Calcium/calmodulin-regulated guanylate cyclase and calcium-permeability in the ciliary membrane from Tetrahymena.
A guanylate cyclase of high specific activity was localized in the ciliary membrane from Tetrahymena pyriformis. Purity of cilia was checked by electron microscopy and purity of membrane fractions isolated by a sucrose density gradient by sodium dodecyl sulfate/polyacrylamide gel electrophoresis. Enzyme activity was due to the presence of endogenous calmodulin as evidenced by the inhibition of guanylate cyclase by addition of antiserum against calmodulin from Tetrahymena or soybean. Removal of endogenous calmodulin by La3+-treatment of ciliary membranes resulted in loss of guanylate cyclase activity. In addition to protozoan calmodulins, the original activity could also be restored by the nonhomologous calmodulins from soybean and pig brain but not by calcium-binding proteins like Dictyostelium calmodulin, parvalbumin, and troponin C, lacking the trimethyllysine characteristic for mammalian calmodulins. However, only calmodulins from the protozoans Tetrahymena and Paramecium stimulated guanylate cyclase activity in excess of the initial activity. This indicates that the guanylate cyclase either contains two binding sites for calmodulin with different specificities or that a single, but only partially occupied binding site is modified possibly by hydrolytic exo-proteases during membrane preparation. The ciliary membrane from Tetrahymena contains a discrete calcium-permeability as demonstrated by calcium-flux measurements using the calcium indicator dye arsenazo III. In analogy to the excitable ciliary membrane of the larger relative Paramecium, the ciliary membrane of Tetrahymena may thus carry the voltage-sensitive calcium-channels known from electrophysiological studies.
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism*; D002923:Cilia / Q000201:enzymology / Q000378:metabolism / Q000648:ultrastructure; D006162:Guanylate Cyclase / Q000378:metabolism*; D008566:Membranes / Q000201:enzymology / Q000378:metabolism; D010539:Permeability; D013768:Tetrahymena / Q000201:enzymology / Q000378:metabolism* / Q000648:ultrastructure
D002147:Calmodulin; D006162:Guanylate Cyclase; D002118:Calcium