A new protein kinase C, nPKC eta', and nPKC theta are expressed in human platelets: involvement of nPKC eta' and nPKC theta in signal transduction stimulated by PAF.
We have detected in human platelets two protein kinase C isozymes that have not been reported previously. Using an anti-nPKC theta antibody and Western blotting, we calculated the molecular weight of platelet nPKC theta as 79K. This molecular weight is identical to that described for nPKC theta in skeletal muscle and in COS cells transfected with the nPKC theta-cDNA. Using an anti-nPKC eta antibody, we determined the molecular weight of an immunoreactive protein, which we called nPKC eta', to be 95K. This molecular weight is higher than that of nPKC eta found in lung and skin tissue of 82K and 78K, and it is higher than nPKC eta of COS cells transfected with the nPKC eta-cDNA expression plasmid. Together with previous reports, these findings make the total number of PKC isozymes in human platelets equal to six. These are the PKC isozymes: alpha, beta, delta and zeta, which have been previously described, and eta' and theta which we describe here. To assess the functionality of these new PKC isoforms, we stimulated platelets with PAF. We found a 200% and 175% increase in the levels of membrane-bound nPKC eta' and nPKC theta, respectively, in human platelets stimulated by PAF. A concomitant decrease in the level of these isoforms in the cytoplasm was observed. This PAF-induced translocation was time-dependent, and it reached its peak after a 1 minute incubation of human platelets with PAF for nPKC theta and 30 seconds for nPKC eta'.
Wang
F
F
Dept. of Anatomy/Cell Biology, SUNY Health Science Center, Brooklyn 11203.
Naik
U P
UP
Ehrlich
Y H
YH
Freyberg
Z
Z
Osada
S
S
Ohno
S
S
Kuroki
T
T
Suzuki
K
K
Kornecki
E
E
D000595:Amino Acid Sequence; D000906:Antibodies; D000911:Antibodies, Monoclonal; D001792:Blood Platelets / Q000187:drug effects / Q000201:enzymology* / Q000502:physiology; D015153:Blotting, Western; D002462:Cell Membrane / Q000201:enzymology; D003600:Cytosol / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000097:blood* / Q000302:isolation & purification; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology; D010972:Platelet Activating Factor / Q000494:pharmacology*; D011493:Protein Kinase C / Q000097:blood* / Q000302:isolation & purification; D015398:Signal Transduction / Q000187:drug effects*; D013997:Time Factors
D000906:Antibodies; D000911:Antibodies, Monoclonal; D007527:Isoenzymes; D010455:Peptides; D010972:Platelet Activating Factor; D011493:Protein Kinase C