Membranes were isolated from B cells stimulated with phorbol 12-myristate 13-acetate (PMA) for a time sufficient to allow maximal redistribution and activation of protein kinase C (PKC). Exposure of such membranes to a short incubation with [γ-32P]ATP resulted in the detection of at least nine unique or hyperphosphorylated membrane proteins by SDS-PAGE and autoradiography. The appearance of these phosphoproteins was blocked by pretreatment of the membranes with H-7 or sangivamycin, two selective inhibitors of PKC. In addition, membranes purified from B cells treated with an inactive phorbol ester or stimulated with dibutyryl cAMP failed to exhibit a pattern of new phosphoproteins. These results are consistent with the involvement of PKC in the phosphorylation of the proteins. These phosphoproteins are also candidates for proteins whose functions are modified as a consequence of early signal delivery to resting B cells following membrane immunoglobulin occupancy. This system was utilized to identify the heavy chain of MHC class I molecules as one of the membrane proteins phosphorylated by PKC. The MHC class II molecules were not phosphorylated in membranes isolated from PMA-treated normal B cells or from PMA-treated B cells which had previously been exposed to IL-4. These results indicate that class I, but not class II, MHC molecules are phosphorylated by PKC. It is possible that such a modification of cell surface class I molecules may be involved during the process of signal transduction leading to B cell activation.
|Number of pages||10|
|State||Published - Dec 1989|
Bibliographical noteFunding Information:
*This work was supported by PHS grants AGO476101 and AI23907 from the National Institutes of Health, a grant from the UKMC Research Fund, and a Travel Award from the Burroughs Wellcome Fund. Abbreviations: DAG, diacylglycerol; EDTA, ethylene-diaminetetra-acetic acid, H-7, l-(%soquinolinesul-fonyl)-2-methylpiperazine; IP,, inositol triphosphate; mIg, membrane immunoglobulin; NP-40, Nonidet P-40; ccPDD. 4anhorbol 12,13-didecanoate: PI, _ ohosuhatidvl-. inositol; PIPES, piperazine-N,N’-bis(2-ethanesulfonic acid); PKC, protein kinase C; PMA, phorbol 12-myris-tate 13-acetate; SDS, sodium dodecyl sulfate.
ASJC Scopus subject areas
- Molecular Biology