RAT MAST-CELL PROTEASE-I ALTERS CELL-METABOLISM

Rat connective tissue mast cells are known to store significant amount s of mast cell protease I (RMCP I), which suppresses normal cell growt h and mediates cytotoxicity against tumor cell lines, including the fi brosarcoma cell line FL. To better define its effects on FL cells, RMC P I was added to FL cultures for 30 min. Analysis of de novo nuclear p rotein synthesis revealed that RMCP I suppressed the expression of thr ee proteins (41, 46, and 69 kD) and enhanced the expression of two oth er proteins (25 and 32 kD). Treatment of FL cells with diisopropylfluo rophosphate (DFP)-inactivated RMCP I proved that these effects were la rgely independent of the protease catalytic site. Western blot hybridi zation, using a monoclonal antibody to phosphotyrosine-containing prot eins, revealed that RMCP I inhibited phosphorylation of a nuclear and a cytoplasmic 81-kD tyrosylprotein. Inhibition of nuclear tyrosine kin ase activity by RMCP I appeared to be catalytic site dependent, wherea s cytoplasmic tyrosine kinase inhibition was independent of RMCP I pro teolytic activity. Biotinylated RMCP I was used to identify potential surface-binding proteins. Three specific binding complexes (130, 150, and 210 kD) were detected. The binding of biotinylated RMCP I to these surface proteins was inhibited by excess unlabeled RMCP I, but not by trypsin or chymotrypsin. We speculate that the binding proteins may b e critical in initiating RMCP I-induced metabolic changes on FL cells. The ability of RMCP I to alter the metabolism of cells suggests that it may have an important role in regulating their functions.