A Nuclear SH3 Domain-binding Protein That Colocalizes with mRNA Splicing Factors and Intermediate Filament-containing Perinuclear Networks*

A Nuclear SH3 Domain-binding Protein That Colocalizes with mRNA Splicing Factors and Intermediate Filament-containing Perinuclear Networks*

Journal of Biological Chemistry, May 2001

11375989

Graham Craggs, Peter M. Finan, Durward Lawson, Jonathan Wingfield, Timothy Perera, Smita Gadher, Nicholas F. Totty, Stuart Kellie

A protein (SNP70) has been isolated that binds to the Src homology domain 3 of p47(phox), p85alpha, and c-src. Cloning and sequencing of the polypeptide revealed it to be a 70-kDa protein that has a number of potential domains, including Src homology 3 binding motifs and several nuclear localization signals. Immunofluorescence using anti-peptide antibodies revealed SNP70 to be primarily concentrated in the nucleus but excluded from nucleoli, in interphase cells. However, it was distributed throughout the cytoplasm in dividing cells. Extraction and subfractionation experiments indicated that SNP70 did not bind directly to DNA but did bind to poly(G)-rich oligonucleotides and was resistant to extraction with non-ionic detergents but was solubilized by treatment with RNase, high salt, or ammonium sulfate. Double-immunofluorescence experiments showed that SNP70 co-localized with two pre-mRNA splicing factors SC35 and U2B" within the nucleus. A population of SNP70 was found outside the nucleus, and double-immunofluorescence and immunoelectron microscopy demonstrated that it associated with vimentin-containing intermediate filaments, particularly those surrounding the nucleus. The data suggest that SNP70 associates with nuclear or perinuclear filaments and may play a role in the regulation of pre-mRNA processing.