A cytosolic splice variant of Cab45 interacts with Munc18b and impacts on amylase secretion by pancreatic acini.
We identified in a yeast two-hybrid screen the EF-hand Ca(2+)-binding protein Cab45 as an interaction partner of Munc18b. Although the full-length Cab45 resides in Golgi lumen, we characterize a cytosolic splice variant, Cab45b, expressed in pancreatic acini. Cab45b is shown to bind (45)Ca(2+), and, of its three EF-hand motifs, EF-hand 2 is demonstrated to be crucial for the ion binding. Cab45b is shown to interact with Munc18b in an in vitro assay, and this interaction is enhanced in the presence of Ca(2+). In this assay, Cab45b also binds the Munc18a isoform in a Ca(2+)-dependent manner. The endogenous Cab45b in rat acini coimmunoprecipitates with Munc18b, syntaxin 2, and syntaxin 3, soluble N-ethylmaleimide-sensitive factor attachment protein receptors with key roles in the Ca(2+)-triggered zymogen secretion. Furthermore, we show that Munc18b bound to syntaxin 3 recruits Cab45b onto the plasma membrane. Importantly, antibodies against Cab45b are shown to inhibit in a specific and dose-dependent manner the Ca(2+)-induced amylase release from streptolysin-O-permeabilized acini. The present study identifies Cab45b as a novel protein factor involved in the exocytosis of zymogens by pancreatic acini.
Lam
Patrick P L
PP
Department of Medicine, University of Toronto, Toronto, Ontario, Canada.
Hyvärinen
Kati
K
Kauppi
Maria
M
Cosen-Binker
Laura
L
Laitinen
Saara
S
Keränen
Sirkka
S
Gaisano
Herbert Y
HY
Olkkonen
Vesa M
VM
D017398:Alternative Splicing / Q000187:drug effects / Q000235:genetics*; D000681:Amylases / Q000378:metabolism*; D000818:Animals; D000906:Antibodies; D001426:Bacterial Proteins / Q000494:pharmacology; D016466:CHO Cells; D019556:COS Cells; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002463:Cell Membrane Permeability / Q000187:drug effects; D002522:Chlorocebus aethiops; D006224:Cricetinae; D003412:Cricetulus; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism*; D004285:Dogs; D020832:EF Hand Motifs; D020869:Gene Expression Profiling; D006023:Glycoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051938:Munc18 Proteins / Q000378:metabolism*; D046790:Pancreas, Exocrine / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D013301:Streptolysins / Q000494:pharmacology
D000906:Antibodies; D001426:Bacterial Proteins; D002135:Calcium-Binding Proteins; D006023:Glycoproteins; D051938:Munc18 Proteins; D012333:RNA, Messenger; D013301:Streptolysins; C025964:streptolysin O; C099100:SDF4 protein, human; D000681:Amylases; D002118:Calcium