A single residue in the C1 domain sensitizes novel protein kinase C isoforms to cellular diacylglycerol production.
The C1 domain mediates the diacylglycerol (DAG)-dependent translocation of conventional and novel protein kinase C (PKC) isoforms. In novel PKC isoforms (nPKCs), this domain binds membranes with sufficiently high affinity to recruit nPKCs to membranes in the absence of any other targeting mechanism. In conventional PKC (cPKC) isoforms, however, the affinity of the C1 domain for DAG is two orders of magnitude lower, necessitating the coordinated binding of the C1 domain and a Ca2+-regulated C2 domain for translocation and activation. Here we identify a single residue that tunes the affinity of the C1b domain for DAG- (but not phorbol ester-) containing membranes. This residue is invariant as Tyr in the C1b domain of cPKCs and invariant as Trp in all other PKC C1 domains. Binding studies using model membranes, as well as live cell imaging studies of yellow fluorescent protein-tagged C1 domains, reveal that Trp versus Tyr toggles the C1 domain between a species with sufficiently high affinity to respond to agonist-produced DAG to one that is unable to respond to physiological levels of DAG. In addition, we show that while Tyr at this switch position causes cytosolic localization of the C1 domain under unstimulated conditions, Trp targets these domains to the Golgi, likely due to basal levels of DAG at this region. Thus, Trp versus Tyr at this key position in the C1 domain controls both the membrane affinity and localization of PKC. The finding that a single residue controls the affinity of the C1 domain for DAG-containing membranes provides a molecular explanation for why 1) DAG alone is sufficient to activate nPKCs but not cPKCs and 2) nPKCs target to the Golgi.
Dries
Daniel R
DR
Biomedical Sciences Graduate Program and Department of Pharmacology, University of California at San Diego, La Jolla, California 92093, USA.
Gallegos
Lisa L
LL
Newton
Alexandra C
AC
D000818:Animals; D019556:COS Cells; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000201:enzymology; D002522:Chlorocebus aethiops; D017124:Conserved Sequence; D004075:Diglycerides / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D007527:Isoenzymes / Q000737:chemistry* / Q000378:metabolism*; D008968:Molecular Conformation; D011493:Protein Kinase C / Q000737:chemistry* / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D014364:Tryptophan / Q000737:chemistry / Q000378:metabolism; D014443:Tyrosine / Q000737:chemistry / Q000378:metabolism
D004075:Diglycerides; D007527:Isoenzymes; D014443:Tyrosine; D014364:Tryptophan; D011493:Protein Kinase C; D002118:Calcium