Characterization of a 58 kDa cis-Golgi protein in pancreatic exocrine cells.
We have studied the biochemical characteristics and localization of a 58 kDa cis-Golgi marker protein (p58) in rat pancreatic exocrine cells. The protein remained associated with membranes after extraction at alkaline pH and was largely resistant to proteases, added to intact microsomes. By electrophoresis p58 could be resolved into two bands which in two-dimensional gels separated into several charge variants around pI 5.5. This size and charge heterogeneity of p58 did not appear to be due to acylation, glycosylation or phosphorylation. In non-reduced gels p58 migrated as two kinetically related, high relative molecular mass forms, apparently corresponding to disulfide-linked homo-dimers and -hexamers. Immuno-electron microscopy localized p58 to both the fenestrated cis-Golgi cisternae and small Golgi vesicles or buds as well as large, pleiomorphic structures, scattered throughout the cells and associated with distinct smooth ER (endoplasmic reticulum) clusters. These findings correlated with cell fractionation results showing the concentration of p58 in two microsomal subfractions, banding at intermediate densities between the rough ER and trans-Golgi in sucrose gradients. Our results indicate that p58 is a major component of pre- and cis-Golgi elements and could be part of the transport machinery that operates in these membranes. Together with results obtained with other cell types, these observations suggest that the peripheral smooth ER clusters are involved in the early stages of the secretory pathway in the pancreatic acinar cells.
Lahtinen
U
U
Ludwig Institute for Cancer Research, Stockholm Branch, Sweden.
Dahllöf
B
B
Saraste
J
J
D000818:Animals; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D016253:Microscopy, Immunoelectron; D008861:Microsomes / Q000378:metabolism; D008970:Molecular Weight; D010179:Pancreas / Q000378:metabolism* / Q000648:ultrastructure; D011487:Protein Conformation; D051381:Rats; D013347:Subcellular Fractions / Q000378:metabolism; D014407:Tumor Cells, Cultured / Q000378:metabolism
D008565:Membrane Proteins