In vivo and in vitro phosphorylation of the T lymphocyte type n (Kv1.3) potassium channel.
The major species of voltage-gated potassium channel found on mammalian T lymphocytes is referred to as the type n channel. This potassium channel exhibits unique functional properties which distinguish it from other species of potassium channels, including a potential role in the onset of cellular events associated with T cell activation. As a first step in characterizing specific biochemical properties of the type n channel, we have generated polyclonal antisera against bacterial fusion proteins containing peptide regions unique to the mouse and human type n channel. From membranes of T cell lines derived from both mouse (SAK 8 cell line) and human (Jurkat cell line), the type n channel can be immunoprecipitated following either surface labeling with 125I or metabolic labeling with 32P. The apparent molecular mass of the immunoprecipitated type n channel is approximately 65 kDa, significantly greater than that of the 58-kDa in vitro translated product, and suggestive of post-translational modification events. Phosphoamino acid analysis of the metabolically labeled Jurkat type n channel reveals phosphorylation of serine residues exclusively. In vitro studies also describe the ability of both protein kinase A and protein kinase C to phosphorylate the Jurkat type n channel. The former kinase also appears to phosphorylate a 40-kDa protein which co-immunoprecipitates with the type n channel. These data suggest that direct phosphorylation of the T lymphocyte type n potassium channel or its associated 40-kDa subunit may serve as a means by which channel activity is regulated.
Cai
Y C
YC
Vollum Institute, Oregon Health Sciences University, Portland 97201.
Douglass
J
J
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002462:Cell Membrane; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D006801:Humans; D007158:Immunologic Techniques; D066298:In Vitro Techniques; D051379:Mice; D008969:Molecular Sequence Data; D010766:Phosphorylation; D015221:Potassium Channels / Q000378:metabolism*; D011233:Precipitin Tests; D011493:Protein Kinase C / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000276:immunology; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013601:T-Lymphocytes / Q000378:metabolism*
D015221:Potassium Channels; D011993:Recombinant Fusion Proteins; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C