Raver1, a dual compartment protein, is a ligand for PTB/hnRNPI and microfilament attachment proteins

Abstract

By screening a yeast two-hybrid library with COOH-terminal fragments of vinculin/metavinculin as the bait, we identified a new protein termed raver1. Raver1 is an 80-kD multidomain protein and widely expressed but to varying amounts in different cell lines. In situ and in vitro, raver1 forms complexes with the microfilament-associated proteins vinculin, metavinculin, and α-actinin and colocalizes with vinculin/metavinculin and α-actinin at microfilament attachment sites, such as cell–cell and cell matrix contacts of epithelial cells and fibroblasts, respectively, and in costameres of skeletal muscle. The NH2-terminal part of raver1 contains three RNA recognition motifs with homology to members of the heterogeneous nuclear RNP (hnRNP) family. Raver1 colocalizes with polypyrimidine tract binding protein (PTB)/hnRNPI, a protein involved in RNA splicing of microfilament proteins, in the perinucleolar compartment and forms complexes with PTB/hnRNPI. Hence, raver1 is a dual compartment protein, which is consistent with the presence of nuclear location signal and nuclear export sequence motifs in its sequence. During muscle differentiation, raver1 migrates from the nucleus to the costamere. We propose that raver1 may coordinate RNA processing and targeting as required for microfilament anchoring in specific adhesion sites.

Structure-function relationships of the polypyrimidine tract binding protein

“…Hence, mechanisms that release PTB are important for tissue-specific regulation of splicing. Another facet of splicing regulation by PTB is the existence of co-repressors like Raver1 [38,39]. Raver1 recruitment is essential for effective repressor function of PTB in certain pre-mRNAs, for example for the splicing repression of a-tropomyosin exon 3, while in other systems PTB alone is sufficient.…”

Internal ribosome entry segment-mediated translation during apoptosis: the role of IRES-trans-acting factors

“…In the presence of unr, this structure is altered such that some of the stem-loops are opened, and this allows nPTB to bind at two sites (P1 and P2). The interactions of these proteins thus allow the Apaf-1 IRES RNA to attain the correct structural conformation for ribosome recruitment by creating a single-stranded region, which allows the ribosome-landing site to become accessible metvinculin, and also interacts with PTB to regulate splicing; 57,58 hnRNP-L, a PTB homologue that was found to bind PTB in a yeast two-hybrid screen; 59 PSF (PTB-associated splicing factor) and p54 nrb /NonO are related multifunctional nuclear factors that contain both RNA-binding and DNAbinding functions. 60 Other ITAFs that help to regulate IRES-mediated expression of proteins that function during apoptosis include PCBP2 and hnRNPK.…”

Raver1 is an integral component of muscle contractile elements

“…1A), a ubiquitously expressed 80 kDa protein that contains three RNA-recognition motifs (RRMs) with a high sequence homology to members of heterogeneous ribonucleoprotein family (Huttelmaier et al 2001;Jockusch et al 2003). In most cell types, raver1 is found in the nucleus, but displays nucleocytoplasmic shuttling presumably mediated by two nuclear localization signals at the N-and C-termini and a putative export sequence located within the central leucine-rich region.…”

“…Direct binding of raver1 to PTB has been demonstrated in vitro and both proteins colocalize in the perinucleolar compartment (PNC), an RNA-dependent structure that is characterized by the accumulation of PTB (Ghetti et al 1992). Furthermore, raver1-PTB complexes can be immunoprecipitated from vertebrate cells (Huttelmaier et al 2001). While direct RNA binding of raver1 has yet to be demonstrated, the protein may assist in PTB-mediated splicing events in smooth muscle (Gromak et al 2003;Spellman et al 2005).…”

“…While direct RNA binding of raver1 has yet to be demonstrated, the protein may assist in PTB-mediated splicing events in smooth muscle (Gromak et al 2003;Spellman et al 2005). Apart from its association with PTB in the nucleus, raver1 interacts with the microfilament-associated proteins vinculin, its muscle-specific splice variant metavinculin and alpha-actinin (Huttelmaier et al 2001). In fact, raver1 was originally identified through its interaction with metavinculin in a yeast two hybrid screen.…”