The X chromosome and fragile X mental retardation

Fragile X syndrome represents the most common inherited cause of mental retardation. It is caused by a stretch of CGG repeats within the fragile X gene, which increases in length as it is transmitted from generation to generation. Once the repeat exceeds a threshold length, no protein is produced, resulting in the fragile X phenotype. Both X chromosome inactivation and inactivation of the FMR1 gene are the result of methylation. X inactivation occurs earlier than inactivation of the FMR1 gene. The instability to a full mutation is dependent on the sex of the transmitting parent and occurs only from mother to child. For most X-chromosomal diseases, female carriers do not express the phenotype. A clear exception is fragile X syndrome. It is clear that more than 50% of the neurons have to express the protein to ensure a normal phenotype in females. This means that a normal phenotype in female carriers of a full mutation is accompanied by a distortion of the normal distribution of X inactivation.     

Fragile X mental retardation syndrome: structure of the KH1-KH2 domains of fragile X mental retardation protein

Fragile X syndrome is the most common form of inherited mental retardation in humans, with an estimated prevalence of about 1 in 4000 males. Although several observations indicate that the absence of functional Fragile X Mental Retardation Protein (FMRP) is the underlying basis of Fragile X syndrome, the structure and function of FMRP are currently unknown. Here, we present an X-ray crystal structure of the tandem KH domains of human FMRP, which reveals the relative orientation of the KH1 and KH2 domains and the location of residue Ile304, whose mutation to Asn is associated with a particularly severe incidence of Fragile X syndrome. We show that the Ile304Asn mutation both perturbs the structure and destabilizes the protein.     

RNA and microRNAs in fragile X mental retardation

Fragile X syndrome is caused by the loss of an RNA-binding protein called FMRP (for fragile X mental retardation protein). FMRP seems to influence synaptic plasticity through its role in mRNA transport and translational regulation. Recent advances include the identification of mRNA ligands, FMRP-mediated mRNA transport and the neuronal consequence of FMRP deficiency. FMRP was also recently linked to the microRNA pathway. These advances provide mechanistic insight into this disorder, and into learning and memory in general.     

Alternative splicing modulates protein arginine methyltransferase-dependent methylation of fragile X syndrome mental retardation protein

The fragile X mental retardation protein (FMRP) is an RNA binding protein that is methylated by an endogenous methyltransferase in rabbit reticulocyte lysates. We mapped the region of methylation to the C-terminal arginine-glycine-rich residues encoded by FMR1 exon 15. We additionally demonstrated that mutation of R(544) to K reduced the endogenous methylation by more than 80%, while a comparable mutant R(546)-K reduced the endogenous methylation by 20%. These mutations had no effect on the subcellular distribution of FMRP, recapitulating previous results using the methyltransferase inhibitor adenosine-2',3'-dialdehyde. Using purified recombinant protein arginine methyltransferases (PRMTs), we showed that the C-terminal domain could be methylated by PRMT1, PRMT3, and PRMT4 in vitro and that both the R(544)-K mutant and the R(546)-K mutant were refractory toward these enzymes. We also report that truncating the N-terminal 12 residues encoded by FMR1 exon 15, which occurs naturally via alternative splicing, had no effect on FMRP methylation, demonstrating conclusively that phosphorylation of serine residue 500 (S(500)), one of the 12 residues, was not required for methylation. Nevertheless, truncating 13 additional amino acids, as occurs in the smallest alternatively spliced variant of FMR1 exon 15, reduced methylation by more than 85%. This suggests that differential expression and methylation of the FMRP exon 15 variants may be an important means of regulating target mRNA translation, which is consonant with recently demonstrated functional effects mediated by inhibiting FMRP methylation in cultured cells.     

Therapeutic implications of the mGluR theory of fragile X mental retardation

Evidence is reviewed that the consequences of group 1 metabotropic glutamate receptor (Gp1 mGluR) activation are exaggerated in the absence of the fragile X mental retardation protein, likely reflecting altered dendritic protein synthesis. Abnormal mGluR signaling could be responsible for remarkably diverse psychiatric and neurological symptoms in fragile X syndrome, including delayed cognitive development, seizures, anxiety, movement disorders and obesity.     

Balanced X-autosomal translocation and mental retardation. Mapping mental retardation linked to X (excluding fragile X)

From personal observations and reported cases of translocation X-Autosome, a study of the breakpoint showed that Xp11 is more frequently associated to mental retardation. This finding is in agreement with linkage analysis in families with X-linked mental retardation non X-fra.     

