Calmodulin regulates fodrin susceptibility to cleavage by calcium-dependent protease I.
The intracellular calcium-dependent proteolysis of fodrin has been postulated to be central to the regulation of plasticity of the cortical cytoskeleton of many eukaryotic cells. The close proximity of the sites of calmodulin (CaM) binding and calcium-dependent protease I (CDP-I) cleavage in mammalian alpha-fodrin suggested that their action may be linked. In hypotonic and isotonic buffers, CDP-I proteolysis of the beta subunit of fodrin was absolutely dependent upon the presence of active CaM. The stimulation by CaM was inhibited by CaM antagonists. The rate of CDP-I proteolysis of both subunits was enhanced by CaM, while the rate of fodrin proteolysis with other proteases was not influenced by CaM. The increase in the susceptibility of fodrin to CDP-I proteolysis was half-maximal at 80 nM CaM, and maximal at 200 nM CaM. The unusual and differential susceptibility of alpha- and beta-fodrin to proteolysis by CDP-I in the absence of CaM was exploited to investigate the quaternary structure of fodrin in which only the alpha subunit was cleaved. Cleavage of the alpha subunit alone did not destroy the tetrameric form of the molecule, whereas CDP-I cleavage of both subunits rendered the molecule incapable of reforming tetramers. These results provide structural and functional evidence that CaM and CDP-I act synergistically in the regulated proteolysis of fodrin.
Harris
A S
AS
Department of Pathology, Yale University School of Medicine, New Haven, Connecticut 06510.
Croall
D E
DE
Morrow
J S
JS
D000818:Animals; D002147:Calmodulin / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D002154:Calpain / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism*; D002417:Cattle; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D046911:Macromolecular Substances; D008840:Microfilament Proteins / Q000378:metabolism*; D008970:Molecular Weight; D009994:Osmolar Concentration; D013449:Sulfonamides / Q000494:pharmacology; D013049:Spectrin
D002147:Calmodulin; D002154:Calpain; D002352:Carrier Proteins; D046911:Macromolecular Substances; D008840:Microfilament Proteins; D013449:Sulfonamides; D013049:Spectrin; C017967:W 7