SIGNALING PATHWAYS UNDERLYING EOSINOPHIL CELL MOTILITY REVEALED BY USING CAGED PEPTIDES

Insights into structure-function relations of many proteins opens the possibility of engineering peptides to selectively interfere with a pr otein's activity, To facilitate the use of peptides as probes of cellu lar processes, we have developed caged peptides whose influence on spe cific proteins can be suddenly and uniformly changed by near-UV fight. Two peptides are described which, on photolysis of a caging moiety, b lock the action of calcium-calmodulin or myosin light chain kinase (ML CK). The efficacy of theses peptides is demonstrated in vitro and in v ivo by determining their effect before and after photolysis on activit ies of isolated enzymes and cellular functions known to depend on calc ium-calmodulin and MLCK. These caged peptides each were injected into motile, polarized eosinophils, and when exposed to light promptly bloc ked cell locomotion in a similar manner. The results indicate that the action of calcium-calmodulin and MLCK and by inference myosin II, are required for the ameboid locomotion of these cells, This methodology provides a powerful means for assessing the role of these and other pr oteins in a wide range of spatio-temporally complex functions in intac t living cells.