Regulation of acetyl-CoA carboxylase in rat mammary gland. Effects of incubation with Ca2+, Mg2+ and ATP on enzyme activity in tissue extracts.
1. The effect of preincubation of extracts of lactating rat mammary gland with ATP, Mg2+ and micromolar concentrations of Ca2+ on the activity of acetyl-CoA carboxylase was studied. 2. Both Mg2+ and Ca2+ activated the enzyme. Activation with Mg2+ (5 mM) was larger than that with Ca2+ (calculated free Ca2+ concentration = 20-50 microM), but the activity decreased after reaching a peak. The activation obtained with Ca2+ was stable for up to 180 min. 3. Incubation with Ca2+ and Mg2+ together resulted in an activation that was slightly higher than that with Mg2+ only and was stable (compare the results for Ca2+ alone). 4. Preincubation in the absence of Mg2+, but not in the absence of Ca2+, resulted in the impairment of subsequent activation with either Mg2+ (when preincubation was with Ca2+ alone) or Mg2+ plus Ca2+. 5. KF (50 mM) prevented the activation of acetyl-CoA carboxylase by Ca2+ and Mg2+. 6. MgATP2- reversed (Mg2+ + Ca2+)-mediated activation and decreased the activity of acetyl-CoA carboxylase to about 10% of initial activity. Inhibition by ATP was unaffected by addition of cyclic AMP or cyclic AMP-dependent protein kinase inhibitor. 7. 32P was incorporated into acetyl-CoA carboxylase when incubations were carried out in the presence of [gamma-32P]ATP. Subsequent removal of ATP from the incubation medium resulted in rapid loss of 32P from acetyl-CoA carboxylase. 8. It is suggested that extracts of rat mammary gland contain endogenous protein kinase and phosphatase activities that modulate acetyl-CoA carboxylase activity through reversible phosphorylation and dephosphorylation. The phosphatase activity is sensitive to both Mg2+ and micromolar concentrations of Ca2+, whereas the kinase does not appear to be cyclic AMP-dependent.
D000103:Acetyl-CoA Carboxylase / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D002951:Citrates / Q000494:pharmacology; D019343:Citric Acid; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D005459:Fluorides; D066298:In Vitro Techniques; D007774:Lactation; D008025:Ligases / Q000378:metabolism*; D008274:Magnesium / Q000494:pharmacology*; D008321:Mammary Glands, Animal / Q000187:drug effects / Q000201:enzymology*; D011188:Potassium / Q000494:pharmacology; D017680:Potassium Compounds; D011247:Pregnancy; D051381:Rats; D011919:Rats, Inbred Strains
D002951:Citrates; D017680:Potassium Compounds; D019343:Citric Acid; D000255:Adenosine Triphosphate; C033320:potassium fluoride; D008025:Ligases; D000103:Acetyl-CoA Carboxylase; D008274:Magnesium; D005459:Fluorides; D011188:Potassium; D002118:Calcium