Association of the G protein α(q)/α11-subunit with cytoskeleton in adrenal glomerulosa cells: Role in receptor-effector coupling

Abstract

In 3-day primary cultures of rat glomerulosa cells, a 30-min preincubation with either 10 μM colchicine (a microtubule-disrupting agent) or 10 μM cytochalasin B (a microfilament-disrupting agent) decreased angiotensin II (Ang II)-induced inositol phosphate accumulation by 50%. Moreover, both drugs decreased inositol phosphate production induced by fluoroaluminate (a nonspecific activator of all G proteins), indicating that both microtubules and microfilaments are essential for phospholipase C activation. Analysis of microfilament- and microtubule-enriched fractions and immunoprecipitation of actin and tubulin revealed that the α(q)/α11- subunit of the G(q/11) protein was associated with both structures. Ang II stimulation induced a rapid translocation of α(q)/α11, microfilaments, and microtubules to the membrane and induced a time-dependent increase in the level of α(q)/α11 associated with both microfilaments and microtubules. Moreover, double immunofluorescence staining clearly showed a colocalization of the α(q)/α11-subunit of the G(q/11) coupling protein and microfilament distribution. These associations and plasma membrane redistribution under Ang II stimulation indicate that microfilaments and microtubules are both involved in phospholipase C activation and inositol phosphate production. Moreover, our results indicate that the α(q)/α11 protein is closely associated with cytoskeletal elements and is found both at the plasma MeMbrane level as well as on intracellular stress fibers.