Protein kinase C association with the retinal cytoskeleton and phosphorylation of vimentin.
Protein kinase C plays an important role in regulation of the cytoskeleton. We found that protein kinase C activity is associated with the retinal cytoskeleton, and that it catalyses the stoichiometric phosphorylation of two cytoskeletal proteins, with apparent molecular masses of 56 kDa and 100 kDa. The 56-kDa substrate was identified as vimentin on the basis of its apparent molecular mass, pI, solubility, immunoreactivity, pattern of proteolysis by Lys-C and a partial amino acid sequence. Immunomicroscopy was consistent with previous reports that in the retina vimentin has the unusual property of being present in neuronal cells--horizontal cells--as well as non-neuronal cells. The characteristics of protein kinase C phosphorylation of vimentin that was enriched in neuronal vimentin were determined. Hyperactivation of protein kinase C by treatment of retinas with phorbol myristate acetate resulted in the phosphorylation of vimentin in situ, indicating that the phosphorylation is physiologically relevant. In vitro, purified retinal protein kinase C catalysed the incorporation of nearly 2 mol phosphate per mole of monomeric vimentin. The phosphorylation was highly dependent on the presence of phosphatidylserine. Thus, protein kinase C functions in the retinal cytoskeleton, where a major role is in the modification of vimentin. The characterization of the phosphorylation of outer retinal vimentin by protein kinase C provides a basis for further studies on the regulation and function of this cytoskeletal element.
Williams
D S
DS
Laboratory of Cell Biology, School of Optometry, Indiana University, Bloomington 47405.
Park
S
S
Schlamp
C L
CL
Newton
A C
AC
D000818:Animals; D015153:Blotting, Western; D002154:Calpain / Q000494:pharmacology; D002417:Cattle; D003599:Cytoskeleton / Q000201:enzymology*; D007527:Isoenzymes / Q000032:analysis; D009475:Neurons, Afferent / Q000737:chemistry; D010766:Phosphorylation; D011493:Protein Kinase C / Q000032:analysis / Q000502:physiology*; D051381:Rats; D012160:Retina / Q000737:chemistry / Q000201:enzymology*; D013552:Swine; D014746:Vimentin / Q000032:analysis / Q000378:metabolism*
D007527:Isoenzymes; D014746:Vimentin; D011493:Protein Kinase C; D002154:Calpain