Localization of Mitochondrial 60-kD Heat Shock Chaperonin Protein (Hsp60) in Pituitary Growth Hormone Secretory Granules and Pancreatic Zymogen Granules

Abstract

SUMMARYWe used quantitative immunogold electron microscopy and biochemical analysis to evaluate the subcellular distribution of Hsp60 in rat tissues. Western blot analysis, employing both monoclonal and polyclonal antibodies raised against mammalian Hsp60, shows that only a single 60-kD protein is reactive with the antibodies in brain, heart, kidney, liver, pancreas, pituitary, spleen, skeletal muscle, and adrenal gland. Immunogold labeling of tissues embedded in the acrylic resin LR Gold shows strong labeling of mitochondria in all tissues. However, in the anterior pitutary and in pancreatic acinar cells, Hsp60 also localizes in secretory granules. The labeled granules in the pituitary and pancreas were determined to be growth hormone granules and zymogen granules, respectively, using antibodies to growth hormone and carboxypeptidase A. Immunogold labeling of Hsp60 in all compartments was prevented by preadsorption of the antibodies with recombinant Hsp60. Biochemically purified zymogen granules free of mitochondrial contamination are shown by Western blot analysis to contain Hsp60, confirming the morphological localization results in pancreatic acinar cells. In kidney distal tubule cells, low Hsp60 reactivity is associated with infoldings of the basal plasma membrane. In comparison, the plasma membrane in kidney proximal tubule cells and in other tissues examined showed only background labeling. These findings raise interesting questions concerning translocation mechanisms and the cellular roles of Hsp60.

Effects of cadmium chloride on some mitochondria-related activity and gene expression of human MDA-MB231 breast tumor cells

“…The over-expression of hsp chaperones can depend on Cd-promoted denaturation and oxidation of proteins, and is probably linked to the increase of oxidative stress. Although hsp60 is mostly localized in the mitochondrial matrix, it has also been found in other subcellular localizations including the ER, cell surface, and unidentified vesicles and cytoplasmic granules [57] and also found to be over-expressed and localized in the cytoplasm of cancer cells [58]. In the current study, we report the presence of hsp60 in the cytoplasm of MDA-MB231 tumor cells and demonstrate that CdCl 2 exposure modifies the cytosolic level of the protein.…”

Regulation of HSP60 and the role of MK2 in a new model of severe experimental pancreatitis

“…We and others have shown that overexpression and phosphorylation of HSP27 and induction of HSP70 in pancreatic acinar cells protect against cerulein-induced pancreatitis (2,12). It has been found that cellular organelles like mitochondria in acinar cells express HSP60 (5,13). Along the synthesis and secretory pathway of pancreatic enzymes in acinar cells, there is an intimate coincidence of HSP60 with pancreatic lipase, amylase, and trypsin confirmed by immunoprecipitation assays; i.e., when HSP60 antibody is added to the isolated zymogen granule extracts, coprecipitation of HSP60 with digestive enzymes occurs (13,18).…”

Alteration of Cpn60 expression in pancreatic tissue of rats with acute pancreatitis

“…It not only exists in the mitochondria of pancreatic acinar cells, but also presents in the rough endoplasmic reticulum (RER), in Golgi body (G), in zymogen granules (ZG), and in the secretory pathway of pancreatic enzymes in acinar cells (Li et al 2003;Cechetto et al 2000). It is thought that Cpn60 residing in the mitochondria plays a different role from that of the protein located in the RER-G-ZG Keskin et al 2002).…”