Immunocytochemical localization of prohormone convertases PC1/PC3 and PC2 in rat pancreatic islets.
The prohormone convertases PC1/PC3 and PC2 are endoproteases involved in prohormone cleavage at pairs of basic amino acids. To determine the cellular and subcellular distribution of PC1/PC3 and PC2 in the rat pancreas, we generated their polyclonal antisera in rabbits, using as immunogens two synthetic peptide antigens corresponding to amino acids 442-459 (ST-28) of PC1/PC3 and 613-629 (ST-29) of PC2 and two bacterially expressed antigens covering amino acids 145-414 (KN-1) of PC1/PC3 and 385-637 (KN-2) of PC2. Western blot analysis revealed the presence of PC1/PC3 (87 and 68 kDa) and PC2 (75 and 70 kDa) in rat pancreatic islets, indicating that the antisera are specific for the corresponding antigens. Immunocytochemical staining of serial sections demonstrated that the antibody against PC1/PC3 immunostained only insulin-producing cells, whereas the PC2 antibody stained insulin, glucagon-, somatostatin-, and pancreatic polypeptide-producing cells. Double-immunolabeling of the prohormone convertases and pancreatic hormones with gold particles of different sizes revealed that insulin-positive secretory granules were also immunolabeled with PC1/PC3 and PC2 antibodies, whereas glucagon-, somatostatin-, or pancreatic polypeptide-positive granules were labeled only with the PC2 antibody. This differential localization of PC1/PC3 and PC2 provides a further problem on the substrate-specificity of these enzymes in the processing of pancreatic prohormones.
Tanaka
S
S
Department of Cell Biology, Gunma University, Maebashi, Japan.
Kurabuchi
S
S
Mochida
H
H
Kato
T
T
Takahashi
S
S
Watanabe
T
T
Nakayama
K
K
D000595:Amino Acid Sequence; D000818:Animals; D000918:Antibody Specificity; D016282:Aspartic Acid Endopeptidases / Q000378:metabolism*; D015153:Blotting, Western; D007150:Immunohistochemistry; D007515:Islets of Langerhans / Q000201:enzymology*; D008297:Male; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D045706:Proprotein Convertase 2; D043484:Proprotein Convertases; D051381:Rats; D017208:Rats, Wistar; D013347:Subcellular Fractions / Q000201:enzymology; D013379:Substrate Specificity; D013381:Subtilisins / Q000378:metabolism*
D043484:Proprotein Convertases; D013381:Subtilisins; D045706:Proprotein Convertase 2; D016282:Aspartic Acid Endopeptidases