Platelet-derived growth factor stimulation of mitogen-activated protein kinases and cyclin D-1 promoter activity in cultured airway smooth-muscle cells - Role of Ras

We hypothesized that in bovine tracheal myocytes, growth factor treatment i nduces transcription from the cyclin D-1 promoter that is dependent on the activation of both Ras and extracellular signal-related kinase (ERK), We fo und that platelet-derived growth factor (PDGF) treatment induced substantia l activation of ERK2 that was blocked by expression of a dominant-negative Ha-Ras. Further, expression of a constitutively active Ha-Ras induced subst antial ERK2 activity, consistent with the notion that Pas is required and s ufficient for ERK activation. PDGF treatment induced only modest activation of the Jun amino terminal kinase-1 (JNK1) and p38 mitogen-activated protei n kinases (MAPKs). Active Pas induced similar responses, implying that comp lete activation of the JNK and p38 pathways requires additional or alternat ive upstream signaling intermediates besides Pas. In contrast, expression o f a constitutively active Rac1, an alternative guanosine triphosphatase inv olved in intracellular signaling, produced a high level of JNK1 activation, suggesting that Rad is an important upstream activator of JNK in this syst em. 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 m yocytes: (I) activation of MAPK by PDGF is dependent on Pas; (2) active Pas is sufficient for ERK activation but is insufficient for maximal activatio n of JNK or p38; (3) activation of Rac1 is sufficient for maximal JNK activ ation; and (4) Ras, MEK, and ERK constitute a distinct pathway to cyclin D- 1 transcriptional activation.