Dominant negative alpha-subunit of farnesyl- and geranylgeranyl-transferase I inhibits insulin-induced differentiation of 3T3-L1 pre-adipocytes.
OBJECTIVE
To investigate whether the expression of a dominant negative (DN) farnesyl- and geranygeranyl-transferase I (FTase/GGTase I) alpha-subunit in 3T3-L1 pre-adipocytes can inhibit insulin's ability to induce differentiation.


METHODS
3T3-L1 pre-adipocytes were stably transfected with vector alone or vector expressing a mutated DN FTase/GGTase I alpha-subunit (S60A)(S62A) and incubated in serum-free medium in the absence and presence of insulin.


METHODS
Various assays were performed to determine the effect of DN FTase/GGTase I alpha-subunit expression in 3T3-L1 pre-adipocyte on insulin-induced DNA synthesis, cell count, phosphorylation of the FTase/GGTase I alpha-subunit, FTase and GGTase I activity, amounts of prenylated p21Ras and RhoA, phosphorylation of MAP kinase and Akt, and differentiation to mature fat cells.


RESULTS
Expression of DN FTase/GGTase I alpha-subunit inhibited insulin's ability to increase DNA synthesis, cell count, FTase and GGTase I activity, amounts of prenylated p21Ras and RhoA, and magnitude of phosphorylation of MAP kinase. Expression of DN FTase/GGTase I alpha-subunit in 3T3-L1 pre-adipocytes was without effect on insulin-induced Akt phosphorylation.


CONCLUSIONS
Expression of DN FTase/GGTase I alpha-subunit inhibits insulin-induced differentiation of 3T3-L1 pre-adipocytes to mature adipocytes, and thus could indicate potential therapeutic avenues to assuage the deleterious effects of obesity and type 2 diabetes.
Solomon
C S
CS
Research Service, Denver VA Medical Center, Denver, Colorado 80220, USA.
Leitner
J W
JW
Goalstone
M L
ML
D016475:3T3 Cells / Q000187:drug effects; D017667:Adipocytes / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D019883:Alkyl and Aryl Transferases / Q000494:pharmacology*; D000818:Animals; D015153:Blotting, Western; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D003924:Diabetes Mellitus, Type 2 / Q000378:metabolism; D051231:Farnesyltranstransferase; D006801:Humans; D007004:Hypoglycemic Agents / Q000494:pharmacology*; D007328:Insulin / Q000494:pharmacology*; D051379:Mice; D010766:Phosphorylation; D016283:Proto-Oncogene Proteins p21(ras) / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000378:metabolism
D007004:Hypoglycemic Agents; D007328:Insulin; D019883:Alkyl and Aryl Transferases; C083291:geranylgeranyltransferase type-I; D051231:Farnesyltranstransferase; C501009:HRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras); D020742:rhoA GTP-Binding Protein