Platelet-derived growth factor stimulation of mitogen-activated protein kinases and cyclin D1 promoter activity in cultured airway smooth-muscle cells -: Role of Ras

We hypothesized that in bovine tracheal myocytes, growth factor treatment induces transcription from the cyclin D-1 promoter that is dependent on the activation of both Ras and extracellular signal-related kinase (ERK), We found that platelet-derived growth factor (PDGF) treatment induced substantial activation of ERK2 that was blocked by expression of a dominant-negative Ha-Ras. Further, expression of a constitutively active Ha-Ras induced substantial ERK2 activity, consistent with the notion that Pas is required and sufficient for ERK activation. PDGF treatment induced only modest activation of the Jun amino terminal kinase-1 (JNK1) and p38 mitogen-activated protein kinases (MAPKs). Active Pas induced similar responses, implying that complete activation of the JNK and p38 pathways requires additional or alternative upstream signaling intermediates besides Pas. In contrast, expression of a constitutively active Rac1, an alternative guanosine triphosphatase involved in intracellular signaling, produced a high level of JNK1 activation, suggesting that Rad is an important upstream activator of JNK in this system. Active Pas and MAPK/ERK kinase-l (MEK1) (the upstream activator of ERK) each induced cyclin D-1 promoter activity, whereas active stress-activated protein kinase/ERK kinase-l (SEK1), an upstream activator of JNK, did not. Finally, the synthetic MEK inhibitor PD98059 blocked Ras-induced cyclin D-1 promoter activity. Together, these data suggest that in bovine tracheal myocytes: (I) activation of MAPK by PDGF is dependent on Pas; (2) active Pas is sufficient for ERK activation but is insufficient for maximal activation of JNK or p38; (3) activation of Rac1 is sufficient for maximal JNK activation; and (4) Ras, MEK, and ERK constitute a distinct pathway to cyclin D-1 transcriptional activation.