FRAXE-associated mental retardation protein (FMR2) is an RNA-binding protein with high affinity for G-quartet RNA forming structure

CNRS UMR 6097-Institut de Pharmacologie Moléculaire et Cellulaire, Valbonne, France.
Nucleic Acids Research (Impact Factor: 9.11). 02/2009; 37(4):1269-79. DOI: 10.1093/nar/gkn1058
Source: PubMed


FRAXE is a form of mild to moderate mental retardation due to the silencing of the FMR2 gene. The cellular function of FMR2 protein is presently unknown. By analogy with its homologue AF4, FMR2 was supposed to have a role in transcriptional regulation, but robust evidences supporting this hypothesis are lacking. We observed that FMR2 co-localizes with the splicing factor SC35 in nuclear speckles, the nuclear regions where splicing factors are concentrated, assembled and modified. Similarly to what was reported for splicing factors, blocking splicing or transcription leads to the accumulation of FMR2 in enlarged, rounded speckles. FMR2 is also localized in the nucleolus when splicing is blocked. We show here that FMR2 is able to specifically bind the G-quartet-forming RNA structure with high affinity. Remarkably, in vivo, in the presence of FMR2, the ESE action of the G-quartet situated in mRNA of an alternatively spliced exon of a minigene or of the putative target FMR1 appears reduced. Interestingly, FMR1 is silenced in the fragile X syndrome, another form of mental retardation. All together, our findings strongly suggest that FMR2 is an RNA-binding protein, which might be involved in alternative splicing regulation through an interaction with G-quartet RNA structure.

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Available from: Jozef Gecz
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    • "Besides the X-chromosome fragile site A, fragile site E (locus in Xq28) is also associated with intellectual disability [FRAXE; MIM #309548]. This disorder is mainly a non-syndromic form of X-linked intellectual disability and is the result of silencing of the AFF2 gene, as a consequence of an upstream CCG expansion [8]. In the normal population, the number of CCG repeats varies between 6 and 35, while it is increased to more than 200 hyper-methylated triplets in FRAXE/AFF2 intellectually disabled patients. "
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    ABSTRACT: X-linked intellectual disability is a common cause of inherited cognitive deficit affecting mostly males. There are several genetic causes implicated in this condition, which has hampered the establishment of an accurate diagnosis. We developed a multiplex-PCR assay for the mutational hotspot regions of the FMR1, AFF2 and ARX genes. The multiplex-PCR was validated in a cohort of 100 males selected to include known alleles for the FMR1 repetitive region: five full mutations (250--650 CGGs), ten premutations (70--165 CGGs) and eighty-five in the normal range (19--42 CGGs). Sequencing or Southern blotting was used to confirm the results, depending on the allele class. In this cohort, with the exception of one sample showing an AFF2 intermediate-sized allele, all other samples were normal (8--34 CCGs). No ARX variant was found besides the c.429_452dup. The validated assay was applied to 5000 samples (64.4% males and 35.6% females). The normal-allelic range of both FMR1 and AFF2 genes as well as the nature of ARX variants identified was similar in both genders. The rate of homozygosity observed in female samples, 27.5% for FMR1 and 17.8% for AFF2 alleles, is comparable to that published by others. Two FMR1 premutations were identified, in a male (58 CGGs) and a female case [(CGG)47/(CGG)61], as well as several FMR1 or AFF2 intermediate-sized alleles. One AFF2 premutation (68 CCGs) and two putative full expansions were picked up in male subjects, which seems relevant considering the rarity of reported AFF2 mutations found in the absence of a family history. We developed a robust multiplex-PCR that can be used to screen the mutational hotspot regions of FMR1, AFF2 and ARX genes. Moreover, this strategy led to the identification of variants in all three genes, representing not only an improvement in allele-sizing but also in achieving a differential diagnosis. Although the distinction between females who are truly homozygous and those with a second pre- or full mutation sized allele, as well as a definitive diagnosis, requires a specific downstream technique, the use of this multiplex-PCR for initial screening is a cost-effective approach which widens the scope of detection.
    Full-text · Article · Aug 2013 · BMC Medical Genetics
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    • "The interaction between FMRP and the G-quadruplex RNA is mediated by the RGG box domain, while no RNA-binding domain was defined in the sequence of FMR2P (Bensaid et al., 2009). Analyzing the FMR2P sequence by the software "
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    ABSTRACT: Intellectual disability (ID) is the most frequent cause of serious handicap in children and young adults and interests 2-3% of worldwide population, representing a serious problem from the medical, social, and economic points of view. The causes are very heterogeneous. Genes involved in ID have various functions altering different pathways important in neuronal function. Regulation of mRNA metabolism is particularly important in neurons for synaptic structure and function. Here, we review ID due to alteration of mRNA metabolism. Functional absence of some RNA-binding proteins--namely, FMRP, FMR2P, PQBP1, UFP3B, VCX-A--causes different forms of ID. These proteins are involved in different steps of RNA metabolism and, even if a detailed analysis of their RNA targets has been performed so far only for FMRP, it appears clear that they modulate some aspects (translation, stability, transport, and sublocalization) of a subset of RNAs coding for proteins, whose function must be relevant for neurons. Two other proteins, DYRK1A and CDKL5, involved in Down syndrome and Rett syndrome, respectively, have been shown to have an impact on splicing efficiency of specific mRNAs. Both proteins are kinases and their effect is indirect. Interestingly, both are localized in nuclear speckles, the nuclear domains where splicing factors are assembled, stocked, and recycled and influence their biogenesis and/or their organization.
    Full-text · Article · Jan 2012 · Progress in brain research
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    • "However, little is known about the mechanisms by which G4-RNA formation is regulated in the cells. Indeed, only a few proteins have been reported so far to interact with G4-RNAs in vitro (62–66) and, apart from FMRP and FMR2P, experimental data on their biological activity subsequent to their interaction with G4-RNAs is scarce. Of these G4-RNA binding proteins, RHAU is the only protein that exhibits robust in vitro ATPase-dependent G4-RNA resolving activity, in addition to a high affinity and specificity for its target RNAs (42). "
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    ABSTRACT: Guanine-quadruplexes (G4) consist of non-canonical four-stranded helical arrangements of guanine-rich nucleic acid sequences. The bulky and thermodynamically stable features of G4 structures have been shown in many respects to affect normal nucleic acid metabolism. In vivo conversion of G4 structures to single-stranded nucleic acid requires specialized proteins with G4 destabilizing/unwinding activity. RHAU is a human DEAH-box RNA helicase that exhibits G4-RNA binding and resolving activity. In this study, we employed RIP-chip analysis to identify en masse RNAs associated with RHAU in vivo. Approximately 100 RNAs were found to be associated with RHAU and bioinformatics analysis revealed that the majority contained potential G4-forming sequences. Among the most abundant RNAs selectively enriched with RHAU, we identified the human telomerase RNA template TERC as a true target of RHAU. Remarkably, binding of RHAU to TERC depended on the presence of a stable G4 structure in the 5′-region of TERC, both in vivo and in vitro. RHAU was further found to associate with the telomerase holoenzyme via the 5′-region of TERC. Collectively, these results provide the first evidence that intramolecular G4-RNAs serve as physiologically relevant targets for RHAU. Furthermore, our results suggest the existence of alternatively folded forms of TERC in the fully assembled telomerase holoenyzme.
    Full-text · Article · Aug 2011 · Nucleic Acids Research
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