A submembranous matrix of proteoglycans on zymogen granule membranes is involved in granule formation in rat pancreatic acinar cells.
The secretory lectin ZG16p mediated the binding of aggregated zymogens to the granule membrane in pancreatic acinar cells. Using a recently established in vitro condensation-sorting assay, we now show that pretreatment of zymogen granule membranes (ZGM) with either sodium bicarbonate at pH 10 or with phosphatidyl inositol-specific phospholipase C (PI-PLC) reduced the binding efficiency of zymogens to the same extent, as distinct components were liberated from ZGM. Analysis of the composition of the bicarbonate extract revealed the presence of the secretory lectin ZG16p, the serpin ZG46p and the GPI-linked glycoprotein GP-2, together with several unknown proteins, and small amounts of lipase and carboxylester lipase. The unknown proteins detected in 2-D gels represented a group of acidic and basic protein spots, which were positive in a glycan staining reaction and were soluble in methanol. One protein spot of the acidic group and several of the basic group reacted with a monoclonal antibody directed against chondroitin sulfate, indicating that the proteins represented proteoglycans. A staining pattern similar to the glycan reaction was observed in immunoblots using a polyclonal antibody directed against the whole bicarbonate extract. Immunogold electron microscopy revealed that this antibody reacted with components in the periphery of zymogen granules and strongly stained ZGM in the pellet fraction of a standard in vitro condensation-sorting assay. The amino acid composition of isolated components of both the acidic and basic group showed similarities to aggrecan, a cartilage-specific proteoglycan, and to glycine-rich glycoproteins, respectively. We therefore conclude that a submembranous matrix on the ZGM composed of proteoglycans and glycoproteins is involved in granule formation in pancreatic acinar cells.
Schmidt
K
K
Institut für Zytobiologie und Zytopathologie, Philipps Universität, Robert-Koch-Str. 5, D-35033 Marburg, Germany.
Dartsch
H
H
Linder
D
D
Kern
H F
HF
Kleene
R
R
D000818:Animals; D003594:Cytoplasmic Granules / Q000378:metabolism* / Q000648:ultrastructure; D004792:Enzyme Precursors / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism*; D008297:Male; D008854:Microscopy, Electron; D010179:Pancreas / Q000378:metabolism* / Q000648:ultrastructure*; D011509:Proteoglycans / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar
D004792:Enzyme Precursors; D011509:Proteoglycans