Abstract
Macrophage colony stimulating factor (CSF‐1) and 1,25‐dihydroxyvitamin D~3~ (1,25(OH)~2~D~3~) are potent inducers of macrophage differentiation. Both appear to modulate protein phosphorylation, at least in part, through protein kinase C (PKC) raising the question as to whether they concurrently impact on macrophage‐like cells. In this regard, we utilized the CSF‐1 dependent murine macrophage‐like line BAC 1.25F5. CSF‐1 treatment of these cells for 30 min leads to particular phosphorylation of a 165 kDa protein, the putative CSF‐1 receptor, and a 210 kDa moiety. 1,25(OH)~2~D~3~ exposure for 24 h prior to addition of CSF‐1 enhances phosphorylation of the 165 kDa species and, especially, the 210 kDa protein. Phosphorylation of the latter protein is 1,25(OH)~2~D~3~ dose‐ and time‐dependent and the molecule is specifically immunoprecipitated with a rabbit polyclonal anti‐talin antibody. Experiments with okadaic acid show that the enhanced phosphorylation of talin does not result from serine phosphatase inhibition. CSF‐1 and 1,25(OH)~2~D~3~, alone or in combination, do not increase talin protein expression. The tyrosine kinase inhibitor, genestein, blocks 1,25(OH)~2~D~3~/CSF‐1 induced phosphorylation of the putative CSF‐1 receptor but has no effect on talin phosphorylation which occurs exclusively on serine. In contrast to genestein, staurosporin, an inhibitor of PKC, inhibits phosphorylation of talin. Moreover, exposure of 1,25(OH)~2~D~3~ pretreated cells to phorbol 12‐myristate 13‐acetate (PMA) in place of CSF‐1 also prompts talin phosphorylation. Finally, 1,25(OH)~2~D~3~ enhances ^3^[H]PDBu binding, indicating that the steroid increases PMA receptor capacity. Thus, CSF‐1 and 1,25(OH)~2~D~3~ act synergistically via PKC to phosphorylate talin, a cytoskeletal‐associated protein.