Characterization of the phosphorylation of rat mammary ATP-citrate lyase and acetyl-CoA carboxylase by Ca2+ and calmodulin-dependent multiprotein kinase and Ca2+ and phospholipid-dependent protein kinase.
ATP-citrate lyase and acetyl-CoA carboxylase purified from lactating rat mammary gland are phosphorylated stoichiometrically by the calmodulin-dependent multiprotein kinase from rabbit skeletal muscle. The reactions are completely dependent on the presence of both Ca2+ and calmodulin. ATP-citrate lyase and acetyl-CoA carboxylase are also phosphorylated stoichiometrically by the Ca2+- and phospholipid-dependent protein kinase (protein kinase C) purified from bovine brain. Phosphorylation of these substrates is stimulated 6-fold and 40-fold respectively by Ca2+ and phosphatidylserine. The calmodulin-dependent and phospholipid-dependent protein kinases phosphorylate the same serine residue on ATP-citrate lyase that is phosphorylated by cyclic-AMP-dependent protein kinase. The sequence of the tryptic peptide containing this site on the mammary enzyme is identical with the sequence of the peptide containing the site on ATP-citrate lyase that is phosphorylated in isolated hepatocytes in response to insulin and/or glucagon. The calmodulin-dependent, phospholipid-dependent and cyclic-AMP-dependent protein kinases phosphorylate distinct sites on acetyl-CoA carboxylase. However, one of the three phosphorylated tryptic peptides derived from enzyme treated with the phospholipid-dependent kinase is identical with the major phosphopeptide (T1) derived from enzyme treated with cyclic-AMP-dependent protein kinase. Phosphorylation of acetyl-CoA carboxylase by the phospholipid-dependent protein kinase inactivates acetyl-CoA carboxylase in a similar manner to cyclic-AMP-dependent protein kinase. With either protein kinase slightly greater phosphorylation and inactivation is seen after pretreatment of acetyl-CoA carboxylase with protein phosphatase-2A, but the effects of the protein phosphatase treatment are not completely reversed. Inactivation by the phospholipid-dependent protein kinase is Ca2+- and phospholipid-dependent, is reversed by protein phosphatase-2A, and correlates with the degree of phosphorylation. The relevance of these findings to insulin- and growth-factor-promoted phosphorylation of ATP-citrate lyase and acetyl-CoA carboxylase in intact cells is discussed.
Hardie
D G
DG
Carling
D
D
Ferrari
S
S
Guy
P S
PS
Aitken
A
A
D001275:ATP Citrate (pro-S)-Lyase / Q000378:metabolism*; D000103:Acetyl-CoA Carboxylase / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001923:Brain Chemistry; D002118:Calcium / Q000494:pharmacology*; D002147:Calmodulin / Q000494:pharmacology*; D000242:Cyclic AMP / Q000494:pharmacology; D005260:Female; D007700:Kinetics; D007774:Lactation; D008025:Ligases / Q000378:metabolism*; D008321:Mammary Glands, Animal / Q000201:enzymology; D009132:Muscles / Q000201:enzymology; D010446:Peptide Fragments / Q000378:metabolism; D010743:Phospholipids / Q000494:pharmacology*; D010766:Phosphorylation; D011247:Pregnancy; D011494:Protein Kinases / Q000378:metabolism*; D011817:Rabbits; D051381:Rats; D012694:Serine / Q000378:metabolism; D012756:Sheep; D014357:Trypsin
D002147:Calmodulin; D010446:Peptide Fragments; D010743:Phospholipids; D012694:Serine; D000242:Cyclic AMP; D001275:ATP Citrate (pro-S)-Lyase; D011494:Protein Kinases; D014357:Trypsin; D008025:Ligases; D000103:Acetyl-CoA Carboxylase; D002118:Calcium