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Phorbol Ester‐ and Retinoic Acid‐Induced Regulation of the Protein Kinase C Substrate MARCKS in Immortalized Hippocampal Cells
Author(s) -
Watson D. G.,
Wainer B. H.,
Lenox R. H.
Publication year - 1994
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.1994.63051666.x
Subject(s) - marcks , protein kinase c , retinoic acid , phorbol , chemistry , tretinoin , cell culture , microbiology and biotechnology , biology , biochemistry , phosphorylation , genetics
The expression of MARCKS, a major protein kinase C (PKC) substrate, was examined in the immortalized hippocampal cell line HN33, following differentiation using phorbol esters or retinoic acid. In cells exposed to phorbol esters, MARCKS protein levels were reduced through an apparent PKC‐dependent mechanism. Exposure to 1 µ M phorbol 12‐myristate 13‐acetate (PMA) for 10 min resulted in a rapid loss of PKC activity in the soluble fraction with a concurrent increase in membrane‐associated PKC activity. PKC activity was reduced to <20% of control values in both soluble and membrane fractions following 1 h of PMA exposure. Significant reductions in MARCKS protein levels were initially observed in membrane and soluble fractions following PMA exposure for 4 and 8 h, respectively. The reduction in MARCKS protein levels was maximal following 24 h of PMA exposure. MARCKS protein expression was also down‐regulated in a dose‐dependent manner on exposure of HN33 cells to retinoic acid. In cells exposed to 10 µ M retinoic acid, the MARCKS protein level was reduced in the membrane fraction within 4 h. Reduction of MARCKS protein levels was maximal (>90%) by 12 h with no evidence for any alteration in PKC activity. Reduced levels of MARCKS protein were also observed in the soluble fraction of retinoic acid‐exposed cells, but to a significantly lesser extent. Addition of the PKC inhibitor GF109203X blocked the down‐regulation of MARCKS protein in PMA‐treated cultures but not in retinoic acid‐treated cells. These findings suggest that the down‐regulation of MARCKS may play an important role in both phorbol ester‐ and retinoic acid‐induced differentiation in cells of neuronal origin.