Involvement of Rho GTPases and their effectors in the secretory process of PC12 cells.
We investigated the involvement of Rho GTPases in the secretory process of PC12 cells. Overexpression of wild-type RhoA, Rac1, or Cdc42 did affect exocytosis. In contrast, secretion elicited by depolarizing K(+) concentrations was enhanced by the dominant negative mutants RhoA(N19), Rac1(N17), and Cdc42(N17) and was diminished by the constitutively active mutants RhoA(V14), Rac1(V12), and Cdc42(V12). The inhibition observed in the presence of RhoA(V14) was likely a result of the activation of ROK(alpha), since the catalytic domain of this kinase was able to mimic both the reorganization of the actin cytoskeleton and the decrease in exocytosis induced by the RhoA mutant. Part of the effect of Rac1(V12) may be due to POR1 activation. Thus, overexpression of full-length POR1 diminished K(+)-stimulated exocytosis, and a point mutation in the effector domain of Rac1(V12) that prevents the interaction with POR1 abolished the inhibitory effect of the GTPase. We also searched for the Cdc42(V12) target but overexpression of the Cdc42 effector WASP did not mimic the inhibition of exocytosis observed in cells transfected with the activated GTPase. Our findings indicate that different signaling cascades resulting in the activation of RhoA, Rac1, or Cdc42 can modulate the exocytotic process of neuroendocrine cells.
Frantz
Christian
C
Institut de Biologie Cellulaire et de Morphologie, University of Lausanne, Lausanne, Switzerland.
Coppola
Thierry
T
Regazzi
Romano
R
D000199:Actins / Q000378:metabolism; D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D003599:Cytoskeleton / Q000502:physiology; D005089:Exocytosis / Q000502:physiology*; D019382:Human Growth Hormone / Q000378:metabolism; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D016297:Mutagenesis, Site-Directed; D016716:PC12 Cells; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011506:Proteins / Q000235:genetics / Q000378:metabolism; D051381:Rats; D015398:Signal Transduction; D051316:Wiskott-Aldrich Syndrome Protein; D020764:cdc42 GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*; D054460:rho-Associated Kinases; D020742:rhoA GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*
C101158:ARFIP2 protein, human; D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D047908:Intracellular Signaling Peptides and Proteins; D011506:Proteins; C495767:WAS protein, human; C495769:Was protein, rat; D051316:Wiskott-Aldrich Syndrome Protein; D019382:Human Growth Hormone; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D020764:cdc42 GTP-Binding Protein; D020830:rac1 GTP-Binding Protein; D020742:rhoA GTP-Binding Protein