Nuclear Translocation of Insulin Receptor Substrate-1 by Oncogenes And Igf-I

Abstract

The insulin receptor substrate-1 (IRS-1) is one of the major substrates of both the insulin and IGF-I receptors and is generally localized in the cytosol/membrane fraction of the cell. We show here that a substantial fraction of IRS-1 is translocated to the nucleus in mouse embryo fibroblasts (MEF) expressing the simian virus 40 T antigen. Nuclear translocation of IRS-1 occurs also in MEF stimulated with IGF-I or in MEF expressing the oncogene v-src. Nuclear translocation of IRS-1 can be demonstrated by confocal microscopy, immunohistochemistry, or subcellular fractionation. An antibody to IRS-1 immunoprecipitates from nuclear fractions (but not from cytosolic fractions) the upstream binding factor, which is a key regulator of RNA polymerase I activity and ribosomal RNA (rRNA) synthesis. In agreement with this finding, in 32D murine hemopoietic cells, nuclear translocation of IRS-1 correlates with a markedly increased rRNA synthesis. Our experiments suggest that nuclear IRS-1 may play a specialized role in rRNA synthesis and/or processing.The insulin receptor substrate (IRS) 1 proteins are a family of docking proteins, which include IRS-1-4, Gab-1, and p62 dok (1). IRS-1 was the first to be identified as a docking protein for both the insulin and the IGF-I receptors. It transmits a signal from both receptors by interacting with a number of partners, including phosphatidylinositol 3-kinase, SHP2, Grb2, Crk, and others (2, 3). Tyrosine kinase activity of the receptors and phosphorylation of IRS-1 are essential steps in signal transduction. Of the downstream signals generated by IRS-1, the best studied is the phosphatidylinositol 3-kinase signaling pathway, which plays a major role in a number of biological responses to growth factors (1, 4). IRS-1 interacts directly with both the insulin and the IGF-I receptors, and the domains required for their interaction have also been identified (5). Because of its direct interaction with the receptors, its size, and its downstream signaling, it has been generally assumed that IRS-1 is an exclusively cytosolic (or plasma membrane) protein (6, 7). However, IRS-1 is known to interact with the SV40 T antigen (8, 9) and nucleolin (10). T antigen and nucleolin are predominantly nuclear proteins, although minor fractions of either protein can be found in the cytosol (11-13). It has been therefore tacitly assumed that IRS-1, anchored to the receptor, was interacting with the minor cytosolic fractions of T antigen and nucleolin. There is evidence, however, that signal-transduction molecules can translocate to the nucleus. They include mitogen-activated protein kinase (14), p70 S6K /TOR (15, 16), the STAT proteins (17, 18), Akt (19), ␤-catenin (20), the epidermal growth factor receptor (21), phosphatases (22), IRS-3 (23), and a cleaved ErbB-4 receptor (24). Indeed, Jans and Hassan (25) have summarized in a review the evidence that growth factors and their receptors can accumulate in the nuclei of cells. IGF-I, IGF-IR, and IRS-1 are not mentioned in that review, but insulin is (26).T...

Nuclear insulin receptor substrate-1 activates promoters of cell cycle progression genes

“…Activation of the cyclin D1 and c-myc promoters in R þ and RÀ cells stimulated by IGF-1 In R þ cells stimulated with IGF-1, IRS-1 is nuclear, and rDNA promoter activity and rRNA synthesis increase sharply (Tu et al, 2002;Wu et al, 2005). By ChIPs, IRS-1 is detectable in the rDNA promoter (Chen et al, 2005).…”

“…In transformed MEFs with high IGF-IR numbers, like R þ cells (Sell et al, 1993), IRS-1 is largely nuclear (Tu et al, 2002). In cells with lower numbers of IGF-IR (R12 cells, and other cell types), IRS-1 is confined to the cytoplasm.…”

“…By ChIPs, IRS-1 is detectable in the rDNA promoter (Chen et al, 2005). IGF-1 fails to translocate IRS-1 to nuclei of RÀ cells (Tu et al, 2002) that are MEFs with a targeted disruption of the IGF-IR genes (Sell et al, 1993). We asked whether nuclear IRS-1 in R þ cells would also increase transcription from RNA polymerase II promoters.…”

“…We monitored promoter occupancy in these cells, and the results are shown in Figure 4b. The only protein detected in the rDNA, cyclin D1 and c-myc promoters of R12 cells is the Nuclear IRS-1 and transcription A Wu et al UBF protein, an exquisitely nuclear/nucleolar protein (Tu et al, 2002).…”

“…More recently, IRS-1 has been found in nuclei and nucleoli of cells, either after stimulation with IGF-1 (Tu et al, 2002), or in association with the T antigens of SV40 or its human equivalent (Lassak et al, 2002;Prisco et al, 2002). Nuclear IRS-1 binds the upstream binding factor 1 (UBF1), one of the proteins that regulate RNA polymerase I activity (Tu et al, 2002).…”