTAS) is a neurodegenerative disorder caused by expansion of 55-200 CGG repeats in the 5 0 -UTR of the FMR1 gene. FXTAS is characterized by action tremor, gait ataxia and impaired executive cognitive functioning. It has been proposed that FXTAS is caused by titration of RNA-binding proteins by the expanded CGG repeats. Sam68 is an RNAbinding protein involved in alternative splicing regulation and its ablation in mouse leads to motor coordination defects. Here, we report that mRNAs containing expanded CGG repeats form large and dynamic intranuclear RNA aggregates that recruit several RNA-binding proteins sequentially, first Sam68, then hnRNP-G and MBNL1. Importantly, Sam68 is sequestered by expanded CGG repeats and thereby loses its splicing-regulatory function. Consequently, Sam68-responsive splicing is altered in FXTAS patients. Finally, we found that regulation of Sam68 tyrosine phosphorylation modulates its localization within CGG aggregates and that tautomycin prevents both Sam68 and CGG RNA aggregate formation. Overall, these data support an RNA gain-of-function mechanism for FXTAS neuropathology, and suggest possible target routes for treatment options.

Fragile X syndrome (FXS) is the leading inherited form of intellectual disability and autism spectrum disorder, and patients can present with severe behavioural alterations, including hyperactivity, impulsivity and anxiety, in addition to poor language development and seizures. FXS is a trinucleotide repeat disorder, in which >200 repeats of the CGG motif in FMR1 leads to silencing of the gene and the consequent loss of its product, fragile X mental retardation 1 protein (FMRP). FMRP has a central role in gene expression and regulates the translation of potentially hundreds of mRNAs, many of which are involved in the development and maintenance of neuronal synaptic connections. Indeed, disturbances in neuroplasticity is a key finding in FXS animal models, and an imbalance in inhibitory and excitatory neuronal circuits is believed to underlie many of the clinical manifestations of this disorder. Our knowledge of the proteins that are regulated by FMRP is rapidly growing, and this has led to the identification of multiple targets for therapeutic intervention, some of which have already moved into clinical trials or clinical practice.