X-linked mental retardation with the fragile X. A study of 15 families

Summary The coinical and cytogenetic features of 15 families with mental retardation linked to the fragile site on the X chromosome are presented.The 15 propositi were all prepubertal, and one was a girl. Although the clinical picture varied in severity, it was sufficiently constant to suggest the diagnosis from the facial features and the encephalopathy with language retardation and disturbed behavior. Macroorchidism was not seen before puberty.The fragile X chromosome was found in seven of the nine mothers studied and in two mildly retarded sisters and has also been demonstrated in fibroblasts in eleven subjects with the abnormality.Supported by grants from INSERM (ATP 79-110)     

Dissociation between mental retardation and fragile site expression in a family with fragile X-linked mental retardation

Summary We report an extended family in which two brothers with a fragile X chromosome are mentally retarded while a third brother with the fragile site is both phenotypically and mentally normal. The study of six probes detecting restriction fragment length polymorphisms on either sides of the fragile site Xq27 confirmed that the fragile X regions inherited by these three brothers were identical from DXS 102 to the telomere. These data highlight the heterogeneity of the fragile X syndrome, which is discussed in the framework of the different hypotheses previously proposed.     

The fragile X mental retardation syndrome protein interacts with novel homologs FXR1 and FXR2.

Fragile X Mental Retardation Syndrome is the most common form of hereditary mental retardation, and is caused by defects in the FMR1 gene. FMR1 is an RNA-binding protein and the syndrome results from lack of expression of FMR1 or expression of a mutant protein that is impaired in RNA binding. The specific function of FMR1 is not known. As a step towards understanding the function of FMR1 we searched for proteins that interact with it in vivo. We have cloned and sequenced a protein that interacts tightly with FMR1 in vivo and in vitro. This novel protein, FXR2, is very similar to FMR1 (60% identity). FXR2 encodes a 74 kDa protein which, like FMR1, contains two KH domains, has the capacity to bind RNA and is localized to the cytoplasm. The FXR2 gene is located on human chromosome 17 at 17p13.1. In addition, FMR1 and FXR2 interact tightly with the recently described autosomal homolog FXR1. Each of these three proteins is capable of forming heteromers with the others, and each can also form homomers. FXR1 and FXR2 are thus likely to play important roles in the function of FMR1 and in the pathogenesis of the Fragile X Mental Retardation Syndrome.     

Molecular diagnosis of the fragile X and FRAXE syndromes in patients with mental retardation of unknown cause in Mexico

The fragile X syndrome (Fra-X) is the most common cause of inherited mental retardation with X-linked semi-dominant inheritance. The prevalence of Fra-X in the Mexican population is unknown. The aim of this population screening study was to determine if Fra-X or FRAXE mutations are the cause of a number of cases of mental retardation in a sample of Mexican children with mental retardation of unknown cause (MRUC) and to stress the importance of performing molecular analysis of the FMR-1 gene in all patients with MRUC. We report here the direct analysis of CGG and GCC repeats within the FMR-1 and FMR-2 genes, respectively, in 62 unrelated patients with MRUC. Two male index cases had the CGG expansion, although they did not express the Xq27.3 fragile site cytogenetically. Fra-X diagnosis was highly suspected on a clinical basis in one of the patients, but not in the other. Both mothers were found to be premutation carriers. The molecular studies of FMR-1 showed that the proportion of MRUC patients with Fra-X is 3.2%. This frequency was not significantly different to that reported in most populations. As reported in other series, no patients with FRAXE were found in our sample. Our findings confirm that the molecular analysis of the FMR-1 gene is necessary in MRUC patients to achieve unequivocal diagnosis of fragile X syndrome, carrier premutation detection and for accurate genetic counseling.     

Spermatogenesis in two patients with the fragile X syndrome

Summary Light and electron microscopic studies on testicular biopsies were carried out in two men, 40 and 44 year old, with the fra(X) form of mental retardation and macroorchidism. Distinct interstitial edema, an increased amount of lysosomal inclusions in Sertoli cells, and disturbance of spermatid differentiation were found in both probands. Additionally, some extent of tubular atrophy was demonstrated in one patient. The impairment of spermatogenesis is discussed with respect to pressure effects on the germinal epithelium due to the edema.     

Spermatogenesis in two patients with the fragile X syndrome

Summary Chromosomes at first meiosis from two males with the fra(X) form of mental retardation were studied using pachytene surface spreads and air-dried preparations. The pachytene sex bivalents showed no discontinuation of the synaptonemal complex in the terminal part of Xq corresponding to band Xq27–28 of the mitotic chromosomes. In both cases the frequency of a secondary association of Xq and Yq appeared to be increased compared with controls. The pairing behavior of autosomal bivalents in pachytene and the frequency and distribution of chiasmata in diakinesis were normal. The impairment of spermatogenesis found in these males may not be caused by a meiotic disorder, but could be related to peritubular or intratubular pressure effects on germ cells.     

High incidence of mental retardation in Turner syndrome patients with ring chromosome X formation

In this report we describe and comment the high incidence of mental subnormality in a series of 21 Turner syndrome patients with ring chromosome X, diagnosed in Leuven in the period 1965-1989. In 7 of the 21 (one third) a varying degree of mental retardation, from borderline intelligence to severe mental retardation was found. In 4 of them (18.5%) mental retardation was moderate to severe.     

