Regulation of non-classical protein kinase C isoenzymes in a human T cell line.
We have examined the expression and responses to activation, of novel/atypical protein kinase C (PKC) isoforms epsilon, zeta, and delta in the T cell lymphoma cell line K-4. The effects of 1-h phorbol 12-myristate 13-acetate (PMA) and OKT3 activation of K-4 cells on PKC isoform distribution were examined. In addition, the effects of PMA-mediated down-regulation on the expression of PKC epsilon and zeta were determined using high concentrations of PMA over 24- and 48-h time periods in these cells. PKC zeta expression was not altered by incubation of K-4 cells with up to 200 ng/ml PMA over a 24- or 48-h period. PKC epsilon was down-regulated in a concentration-dependent manner by PMA after both 24- and 48-h of activation. Expression of PKC epsilon was not completely depressed, however, even at the highest concentration of the phorbol ester after 48-h incubation with PMA. The presence of PKC epsilon, zeta, and delta was confirmed by immunohistochemistry with distinct patterns of expression observed. PMA-induced PKC activation for a 1-h period resulted in a translocation of PKC delta from resting cytoplasmic/nuclear staining to a cytoplasmic aggregate. Following 1-h activation through the T cell receptor-associated complex CD3, PKC delta translocated from a peri-nuclear/cytoplasmic compartment to a putative cytoskeletal location in K-4 cells. This translocation was time dependent and redistributed to a cytoplasmic aggregate prior to the cytoskeleton. Similarly, following 1-h activation through the T cell receptor, PKC zeta redistributed directly to what is possibly a cytoskeletal cell compartment. The cytoplasmic distribution of PKC zeta was unaltered following activation with PMA over a 1-h time period. There was no apparent redistribution of PKC epsilon cytoplasmic staining pattern following a 1-h direct or indirect activation. These results underline the differences in individual PKC isoform distribution, and responses to different stimuli, thereby providing additional evidence for the use of discrete PKC isoform signaling pathways in T cells. Furthermore, this data underlines the differences in PMA-mediated PKC activation and activation through the T cell receptor.
Keenan
C
C
Department of Clinical Medicine, Trinity College Dublin, St. James's Hospital, Ireland.
Kelleher
D
D
Long
A
A
D001692:Biological Transport; D017252:CD3 Complex / Q000276:immunology; D002460:Cell Line; D015536:Down-Regulation; D005455:Fluorescent Antibody Technique; D006801:Humans; D015151:Immunoblotting; D007527:Isoenzymes / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D013601:T-Lymphocytes / Q000201:enzymology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D017252:CD3 Complex; D007527:Isoenzymes; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate