Stimulus-dependent phosphorylation of MacMARCKS, a protein kinase C substrate, in nerve termini and PC12 cells.
MacMARCKS (also known as myristoylated alanine-rich C kinase substrate (MARCKS)-related protein) is a member of the MARCKS family of protein kinase C substrates, which binds Ca2+/calmodulin in a phosphorylation-dependent manner. Immunoprecipitation demonstrated that MacMARCKS is present in both PC12 cells and in neurons. Upon depolarization of PC12 cells with 60 mM KCl, MacMARCKS phosphorylation increased 4-fold over basal levels in a Ca(2+)-dependent manner. By immunofluorescence microscopy, MacMARCKS was colocalized in PC12 cells to neurite tips with the synaptic vesicle membrane protein synaptophysin and to vesicles in the perinuclear region. Subcellular fractionation demonstrated that MacMARCKS associates tightly with membranes in PC12 cells. In Percoll-purified rat cerebrocortical synaptosomes, depolarization with 60 mM KCl in the presence of exogenous Ca2+ transiently increased MacMARCKS phosphorylation, whereas phorbol ester promoted a sustained increase in MacMARCKS phosphorylation. Subcellular fractionation of rat brain indicated that MacMARCKS was present in both soluble and particulate fractions; particulate MacMARCKS was associated with both small vesicles and highly purified synaptic vesicles. These results are consistent with a role for MacMARCKS in integrating Ca(2+)-calmodulin and protein kinase C-dependent signals in the regulation of neurosecretion.
Chang
S
S
Laboratory of Signal Transduction, Rockefeller University, New York, New York 10021-6399, USA.
Hemmings
H C
HC
Aderem
A
A
D000818:Animals; D002118:Calcium / Q000378:metabolism; D008565:Membrane Proteins; D009411:Nerve Endings / Q000378:metabolism*; D016716:PC12 Cells; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D013347:Subcellular Fractions / Q000378:metabolism; D033921:Vesicular Transport Proteins
C512942:Marcksl1 protein, rat; D008565:Membrane Proteins; D011506:Proteins; D033921:Vesicular Transport Proteins; D011493:Protein Kinase C; D002118:Calcium