Published online 14 December 2006 Nucleic Acids Research, 2007, Vol. 35, No. 1 299–306 doi:10.1093/nar/gkl1021 Fragile X related protein 1 isoforms differentially modulate the affinity of fragile X mental retardation protein for G-quartet RNA structure Elias Bechara, Laetitia Davidovic1, Mireille Melko, Mounia Bensaid, Sandra Tremblay 1 , Josiane Grosgeorge, Edouard W. Khandjian 1 , Enzo Lalli2 and Barbara Bardoni* CNRS UMR 6543, Faculte´ de Me´ decine–Universite´ de Nice Sophia-Antipolis, 28 Avenue De Valombrose, 06107, Nice, France, 1Unite´ de Recherche en Ge´ ne´ tique Humaine et Mole´ culaire, Centre de Recherche Hoˆ pital Saint-Franc¸ois d’Assise, le CHUQ, Que´ bec, Canada G1L 3L5, De´ partement de Biologie Me´ dicale, Faculte´ de Me´ decine, Universite´ Laval, Que´ bec, Canada and 2Institut de Pharmacologie Mole´ culaire et Cellulaire, CNRS UMR6097, Universite´ de Nice Sophia-Antipolis, 660 Route des Lucioles, 06560 Valbonne Sophia-Antipolis, France Received September 6, 2006; Revised and Accepted November 13, 2006 ABSTRACT Fragile X syndrome, the most frequent form of inherited mental retardation, is due to the absence of expression of the Fragile X Mental Retardation Protein (FMRP), an RNA binding protein with high specificity for G-quartet RNA structure. FMRP is involved in several steps of mRNA metabolism: nucleocytoplasmic trafficking, translational control and transport along dendrites in neurons. Fragile X Related Protein 1 (FXR1P), a homologue and inter- actor of FMRP, has been postulated to have a function similar to FMRP, leading to the hypothesis that it can compensate for the absence of FMRP in Fragile X patients. Here we analyze the ability of three isoforms of FXR1P, expressed in different tissues, to bind G-quartet RNA structure specific- ally. Only the longest FXR1P isoform was found to be able to bind specifically the G-quartet RNA, albeit with a lower affinity as compared to FMRP, whereas the other two isoforms negatively regulate the affinity of FMRP for G-quartet RNA. This result is important to decipher the molecular basis of fragile X syndrome, through the understanding of FMRP action in the context of its multimolecular complex in different tissues. In addition, we show that the action of FXR1P is synergistic rather than compen- satory for FMRP function. INTRODUCTION Fragile X related genes are members of a small gene family whose founding member is the Fragile X Mental Retardation 1 gene (FMR1). Inactivation of FMR1 causes Fragile X syn- drome, the most common cause of inherited mental retarda- tion (1,2). The other members of this family, FXR1 and FXR2, are autosomal and have not been associated so far with any human disease (2–4). Animal models have been generated for Fmr1 deﬁciency, recapitulating the phenotype of Fragile X syndrome (5,6). Fxr2 mice are viable and show some behavioral phenotypes, such as hyperactivity, similar to those observed in Fmr1 knockout mice (7). Fxr1 mice die shortly after birth most likely because of heart and/or respiratory failure due to alterations in muscle development (8). In Xenopus, complete or partial inactivation of xFxr1 expression has dramatic muscle-speciﬁc effects (9). In vertebrates, members of the FXR protein family are structurally very similar and share a high degree of sequence homology in clustered regions corresponding to functional domains (2–4). Like FMRP, FXR1P contains several RNA binding domains: two KH domains and one RGG box. It also contains a nuclear localization signal (NLS), a nuclear export signal (NES) and a protein–protein interaction domain (2,10). They also share the same gene structure, derived from their common ancestor in Drosophila melanogaster (11). FXR proteins are able to bind RNA (3,4), but binding speci- ﬁcity has been studied in detail only for FMRP. Indeed, even if a few hundreds of different RNAs have been proposed to be putative targets of FMRP, only two structures are speciﬁcally bound by this protein, the G-quartet and the kissing complex (12–14) and one sequence, a poly(U) stretch (15). FXR1P has been reported to bind AU rich element (ARE) and, through the interaction with this element, to regulate the expression of the proinﬂammatory cytokine tumor necrosis factor (TNFa) in macrophages (16). In the cytoplasm the three FXR proteins are associated with polyribosomes (17), while they share only two interacting proteins, CYFIP2 and MSP58, with FMRP (2,18,19). The FXR1 primary transcript is alternatively *To whom correspondence should be addressed. Tel: +0033 493377786; Fax: +0033 493377033; Email: bardoni@unice.fr Present address: Laetitia Davidovic, CNRS UMR 6543, Faculte´ de Me´decine–Universite´ de Nice Sophia-Antipolis, 28 Avenue De Valombrose, 06107, Nice, France 2006 The Author(s). This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/ by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.