Antisense oligodeoxynucleotides to G_s protein α-subunit sequence accelerate differentiation of fibroblasts to adipocytes

Abstract

FULLY-DIFFERENTIATED mouse 3T3-L1 fibroblasts accumulate large amounts of lipid at 7–10 days after induction by insulin or by dexamethasone and a methyl xanthine^1–3. G proteins mediate transmembrane signalling from a diverse group of cell-surface receptors to effector units that include phospholipase C, adenylyl cyclase and ion channels^4–6. They are also targets of regulation themselves^7–10.3T3-L1 fibroblasts display marked changes in levels of G protein when induced to differentiate to adipocytes^11–15. Here we show that cholera toxin, which ADP-ribosylates and activates the G protein subunit G_sα, blocks the induction of differentiation, whereas increasing intracellular cyclic AMP directly with the dibutyryl analogue or indirectly with pertussis toxin or forskolin does not affect differentiation. Oligodeoxynucleotides antisense to the sequence encoding G_sα accelerate differentiation markedly. The time course of adipogenesis declined from 7–10 days in controls to roughly 3 days in cultures treated with antisense-G_sα oligodeoxynucleotides, whereas oligodeoxynucleotides, antisense to G_iα1, G_iα3, and sense and missense to G_sα, had no such effect. Antisense-G_sα alone induced differentiation by day 7, indicating that G_sα activity modulates differentiation in 3T3-L1 cells, acting in a new role which is independent of increased intracellular cAMP.