The fragile X-related proteins FXR1P and FXR2P contain a functional nucleolar-targeting signal equivalent to the HIV-1 regulatory proteins

Fragile X syndrome is caused by the absence of the INTRODUCTION fragile X m ental-retardation protein (FMRP). FMRP and the fragile X-related proteins 1 and 2 (FXR1P and FXR2P) form a gene family with functional similarities, s uch as RNA binding, polyribosomal association and nucleocytoplasmic shuttli ng. In a previous study, we found that FMRP and FXR1P shuttle between cytop lasm and nucleoplasm, while FXR2P shuttles between cytoplasm and nucleolus. The nuclear and nucleolar-targeting properties of these proteins were inve stigated further. Here, we show that FXR2P contains in its C-terminal part, a stretch of basic amino acids 'RPQRRNRSRRRRFR' that resemble the nucleola r-targeting signal (NoS) of the viral protein Rev. This particular sequence is also present within exon 15 of the FXR1 gene. This exon undergoes alter native splicing and is therefore only present in some of the FXR1P isoforms , We investigated the intracellular distribution of various FXR1P isoforms with (iso-e and iso-f) and without (iso-d) the potential NoS in transfected COS cells treated with the nuclear export inhibitor teptomycin-B. Both iso -e and iso-f showed a nucleolar localization, as observed for FXR2P; iso-d was detected in the nucleoplasm outside the nucleoli, Further, when a label led le-residue synthetic peptide corresponding to the NoS of FXR1P was adde d to human fibroblast cultures a clear nucleolar signal was observed. Based on these data we argue that the intranuclear distribution of FXR2P and FXR 1P isoforms is very likely to be mediated by a similar NoS localized in the ir C-terminal region. This domain is absent in some FXR1P isoforms as well as in all FMRP isoforms, suggesting functional differences for this family of proteins, possibly related to RNA metabolism in different tissues.