The Ca2+-binding protein ALG-2 is recruited to endoplasmic reticulum exit sites by Sec31A and stabilizes the localization of Sec31A.
The formation of transport vesicles that bud from endoplasmic reticulum (ER) exit sites is dependent on the COPII coat made up of three components: the small GTPase Sar1, the Sec23/24 complex, and the Sec13/31 complex. Here, we provide evidence that apoptosis-linked gene 2 (ALG-2), a Ca(2+)-binding protein of unknown function, regulates the COPII function at ER exit sites in mammalian cells. ALG-2 bound to the Pro-rich region of Sec31A, a ubiquitously expressed mammalian orthologue of yeast Sec31, in a Ca(2+)-dependent manner and colocalized with Sec31A at ER exit sites. A Ca(2+) binding-deficient ALG-2 mutant, which did not bind Sec31A, lost the ability to localize to ER exit sites. Overexpression of the Pro-rich region of Sec31A or RNA interference-mediated Sec31A depletion also abolished the ALG-2 localization at these sites. In contrast, depletion of ALG-2 substantially reduced the level of Sec31A associated with the membrane at ER exit sites. Finally, treatment with a cell-permeable Ca(2+) chelator caused the mislocalization of ALG-2, which was accompanied by a reduced level of Sec31A at ER exit sites. We conclude that ALG-2 is recruited to ER exit sites via Ca(2+)-dependent interaction with Sec31A and in turn stabilizes the localization of Sec31A at these sites.
Yamasaki
Akinori
A
Department of Biological Sciences, Tokyo Institute of Technology, Yokohama 226-8501, Japan.
Tani
Katsuko
K
Yamamoto
Akitsugu
A
Kitamura
Naomi
N
Komada
Masayuki
M
D051017:Apoptosis Regulatory Proteins / Q000737:chemistry / Q000378:metabolism*; D020126:Brefeldin A / Q000494:pharmacology; D002135:Calcium-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000378:metabolism; D022083:Cytoplasmic Structures / Q000187:drug effects / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism*; D056827:Endosomal Sorting Complexes Required for Transport; D006367:HeLa Cells; D006801:Humans; D020559:Monomeric GTP-Binding Proteins / Q000378:metabolism; D011392:Proline / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D034741:RNA, Small Interfering; D013816:Thermodynamics; D033921:Vesicular Transport Proteins
D051017:Apoptosis Regulatory Proteins; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D056827:Endosomal Sorting Complexes Required for Transport; C097753:PDCD6 protein, human; C117020:PDCD6IP protein, human; D034741:RNA, Small Interfering; C507455:SEC31A protein, human; D033921:Vesicular Transport Proteins; D020126:Brefeldin A; D011392:Proline; C493083:SAR1A protein, human; D020559:Monomeric GTP-Binding Proteins