Cholinephosphotransferase activities in microsomes and neuronal nuclei isolated from immature rabbit cerebral cortex: the use of endogenously generated diacylglycerols as substrate.
A neuronal nuclear fraction (N1) and a microsomal fraction (P3) were isolated from homogenates of cerebral cortices of 15-day-old rabbits. A nuclear envelope fraction (E) was prepared from N1. To assay cholinephosphotransferase, diacylglycerols were first generated in the membranes of these subfractions using a phospholipase C (Bacillus cereus) preincubation. With levels of endogenous diacylglycerols producing maximal specific cholinephosphotransferase activities, an activity ratio of 1:1:5 was found for N1, P3, and E, respectively. An independent neuronal nuclear cholinephosphotransferase, concentrated in nuclear membranes, is indicated. With regard to changes in pH and concentrations of MgCl2 and CDP-choline, N1 and P3 activities responded in a similar manner. However, in contrast to P3, N1 activities we much more profoundly inhibited at low levels of Triton X-100 (0.01-0.02 w/v%) and N1 showed quite significant levels of cholinephosphotransferase activity in the absence of a phospholipase C preincubation. Choline phosphotransferase in N1 and P3 showed Km values for CDP-choline (0.028 and 0.031 mM, respectively) which were much lower than corresponding literature values determined using exogenous diacylglycerols as substrates for this enzyme. The presence of cholinephosphotransferase in neuronal nuclear membranes reflects a rather exceptional nuclear autonomy. This may be related to a need to maintain nuclear phospholipid in the absence of a well-developed endoplasmic reticulum at early stages of neuronal development or to synthesize phospholipid in response to functions unique to the nucleus.
D000818:Animals; D002467:Cell Nucleus / Q000201:enzymology; D002540:Cerebral Cortex / Q000166:cytology / Q000201:enzymology*; D055598:Chemical Phenomena; D002621:Chemistry; D002798:Diacylglycerol Cholinephosphotransferase / Q000378:metabolism*; D004075:Diglycerides; D005227:Fatty Acids; D006863:Hydrogen-Ion Concentration; D008861:Microsomes / Q000201:enzymology; D009474:Neurons / Q000201:enzymology; D009685:Nuclear Envelope / Q000201:enzymology; D010770:Phosphotransferases / Q000378:metabolism*; D011817:Rabbits
C011439:1,2-diacylglycerol; D004075:Diglycerides; D005227:Fatty Acids; D010770:Phosphotransferases; D002798:Diacylglycerol Cholinephosphotransferase