Stomach-specific calpain, nCL-2, localizes in mucus cells and proteolyzes the beta-subunit of coatomer complex, beta-COP.
Calpain is a Ca2+-regulated cytosolic protease. Mammals have 14 calpain genes, half of which are predominantly expressed in specific organ(s); the rest are expressed ubiquitously. A defect in calpains causes lethality/pathogenicity, indicating their physiological indispensability. nCL-2/calpain-8a was identified as a stomach-specific calpain, whose physiological functions are unclear. To elucidate these, we characterized nCL-2 in detail. Unexpectedly, nCL-2 was localized strictly to the surface mucus cells in the gastric epithelium and the mucus-secreting goblet cells in the duodenum. Yeast two-hybrid screening identified several nCL-2-interacting molecules. Of these, the beta-subunit of coatomer complex (beta-COP) occurs in the stomach pit cells and is proteolyzed by nCL-2 in vitro. Furthermore, beta-COP and nCL-2 co-expressed in COS7 cells co-localized in the Golgi, and Ca2+-ionophore stimulation caused the proteolysis of beta-COP near the linker region, resulting in the dissociation of beta-COP from the Golgi. These results strongly suggest novel functions for nCL-2 that involve the membrane trafficking of mucus cells via interactions with coat protein.
Hata
Shoji
S
Department of Enzymatic Regulation for Cell Functions, The Tokyo Metropolitan Institute of Medical Science (Rinshoken), Tokyo 113-8613, Japan.
Koyama
Suguru
S
Kawahara
Hiroyuki
H
Doi
Naoko
N
Maeda
Tatsuya
T
Toyama-Sorimachi
Noriko
N
Abe
Keiko
K
Suzuki
Koichi
K
Sorimachi
Hiroyuki
H
D000818:Animals; D015152:Blotting, Northern; D019556:COS Cells; D002118:Calcium / Q000378:metabolism; D002154:Calpain / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D018076:DNA, Complementary / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004848:Epithelium / Q000378:metabolism; D005753:Gastric Mucosa / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism; D015151:Immunoblotting; D047468:Immunoprecipitation; D017403:In Situ Hybridization; D007476:Ionophores / Q000494:pharmacology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008954:Models, Biological; D009093:Mucus / Q000378:metabolism*; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D013997:Time Factors; D015533:Transcriptional Activation; D020798:Two-Hybrid System Techniques
D017931:DNA Primers; D018076:DNA, Complementary; D007476:Ionophores; D012333:RNA, Messenger; D005982:Glutathione Transferase; D002154:Calpain; C120543:Capn8 protein, mouse; D002118:Calcium