EVIDENCE FOR RAP1 IN VASCULAR SMOOTH-MUSCLE CELLS - REGULATION OF THEIR EXPRESSION BY PLATELET-DERIVED GROWTH-FACTOR BB

The effect of platelet-derived growth factor (PDGF) on Rapl expression was investigated in rat vascular smooth muscle cells (SMC). First, ev idence for Rapl proteins was shown by their: (i) detection in membrane s using a specific anti-Rapl antibody, (ii) typical shift in electroph oretic mobility as a consequence of reduction, and (iii) cAMP-induced phosphorylation and immunoprecipitation. Then, the mitogenic activity of 10 ng/ml PDGF AA and BB for 48 h, resulting in a 2- and 5-fold incr ease in H-3thymidine incorporation, was correlated with that of tota l Rapl protein expression which was found to be 99% +/- 36% and 260% /- 70%, respectively. Further time-course studies established that thi s up-regulation of Rapl proteins was only observed after 48 h of PDGF BB treatment. Lastly, comparative RT-PCR of both rapla and raplb mRNAs showed that PDGF BB also up-regulated the rapla mRNA species, which w as 1.5-fold increased in contrast with the raplb mRNA species. It is c oncluded that the PDGF BB-induced SMC proliferation is associated with an up-regulation of Rapla protein.