Fragile X Mental Retardation protein determinants required for its association with polyribosomal mRNPs.
Fragile X Mental Retardation protein (FMRP) is an RNA-binding protein that contains multiple domains with apparently differential affinity to mRNA and to the ribonucleotide homopolymer poly(G). Attempts have been made to map the RNA-binding sites along the protein sequence with a view to determining which of the KH1, KH2 and RGG domains are required to recognize and bind to RNA. While these studies have greatly contributed to the delineation of domains that bind homopolymers or mRNA in vitro, little is known concerning their implications in FMRP function(s) in vivo. To address this question, we have prepared a series of FMRP versions, in which each known in vitro functional domain has been individually deleted, leaving the rest of the protein intact. Constructs with deletions in the protein-protein interaction and RNA-binding as well as in the phosphorylation domains were expressed in STEK-KO cells lacking FMRP and their recruitment into polyribosomal mRNPs and their intra-cellular localization were determined. Our results indicate that the KH RNA-binding domains and the Protein-Protein Interacting domain are essential for FMRP to associate with polyribosomal mRNPs, while the RGG box and the phosphorylated domains are dispensable.
Mazroui
Rachid
R
Unité de Recherche en Génétique Humaine et Moléculaire, Centre de Recherche Hôpital St-François d'Assise, Le CHUQ, Québec, Qc G1L 3L5, Canada.
Huot
Marc-Etienne
ME
Tremblay
Sandra
S
Boilard
Nathalie
N
Labelle
Yves
Y
Khandjian
Edouard W
EW
D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D051860:Fragile X Messenger Ribonucleoprotein 1; D005822:Genetic Vectors; D009419:Nerve Tissue Proteins / Q000235:genetics* / Q000378:metabolism; D010766:Phosphorylation; D011068:Poly G / Q000378:metabolism; D011485:Protein Binding; D025941:Protein Interaction Mapping; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012261:Ribonucleoproteins / Q000235:genetics / Q000378:metabolism*
D051860:Fragile X Messenger Ribonucleoprotein 1; D009419:Nerve Tissue Proteins; D011068:Poly G; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D012261:Ribonucleoproteins; C026452:messenger ribonucleoprotein