Secretory vesicle swelling by atomic force microscopy.
The swelling of secretory vesicles has been implicated in exocytosis, but the underlying mechanism of vesicle swelling remained unknown. Earlier studies from our laboratory demonstrated the association of the alpha-subunit of heterotrimeric GTP-binding protein G(alphai3) with zymogen granule membrane and implicated its involvement in vesicle swelling. Mas7, an active mastoparan analog known to stimulate Gi proteins, was found to stimulate the GTPase activity of isolated zymogen granules and cause swelling. Increase in vesicle size in the presence of GTP, NaF, and Mas7 were irreversible and found to be KCl sensitive. However, Ca2+ had no effect on zymogen granule size. Taken together, these results indicated that zymogen granules, the membrane-bound secretory vesicles in exocrine pancreas, swell in response to GTP mediated by a G(alphai3) protein. Subsequently, our studies demonstrated that the water channel aquaporin-1 (AQP1) is also present at the zymogen granule membrane and participates in rapid GTP-induced and G(alphai3)-mediated vesicular water gating and swelling. Isolated zymogen granules exhibit low basal water permeability. However, exposure of granules to GTP results in a marked potentiation of water entry. Treatment of zymogen granules with the known water channel inhibitor Hg2+ is accompanied by a reversible loss in both the basal and GTP-stimulable water entry and vesicle swelling. Introduction of AQP1-specific antibody raised against the carboxy-terminal domain of AQP1 blocked GTP-stimulable swelling of vesicles. Our results demonstrate that AQPI associated at the zymogen granule membrane is involved in basal GTP-induced and G(alphai3)-mediated rapid gating of water into zymogen granules of the exocrine pancreas.
Cho
Sang-Joon
SJ
Department of Physiology, Wayne State University School of Medicine, Detroit, MI, USA.
Jena
Bhanu P
BP
D000818:Animals; D051398:Aquaporin 1 / Q000378:metabolism / Q000648:ultrastructure; D005089:Exocytosis / Q000502:physiology; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism; D018625:Microscopy, Atomic Force; D016253:Microscopy, Immunoelectron; D010179:Pancreas / Q000166:cytology; D010316:Particle Size; D051381:Rats; D022142:Secretory Vesicles / Q000378:metabolism / Q000648:ultrastructure
D051398:Aquaporin 1; D019206:GTP-Binding Protein alpha Subunits, Gi-Go