G-ALPHA(I-2) MEDIATES RENAL LLC-PK1 GROWTH BY A RAF-INDEPENDENT ACTIVATION OF P42 P44 MAP KINASE/

The protooncogene G alpha(i-2) plays a pivotal role in signaling pathw ays that control renal cell growth and differentiation. Mitogen-activa ted protein kinases (MAPKs) are potential downstream effecters for G a lpha(i-2) in these pathways. In predifferentiated LLC-PK1 renal cells, the temporal maximal expression of G alpha(i-2) coincided with maxima l activation of MAPK((p42/p44)). By contrast, pertussis toxin treatmen t of these cells inhibited cell growth and reduced MAPK((p42/p44)) act ivity by 30%. These findings reflected upstream activation of MAPK kin ase (MEK1), as transient transfection of cells with a plasmid encoding a constitutively active form of MEK1 increased MAPK((p42/p44)) activi ty and cell growth, whereas treatment with PD-098059, an inhibitor of MEK1 activity, reduced MAPK((p42/p44)) activity and cell growth. Expre ssion of a guanosinetriphosphatase (GTPase)-deficient G alpha(i-2) in these cells increased MAPK((p42/p44)) activity and correspondingly red uced cell doubling time from 24 to 10 h without altering the activity of Raf-1 or c-Jun/stress-activated protein kinases (SAPKs). By contras t, expression of a GTPase-deficient G alpha(i-3) in these cells reduce d both their cell doubling time by 30% and MAPK((p42/p44)) activity by 60%. As the known MEKK isoforms (MEKK1, -2, and -3) can also activate SAPKs, these findings suggest the GTP-charged G alpha(i-2) subunit tr ansduces growth signals in renal cells via activation of MAPK((p42/p44 )) and that such activation may be linked to pathways containing novel MEKK isoforms that preferentially activate MEKs.