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Acta Physiologica Congress

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Acta Physiologica 2013; Volume 207, Supplement 694
92nd Annual Meeting of the German Physiological Society
3/2/2013-3/5/2013
Heidelberg, Germany


ACTIN COATING OF LAMELLAR BODIES DURING THE POST-FUSION PHASE OF EXOCYTOSIS IS MEDIATED BY RHO GTPASES
Abstract number: P014

Ehinger 1   *K. , Neuland 1  K., Timmler 1  M., Wittekindt 1  O.H., Dietl 1  P., Frick 1  M., Miklavc 1  P.

1 University of Ulm, Institute of General Physiology, Ulm, Germany

Actin plays an important role for regulated exocytosis of secretory vesicles. During the post-fusion phase of exocytosis, actin can stabilize the fusion pore and/or compress the exocytotic vesicle, promoting release of secretory material. Alveolar type II (ATII) pneumocytes contain lamellar bodies (LBs), large secretory vesicles that store pulmonary surfactant, a complex mixture of proteins and mostly lipids. Fusion of LBs with the plasma membrane is followed by formation of an actin coat around the vesicle. Compression of the actin coat is necessary for surfactant release. Here, we show that Rho GTPases are required for actin coat formation in primary ATII cells. Inhibition of small GTPases by Clostridium difficile toxin B and inhibition of the Rho GTPase subfamily by C3 transferase prevented actin coating and subsequent vesicle contraction. In contrast, inhibition of the Rac or cdc42 subfamilies with NSC23766 and secramine, respectively, showed no effect. Expression of a dominant-negative Rho construct significantly reduced the frequency of vesicle coating with actin. Using fluorescently-tagged rhotekin GTP-binding domain (rGBD) in real-time, live-cell imaging experiments we observed activation of Rho GTPases on the limiting membrane of LBs following fusion. Activation was transient and preceded actin coat formation. We also observed translocation of GFP-labeled RhoB, RhoD, and RhoG to the LB membrane after exocytosis. Quantitative PCR analysis confirmed high expression of RhoB and RhoG in ATII cells. Together, our data show that Rho GTPases are transiently activated on LBs after fusion and this activation is responsible for subsequent actin coating.

To cite this abstract, please use the following information:
Acta Physiologica 2013; Volume 207, Supplement 694 :P014

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