Mutant α-subunit of the G protein G12 activates proliferation and inhibits differentiation of 3T3-F442A preadipocytes

We studied the G protein alpha-subunit G alpha 12 in various tissues and cell lines. Significant amounts of G alpha 12 were detected by immunoblots in liver, chromaffin cells, RINm5F cells, 3T3-F442A cells, and preadipocytes, but not in adipocytes, sperm, kidney, NB2A cells, or brain. To study the role of G alpha 12 in adipose tissue differentiation, the preadipocyte cell line 3T3-F442A was transfected with wild-type G alpha 12 or a constitutively activated mutant of G alpha 12. Stable expression of the activated mutant of G alpha 12 stimulated cell growth and inhibited preadipocyte differentiation. In contrast, wild-type G alpha 12 overexpression inhibited preadipocyte differentiation, without any effect on cell proliferation. The role of G alpha 12 on the Raf/MEK/mitogen-activating protein kinase (MAPK) cascade was studied. In confluent preadipocytes, expression of the activated mutant of G alpha 12 induced an increase in B-Raf expression, but no change in MAPK activity. Differentiation was associated with a decrease in MAPK activity in control 3T3-F442A cells. Wild-type G alpha 12 overexpression prevented the decrease in MAPK activity and induced MEK1, but not B-Raf, expression. Moreover, the activated mutant of G alpha 12 induced an increase in MAPK activity and in the expression of both MEK1 and B-Raf. These data indicate that the activated mutant of G alpha 12 stimulates the proliferation of 3T3-F442A preadipocytes, possibly through an increase in B-Raf expression, independently of the MEK/MAPK pathway, but prevents differentiation, probably through an increase in MEK1 expression and MAPK activity.