A comparative study of microsomal and cytosolic S6 phosphatase activities in rat liver.
Spontaneous S6 phosphatase activities dephosphorylating Ser(P)-235 and Ser(P)-236 of the ribosomal protein S6 were measured and compared in microsomes and cytosol of rat liver. The substrate used, small (40S) ribosomal subunits 32P-labelled in vitro by protein kinase A, contained phosphorylated S6 (mainly in the diphosphorylated form) and some minor phosphorylated species. The microsomal and cytosolic S6 phosphatase activities displayed a number of distinct properties. The microsomal activity, representing ca 20% of the S6 phosphatase activity in the post-mitochondrial supernatant, was mainly due to a type-1 phosphatase and dephosphorylated only S6. The remaining post-mitochondrial S6 phosphatase activity, which was fully recovered in the cytosol, and appeared to result from a combination of type-1 (43%) and type 2 (57%) phosphatases, acted on S6 as well as on the minor phosphorylated species. The microsomal activity was 50% inhibited by MgCl2 (10 mM) and was stimulated at least 4.3 fold by MnCl2 (1 mM), while the cytosolic activity was inhibited only 18% by Mg2+ (10 mM) and was increased 2.2 fold by Mn2+ (1 mM). The microsomal activity was increased 10% (P less than 0.06) by lower doses of insulin (25 U/Kg) and 14% (P less than 0.05) by vanadate, but was not significantly (P greater than 0.10) affected by larger doses of insulin (100 U/kg), hepatectomy or cycloheximide. By comparison the cytosolic S6 phosphatase activity was unresponsive to insulin and vanadate, but was decreased 14% and 17% (P less than 0.05) by hepatectomy and cycloheximide.(ABSTRACT TRUNCATED AT 250 WORDS)
Stephenson
G M
GM
Russell Grimwade School of Biochemistry, University of Melbourne, Parkville, Victoria, Australia.
D000818:Animals; D003513:Cycloheximide / Q000494:pharmacology; D003600:Cytosol / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D007328:Insulin / Q000494:pharmacology; D008099:Liver / Q000166:cytology / Q000201:enzymology*; D015636:Magnesium Chloride / Q000494:pharmacology; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism*; D010766:Phosphorylation; D051381:Rats; D038601:Ribosomal Protein S6; D012269:Ribosomal Proteins / Q000378:metabolism*; D014638:Vanadates / Q000494:pharmacology
D007328:Insulin; D038601:Ribosomal Protein S6; D012269:Ribosomal Proteins; D015636:Magnesium Chloride; D014638:Vanadates; D003513:Cycloheximide; D010744:Phosphoric Monoester Hydrolases