Association of the G protein alpha(q)/alpha11-subunit with cytoskeleton in adrenal glomerulosa cells: role in receptor-effector coupling.
In 3-day primary cultures of rat glomerulosa cells, a 30-min pre-incubation with either 10 microM colchicine (a microtubule-disrupting agent) or 10 microM 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 alpha(q)/alpha11-subunit of the G(q/11) protein was associated with both structures. Ang II stimulation induced a rapid translocation of alpha(q)/alpha11, microfilaments, and microtubules to the membrane and induced a time-dependent increase in the level of alpha(q)/alpha11 associated with both microfilaments and microtubules. Moreover, double immunofluorescence staining clearly showed a colocalization of the alpha(q)/alpha11-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 alpha(q)/alpha11 protein is closely associated with cytoskeletal elements and is found both at the plasma membrane level as well as on intracellular stress fibers.
Côté
M
M
Department of Medicine, Faculty of Medicine, University of Sherbrooke, Quebec, Canada.
Payet
M D
MD
Dufour
M N
MN
Guillon
G
G
Gallo-Payet
N
N
D008841:Actin Cytoskeleton / Q000737:chemistry / Q000648:ultrastructure; D000199:Actins / Q000032:analysis; D000450:Aldosterone / Q000378:metabolism; D000535:Aluminum / Q000494:pharmacology; D000704:Analysis of Variance; D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D002462:Cell Membrane / Q000737:chemistry / Q000502:physiology / Q000648:ultrastructure; D002478:Cells, Cultured; D003078:Colchicine / Q000494:pharmacology; D003571:Cytochalasin B / Q000494:pharmacology; D003599:Cytoskeleton / Q000737:chemistry* / Q000502:physiology / Q000648:ultrastructure; D004789:Enzyme Activation; D005260:Female; D005455:Fluorescent Antibody Technique; D005461:Fluorine / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000819:agonists / Q000032:analysis* / Q000502:physiology*; D007295:Inositol Phosphates / Q000378:metabolism; D011233:Precipitin Tests; D051381:Rats; D013997:Time Factors; D014404:Tubulin / Q000032:analysis; D010738:Type C Phospholipases / Q000378:metabolism / Q000502:physiology; D015384:Zona Glomerulosa / Q000737:chemistry* / Q000166:cytology* / Q000648:ultrastructure
D000199:Actins; D007295:Inositol Phosphates; D014404:Tubulin; D000804:Angiotensin II; C053629:fluoroaluminum; D005461:Fluorine; D003571:Cytochalasin B; D000450:Aldosterone; D000535:Aluminum; D010738:Type C Phospholipases; D019204:GTP-Binding Proteins; D003078:Colchicine