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G alpha(i-2) mediates renal LLC-PK1 growth by a Raf-independent activation of p42/p44 MAP kinase
1Department of Medicine, Massachusetts General Hospital, Harvard Medical School, Boston 02114, USA.
Abstract:
The protooncogene G alpha(i-2) plays a pivotal role in signaling pathways that control renal cell growth and differentiation. Mitogen-activated protein kinases (MAPKs) are potential downstream effectors for G alpha(i-2) in these pathways. In predifferentiated LLC-PK1 renal cells, the temporal maximal expression of G alpha(i-2) coincided with maximal activation of MAPK(p42/p44). By contrast, pertussis toxin treatment of these cells inhibited cell growth and reduced MAPK(p42/p44) activity by 30%. These findings reflected upstream activation of MAPK kinase (MEK1), as transient transfection of cells with a plasmid encoding a constitutively active form of MEK1 increased MAPK(p42/p44) activity and cell growth, whereas treatment with PD-098059, an inhibitor of MEK1 activity, reduced MAPK(p42/p44) activity and cell growth. Expression of a guanosinetriphosphatase (GTPase)-deficient G alpha(i-2) in these cells increased MAPK(p42/p44) activity and correspondingly reduced cell doubling time from 24 to 10 h without altering the activity of Raf-1 or c-Jun/stress-activated protein kinases (SAPKs). By contrast, expression of a GTPase-deficient G alpha(i-3) in these cells reduced 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 transduces 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.
Insights
The protooncogene G alpha(i-2) activates mitogen-activated protein kinases (MAPKs) to promote renal cell growth. Inhibiting this pathway with pertussis toxin reduces cell growth and MAPK activity, highlighting G alpha(i-2) as a key regulator.
Area of Science:
- Cellular signaling
- Oncogenes
- Renal cell biology
Background:
- The protooncogene G alpha(i-2) is crucial for renal cell growth and differentiation signaling.
- Mitogen-activated protein kinases (MAPKs) are potential downstream effectors of G alpha(i-2) in these pathways.
Purpose of the Study:
- To investigate the role of G alpha(i-2) in regulating renal cell growth via MAPK signaling.
- To elucidate the specific mechanisms by which G alpha(i-2) influences MAPK activation and cell proliferation.
Main Methods:
- LLC-PK1 renal cells were used to study G alpha(i-2) expression and MAPK activation.
- Pertussis toxin was employed to inhibit G alpha(i-2) signaling.
- Transfection with constitutively active MEK1 and treatment with PD-098059 were used to assess MEK1's role.
- Expression of GTPase-deficient G alpha(i-2) and G alpha(i-3) mutants was analyzed.
Main Results:
- Maximal G alpha(i-2) expression correlated with maximal MAPK(p42/p44) activation in LLC-PK1 cells.
- Pertussis toxin inhibited cell growth and reduced MAPK(p42/p44) activity.
- Constitutively active MEK1 increased MAPK(p42/p44) activity and cell growth, while PD-098059 inhibited both.
- GTPase-deficient G alpha(i-2) enhanced MAPK(p42/p44) activity and reduced cell doubling time.
- GTPase-deficient G alpha(i-3) reduced cell doubling time and MAPK(p42/p44) activity.
Conclusions:
- GTP-charged G alpha(i-2) subunit transduces growth signals in renal cells through MAPK(p42/p44) activation.
- This activation may involve novel MEKK isoforms preferentially activating MEKs.
- G alpha(i-2) plays a significant role in regulating renal cell proliferation via the MAPK pathway.