Multisite phosphorylation of microtubule-associated protein 2 (MAP-2) in rat brain: peptide mapping distinguishes between cyclic AMP-, calcium/calmodulin-, and calcium/phospholipid-regulated phosphorylation mechanisms.
Microtubule-associated protein 2 (MAP-2), a cytoskeletal protein of 280 kilodalton that is highly enriched in dendrites and neuronal perikarya, is subject to both cyclic AMP-, calcium/calmodulin-, and calcium/phospholipid-regulated phosphorylation when incubated with [gamma-32P]ATP in vitro. We have analyzed the different sites in MAP-2 phosphorylated by these three kinases in fresh or boiled cytosol from different regions of the rat brain, in particular the olfactory bulb, where only one form (MAP-2B) is present, and the cerebral cortex, where both forms (MAP-2A and MAP-2B) are equally enriched. Cyclic AMP-dependent protein kinase and calcium/calmodulin-dependent protein kinase II phosphorylated four common phosphorylation sites, as well as a number of distinct sites that were unique to each enzyme. Calcium/phospholipid-dependent protein kinase phosphorylated a minimum of 15 sites, only one of which appeared to be shared with the other protein kinases. Only serine residues were phosphorylated by cyclic AMP-dependent and calcium/phospholipid-dependent protein kinases, while both serine and threonine residues were phosphorylated by calcium/calmodulin-dependent protein kinase II. No differences were observed in the peptide maps of phospho-MAP-2 prepared from different brain regions. These results emphasize the complexity of the phosphorylation systems that may regulate the function of MAP-2 in situ.
Walaas
S I
SI
Laboratory of Molecular and Cellular Neuroscience, Rockefeller University, New York, New York.
Nairn
A C
AC
D000255:Adenosine Triphosphate / Q000378:metabolism; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001921:Brain / Q000378:metabolism*; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D008869:Microtubule-Associated Proteins / Q000302:isolation & purification / Q000378:metabolism*; D009928:Organ Specificity; D010449:Peptide Mapping; D010748:Phosphopeptides / Q000302:isolation & purification; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D011494:Protein Kinases / Q000378:metabolism*; D051381:Rats
D000596:Amino Acids; D008869:Microtubule-Associated Proteins; D010748:Phosphopeptides; D000255:Adenosine Triphosphate; D011494:Protein Kinases; D011493:Protein Kinase C; D017871:Calcium-Calmodulin-Dependent Protein Kinases