Drosophila fragile X mental retardation protein developmentally regulates activity-dependent axon pruning

Fragile X Syndrome (FraX) is a broad-spectrum neurological disorder with symptoms ranging from hyperexcitability to mental retardation and autism. Loss of the fragile X mental retardation 1 (fmr1) gene product, the mRNA-binding translational regulator FMRP, causes structural over-elaboration of dendritic and axonal processes, as well as functional alterations in synaptic plasticity at maturity. It is unclear, however, whether FraX is primarily a disease of development, a disease of plasticity or both: a distinction that is vital for engineering intervention strategies. To address this crucial issue, we have used the Drosophila FraX model to investigate the developmental function of Drosophila FMRP (dFMRP). dFMRP expression and regulation of chickadee/profilin coincides with a transient window of late brain development. During this time, dFMRP is positively regulated by sensory input activity, and is required to limit axon growth and for efficient activity-dependent pruning of axon branches in the Mushroom Body learning/memory center. These results demonstrate that dFMRP has a primary role in activity-dependent neural circuit refinement during late brain development.     

Biology of the fragile X mental retardation protein, an RNA-binding protein.

The fragile X syndrome, an X-linked disease, is the most frequent cause of inherited mental retardation. The syndrome results from the absence of expression of the FMR1 gene (fragile mental retardation 1) owing to the expansion of a CGG trinucleotide repeat located in the 5' untranslated region of the gene and the subsequent methylation of its CpG island. The FMR1 gene product (FMRP) is a cytoplasmic protein that contains two KH domains and one RGG box, characteristics of RNA-binding proteins. FMRP is associated with mRNP complexes containing poly(A)+mRNA within actively translating polyribosomes and contains nuclear localization and export signals making it a putative transporter (chaperone) of mRNA from the nucleus to the cytoplasm. FMRP is the archetype of a novel family of cytoplasmic RNA-binding proteins that includes FXR1P and FXR2P. Both of these proteins are very similar in overall structure to FMRP and are also associated with cytoplasmic mRNPs. Members of the FMR family are widely expressed in mouse and human tissues, albeit at various levels, and seem to play a subtle choreography of expression. FMRP is most abundant in neurons and is absent in muscle. FXR1P is strongly expressed in muscle and low levels are detected in neurons. The complex expression patterns of the FMR1 gene family in different cells and tissues suggest that independent, however similar, functions for each of the three FMR-related proteins might be expected in the selection and metabolism of tissue-specific classes of mRNA. The molecular mechanisms altered in cells lacking FMRP still remain to be elucidated as well as the putative role(s) of FXR1P and FXR2P as compensatory molecules.     

Temporal requirements of the fragile X mental retardation protein in the regulation of synaptic structure

Fragile X syndrome (FraX), caused by the loss-of-function of one gene (FMR1), is the most common inherited form of both mental retardation and autism spectrum disorders. The FMR1 product (FMRP) is an mRNA-binding translation regulator that mediates activity-dependent control of synaptic structure and function. To develop any FraX intervention strategy, it is essential to define when and where FMRP loss causes the manifestation of synaptic defects, and whether the reintroduction of FMRP can restore normal synapse properties. In the Drosophila FraX model, dFMRP loss causes neuromuscular junction (NMJ) synapse over-elaboration (overgrowth, overbranching, excess synaptic boutons), accumulation of development-arrested satellite boutons, and altered neurotransmission. We used the Gene-Switch method to conditionally drive dFMRP expression to define the spatiotemporal requirements in synaptic mechanisms. Constitutive induction of targeted neuronal dFMRP at wild-type levels rescues all synaptic architectural defects in Drosophila Fmr1 (dfmr1)-null mutants, demonstrating a presynaptic requirement for synapse structuring. By contrast, presynaptic dFMRP expression does not ameliorate functional neurotransmission defects, indicating a postsynaptic dFMRP requirement. Strikingly, targeted early induction of dFMRP effects nearly complete rescue of synaptic structure defects, showing a primarily early-development role. In addition, acute dFMRP expression at maturity partially alleviates dfmr1-null defects, although rescue is not as complete as either early or constitutive dFMRP expression, showing a modest capacity for late-stage structural plasticity. We conclude that dFMRP predominantly acts early in synaptogenesis to modulate architecture, but that late dFMRP introduction at maturity can weakly compensate for early absence of dFMRP function.     

The fragile X mental retardation protein interacts with U-rich RNAs in a yeast three-hybrid system

We recently identified several ESTs that bind to the fragile X mental retardation protein (FMRP) in vitro. To determine whether they interacted in vivo we performed three-hybrid screens in a Saccharomyces cerevisiae histidine auxotroph. We demonstrate that two of the ESTs support growth on histidine and transduce beta-galactosidase activity when co-expressed with FMRP under selective growth conditions. In contrast, the iron response element (IRE) RNA does not. Likewise, the ESTs do not support growth or transduce beta-galactosidase activity when co-expressed with the iron response element binding protein (IRP). Each EST is relatively small and has 40% identity with a sequence in FMR1 mRNA harboring FMRP binding determinants. Interestingly, while neither the ESTs contain a G-quartet structural motif they do contain U-rich sequences that are found in mRNA with demonstrated in vitro binding and in vivo association with FMRP. This indicates that U-rich elements comprise another motif recognized by FMRP.