The presence in a mouse T cell line of a 97-kDa protein kinase C (PKC) with characteristics similar to known members of the novel PKC subgroup and its possible role in lymphocyte gene expression.
The receptor for tumor-promoting phorbol esters has been shown to be the Ca+2/phospholipid dependent enzyme protein kinase C (PKC). There are two major groups of PKC, the conventional PKC isotypes alpha, beta I, beta II, gamma) and the novel Ca+2-independent PKC (delta, epsilon, zeta, eta). Phorbol esters previously have been demonstrated to increase human IFN-gamma gene expression after treatment of a murine T cell line (Cl 9) that has been transfected with human IFN-gamma genomic DNA. In contrast, treatment with Ca+2 ionophore alone or in combination with phorbol ester did not enhance IFN-gamma production in a synergistic manner above the level obtained with phorbol ester treatment alone. To determine whether the lack of effect of Ca+2 ionophore is due to a defect in PKC, we compared the level of PKC autophosphorylation in the mouse T cell line (Cl 9), a mouse epidermal cell line (JB6), and purified rat brain PKC by in vitro kinase assays. The results demonstrate that instead of the expected 80-kDa autophosphorylated PKC band seen in purified rat brain PKC or mouse JB6 cell lysates, only a novel 97-kDa Ca+2-independent phosphoprotein was observed in Cl 9 cells. To ascertain if there was any nucleic acid sequence similarity to PKC epsilon, we hybridized Cl 9 poly(A+) RNA with a cloned fragment of the PKC epsilon gene and observed two hybridizing RNA bands (4.4 and 4.0 kb). Our results suggest that the 97-kDa phosphoprotein is similar to, but not identical with, PKC epsilon and is the major PKC expressed in the Cl 9 murine T cell line. These data suggested that the 97-kDa PKC may be responsible for the induction of both the transfected human IFN-gamma gene and the endogenous murine IL-2R alpha-chain.
Simek
S L
SL
Laboratory of Viral Carcinogenesis, NCI-FCRDC, Frederick, MD 21702-1201.
Fields
A P
AP
Colburn
N H
NH
Winkler-Pickett
R
R
Young
H A
HA
D000818:Animals; D002478:Cells, Cultured; D015870:Gene Expression; D066298:In Vitro Techniques; D007371:Interferon-gamma / Q000502:physiology; D007527:Isoenzymes / Q000502:physiology; D051379:Mice; D010703:Phorbol Esters / Q000494:pharmacology; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000235:genetics; D015375:Receptors, Interleukin-2 / Q000502:physiology; D013379:Substrate Specificity; D013601:T-Lymphocytes / Q000502:physiology*
D007527:Isoenzymes; D010703:Phorbol Esters; D010750:Phosphoproteins; D012333:RNA, Messenger; D015375:Receptors, Interleukin-2; D007371:Interferon-gamma; D011493:Protein Kinase C