Studies on phosphoinositide metabolism in retinal rod outer segments
The present work was undertaken to determine whether isolated mammalian photoreceptors (bovine rod outer segments, ROS) were capable of synthesizing and hydrolyzing phosphoinositides, and if so, how synthesis and hydrolysis might be regulated. Synthesis of phosphoinositides was demonstrated by the incorporation of radioactively labeled precursors. ROS incubated with ({gamma}{sup 32}P)ATP produced labeled phosphatidic acid (PA), phosphatidylinositol-4-phosphate (PIP) and phosphatidylinositol-4,5-bisphosphate (PIP{sub 2}). When Mn{sup 2+}, CTP and inositol were added, labeled phosphatidylinositol (PI) was also produced. ({sup 3}H)Inositol was incorporated into PI, PIP and PIP{sub 2}, although prolonged incubation was required for detectable incorporation into PIP{sub 2}. Incorporation of ({sup 3}H)inositol was dependent on CTP, indicating that labeling proceeded via synthesis, not base exchange. Incubation with ({alpha}-{sup 32}P)CTP produced labeled CDP-diacylglycerol, an intermediate in PI synthesis. Incorporation of labeled precursors was stimulated by Mg{sup 2+}, Mn{sup 2+} and spermine, but unaffected by light. Phosphoinositide hydrolysis was measured using exogenous {sup 3}H-labeled substrates. PI, PIP, and PIP{sub 2} were all hydrolyzed but most attention was devoted to PIP{sub 2}. Crude enzyme preparations contained an endogenous inhibitor whose effects were Ca{sup 2+}-dependently relieved by calmodulin antagonists. This inhibitor does not appear to be calmodulin but may be a novel Ca{sup 2+}-binding regulatory protein. No evidence for regulation of PLC activity by light or G-proteins was obtained. The effects on PLC activity of Mg{sup 2+}, Mn{sup 2+}, spermine, pH and detergents were also characterized.
- Research Organization:
- Michigan State Univ., East Lansing, MI (USA)
- OSTI ID:
- 7067399
- Resource Relation:
- Other Information: Thesis (Ph.D)
- Country of Publication:
- United States
- Language:
- English
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PHOSPHOLIPIDS
BIOLOGICAL FUNCTIONS
CATTLE
EXPERIMENTAL DATA
INOSITOL
MAGNESIUM COMPOUNDS
MANGANESE COMPOUNDS
PHOSPHORUS 32
TRACER TECHNIQUES
ALKALINE EARTH METAL COMPOUNDS
ANIMALS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
CARBOHYDRATES
DATA
DAYS LIVING RADIOISOTOPES
DOMESTIC ANIMALS
ESTERS
FUNCTIONS
INFORMATION
INOSITOLS
ISOTOPE APPLICATIONS
ISOTOPES
LIGHT NUCLEI
LIPIDS
MAMMALS
MONOSACCHARIDES
NUCLEI
NUMERICAL DATA
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
ORGANIC PHOSPHORUS COMPOUNDS
PHOSPHORUS ISOTOPES
RADIOISOTOPES
RUMINANTS
SACCHARIDES
TRANSITION ELEMENT COMPOUNDS
VERTEBRATES
550201* - Biochemistry- Tracer Techniques