Vesicular calcium regulates coat retention, fusogenicity, and size of pre-Golgi intermediates.
The significance and extent of Ca(2+) regulation of the biosynthetic secretory pathway have been difficult to establish, and our knowledge of regulatory relationships integrating Ca(2+) with vesicle coats and function is rudimentary. Here, we investigated potential roles and mechanisms of luminal Ca(2+) in the early secretory pathway. Specific depletion of luminal Ca(2+) in living normal rat kidney cells using cyclopiazonic acid (CPA) resulted in the extreme expansion of vesicular tubular cluster (VTC) elements. Consistent with this, a suppressive role for vesicle-associated Ca(2+) in COPII vesicle homotypic fusion was demonstrated in vitro using Ca(2+) chelators. The EF-hand-containing protein apoptosis-linked gene 2 (ALG-2), previously implicated in the stabilization of sec31 at endoplasmic reticulum exit sites, inhibited COPII vesicle fusion in a Ca(2+)-requiring manner, suggesting that ALG-2 may be a sensor for the effects of vesicular Ca(2+) on homotypic fusion. Immunoisolation established that Ca(2+) chelation inhibits and ALG-2 specifically favors residual retention of the COPII outer shell protein sec31 on pre-Golgi fusion intermediates. We conclude that vesicle-associated Ca(2+), acting through ALG-2, favors the retention of residual coat molecules that seem to suppress membrane fusion. We propose that in cells, these Ca(2+)-dependent mechanisms temporally regulate COPII vesicle interactions, VTC biogenesis, cargo sorting, and VTC maturation.
Bentley
Marvin
M
Division of Biological Sciences and Center for Structural and Functional Neuroscience, The University of Montana, Missoula, MT 59812-4824, USA.
Nycz
Deborah C
DC
Joglekar
Ashwini
A
Fertschai
Ismene
I
Malli
Roland
R
Graier
Wolfgang F
WF
Hay
Jesse C
JC
D000818:Animals; D022181:COP-Coated Vesicles / Q000378:metabolism* / Q000648:ultrastructure*; D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000378:metabolism; D002478:Cells, Cultured; D002614:Chelating Agents / Q000737:chemistry / Q000378:metabolism; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism* / Q000648:ultrastructure; D008561:Membrane Fusion / Q000502:physiology*; D015394:Molecular Structure; D051381:Rats; D055571:Secretory Pathway / Q000502:physiology
D002135:Calcium-Binding Proteins; D002614:Chelating Agents; D004533:Egtazic Acid; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; D002118:Calcium