Calcium-dependent phosphorylation of proteins in rabbit ciliary processes.
Calcium-dependent phosphorylation of endogenous substrate proteins in albino rabbit ciliary processes was studied by SDS-polyacrylamide gel electrophoresis and autoradiography. In the soluble fraction, a modest augmentation of phosphorylation was observed by Ca2+ alone and together with the additional activators, calmodulin (CAM) or phorbol myristate acetate (PMA). However, there was a greater enhancement of protein phosphorylation by Ca2+ and activators in the particulate fraction. The degree of Ca2+-CAM-dependent protein phosphorylation was greater than that of Ca2+-PMA-dependent phosphorylation. Endogenous substrate proteins for Ca2+-CAM-dependent protein kinases had apparent molecular sizes of 205,170,150,130,77,58,40,32 and 18 kDa. Phosphorylation of the 58 kDa protein band was strongest. This protein was identified as vimentin on the basis of its behavior with Triton-X100 treatment, and by Western blotting using anti-vimentin antibody. Endogenous substrates of protein kinase C (Ca2+-PMA-dependent) were located at 87 kDa and possibly in the 56 and 54 kDa protein bands. A 50 kDa protein was found to be phosphorylated in the presence of Ca2+ alone, and was not affected by the presence of other activators (CAM or PMA). A Ca2+-dependent dephosphorylation of a 43 kDa protein was observed, and some proteins rapidly phosphorylated by Ca2+-CAM kinase were also relatively quickly dephosphorylated at incubation times greater than 1 min.
Yoshimura
N
N
Department of Ophthalmology, Mount Sinai School of Medicine, CUNY, NY 10029.
Mittag
T W
TW
Podos
S M
SM
D000818:Animals; D002118:Calcium / Q000502:physiology*; D002147:Calmodulin / Q000502:physiology; D002924:Ciliary Body / Q000378:metabolism*; D005136:Eye Proteins / Q000378:metabolism*; D010766:Phosphorylation; D011817:Rabbits; D013347:Subcellular Fractions / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000502:physiology; D014746:Vimentin / Q000378:metabolism
D002147:Calmodulin; D005136:Eye Proteins; D014746:Vimentin; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium