Identification and immunolocalization of phospholipase C in bovine rod outer segments.
Bovine rod outer segments (ROS) contain a phospholipase C (PLC) that hydrolyzes phosphatidylinositol 4,5-bisphosphate. Approximately 60-70% of PLC activity is recovered in soluble extracts of ROS. Moreover, the specific activity of this soluble PLC is approximately 10-fold higher than that of resealed ROS enzyme activity. Peptide-specific antiserum (Ab 1109) directed against a highly conserved sequence of the Y-region found in several PLC isozymes was used to detect any PLC belonging to this family. This antibody specifically recognized a protein of apparent molecular mass of approximately 140 kDa present in immunoblots of soluble extracts of both ROS and whole retina. The elution profile of this 140-kDa antigen from a Sephadex G-150 column coincided with the peak of PLC activity, suggesting PLC activity is associated with the 140-kDa protein. Immunocytochemical studies of bovine retina using Ab 1109 showed pronounced immunoreactive labeling in the photoreceptor layer. In resealed ROS and washed ROS membranes, Ab 1109 recognized an additional protein of apparent molecular mass of 70 kDa not usually detectable in soluble extracts of ROS, suggesting the presence of at least two isozymes of PLC in ROS.
Ghalayini
A J
AJ
Cullen Eye Institute, Baylor College of Medicine, Houston, TX 77030.
Tarver
A P
AP
Mackin
W M
WM
Koutz
C A
CA
Anderson
R E
RE
D000818:Animals; D015153:Blotting, Western; D002417:Cattle; D002850:Chromatography, Gel; D007150:Immunohistochemistry; D007527:Isoenzymes / Q000378:metabolism; D012160:Retina / Q000201:enzymology; D012374:Rod Cell Outer Segment / Q000201:enzymology*; D014018:Tissue Distribution; D010738:Type C Phospholipases / Q000378:metabolism*
D007527:Isoenzymes; D010738:Type C Phospholipases