Monoclonal antibodies show that kinase C phosphorylation of GAP-43 during axonogenesis is both spatially and temporally restricted in vivo.
To study the role of kinase C phosphorylation in the distribution and function of GAP-43 we have generated a panel of mAbs that distinguish between GAP-43 that has been phosphorylated by kinase C and forms that have not. One class of antibodies, typified by 2G12/C7, reacts with only the phosphorylated form of GAP-43; it recognizes the peptide IQAS(PO4)FR equivalent to residues 38-43 that includes the single kinase C phosphorylation site at serine. Another, exemplified by 10E8/E7, reacts with both phosphorylated and nonphosphorylated forms. We have used the antibodies to study the distribution of kinase C-phosphorylated GAP-43 during axonogenesis and in the adult nervous system. Two major findings emerge. First, there is a lag between the initiation of axon outgrowth and the phosphorylation of GAP-43 by kinase C. The extent of this lag period varies between the different structures studied. In some cases, e.g., the trigeminal nerve, our result suggest that kinase C phosphorylation may be correlated with proximity of the growing axon to its target. Second, kinase C-phosphorylated GAP-43 is always spatially restricted to the distal axon. It is never seen either proximally or in cell bodies, even those with high levels of GAP-43 protein. This result also implies that GAP-43 is axonally transported in the non-kinase C phosphorylated form. Thus, kinase C phosphorylation of GAP-43 is not required for axon outgrowth or growth cone function per se and may be more related to interactions of the growth cone with its environment.
Meiri
K F
KF
Department of Pharmacology, State University of New York Health Science Center, Syracuse 13210.
Bickerstaff
L E
LE
Schwob
J E
JE
D000818:Animals; D000831:Animals, Newborn; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D001369:Axons / Q000378:metabolism* / Q000648:ultrastructure; D015153:Blotting, Western; D002490:Central Nervous System / Q000196:embryology / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D000939:Epitopes; D019922:GAP-43 Protein; D007150:Immunohistochemistry; D007700:Kinetics; D008562:Membrane Glycoproteins / Q000276:immunology / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000276:immunology / Q000378:metabolism*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000276:immunology / Q000378:metabolism*; D051381:Rats
D000911:Antibodies, Monoclonal; D000939:Epitopes; D019922:GAP-43 Protein; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; D011493:Protein Kinase C