Thrombin induces the release of the Y-box protein dbpB from mRNA: A mechanism of transcriptional activation

Abstract

We have recently demonstrated that thrombin induces expression of the platelet-derived growth factor B-chain gene in endothelial cells (EC) through activation of the Y-box binding protein DNAbinding protein B (dbpB). We now present evidence that dbpB is activated by a novel mechanism: proteolytic cleavage leading to release from mRNA, nuclear translocation, and induction of thrombin-responsive genes. Cytosolic, full-length dbpB (50 kDa) was rapidly cleaved to a 30-kDa species upon thrombin stimulation of EC. This truncated, ''active'' dbpB exhibited nuclear localization and binding affinity for the thrombin response element sequence, which is distinct from the Y-box sequence. Oligo(dT) affinity chromatography revealed that cytosolic dbpB from control EC, but not active dbpB from thrombin-treated EC, was bound to mRNA. Latent dbpB immunoprecipitated from cytosolic extracts of control EC was activated by ribonuclease treatment. Furthermore, when EC cytosolic extracts were subjected to Nycodenz gradient centrifugation, latent dbpB fractionated with mRNA, whereas active dbpB fractionated with free proteins. The cytosolic retention domain of dbpB, which we localized to the region 247-267, was proteolytically cleaved during its activation. In contrast to full-length dbpB, truncated dbpB stimulated platelet-derived growth factor B-chain and tissue factor promoter activity by over 5-fold when transiently cotransfected with reporter constructs. These results suggest a novel mode of transcription factor activation in which an agonist causes release from mRNA of a latent transcription factor leading to its transport to the nucleus and its regulation of target gene expression.DNA-binding protein B ͉ endothelium ͉ platelet-derived growth factor ͉ RNA-binding proteins

Phosphorylation by Akt disables the anti-oncogenic activity of YB-1

“…This view is supported by recent evidence that shows an accumulation of proteolytically cleaved YB-1 species in various cancer tissues (Sorokin et al, 2005). These truncated YB-1 proteins carry the CSD but lack the C-terminal cytoplasmic retention domain and therefore localize to the cell nucleus (Stenina et al, 2001;Jurchott et al, 2003;. The mechanism by which nuclear YB-1 contributes to oncogenesis is not known but may be related to the transcriptional and chromosomal regulatory activities of nuclear YB-1 (Bader, 2006).…”

Y-Box Protein 1 Mediates PDGF-B Effects in Mesangioproliferative Glomerular Disease

“…To confirm the staining pattern and exclude the detection of a subfragment of YB-1 protein, as has been reported by Stenina et al (4,45), this observation was verified using three different primary antibodies directed against domains located in both the YB-1 N-and C-terminus. The staining pattern changed dramatically during the course of an experimental model of mesangioproliferative GN: A predominant cytoplasmic localization that reversed back to nuclear in parallel to resolution of disease at approximately day 31 was observed.…”

Y-box factor YB1 controls p53 apoptotic function

“…However, in vitro, we found this to be uncommon occurring in less than 10% of cells examined after treatment with several DNA-damaging agents . Nonetheless, several reports indicate that nuclear translocation of YB1 is necessary for it to function as a transcription factor, as might be expected (Bargou et al, 1997;Oda et al, 1998Oda et al, , 2003Stenina et al, 2001;Holm et al, 2002). In addition, nuclear YB1 appears to be important clinically as it has been associated with poor prognosis in several types of human cancer (Bargou et al, 1997;Oda et al, 1998Oda et al, , 2003Kamura et al, 1999;Shibahara et al, 2001).…”