Microinjection of a protein-tyrosine-phosphatase inhibits insulin action in Xenopus oocytes.
A protein-tyrosine-phosphatase (PTPase 1B; protein-tyrosine-phosphate phosphohydrolase, EC 3.1.3.48), specific for phosphotyrosyl residues, was microinjected into Xenopus oocytes. This resulted in a 3- to 5-fold increase in PTPase activity over endogenous levels. The PTPase blocked the insulin-stimulated phosphorylation of tyrosyl residues on endogenous proteins, including a protein having a molecular mass in the same range as the beta subunit of the insulin or insulin-like growth factor I receptor. PTPase 1B also blocked the activation of an S6 peptide kinase--i.e., an enzyme recognizing a peptide having the sequence RRLSSLRA found in a segment of ribosomal protein S6 and known to be activated early in response to insulin. On the other hand, the insulin stimulation of an S6 kinase, detected by using 40S ribosomes as substrate, was unaffected even though PTPase 1B partially prevented the phosphorylation of ribosomal protein S6 in vivo. Mono Q chromatography of insulin-treated oocyte extracts revealed two main peaks of S6 kinase activity. Fractions from the first peak displayed S6 peptide kinase activity that was essentially abolished in profiles from PTPase 1B-injected oocytes. Material from the second peak, which was best revealed by using 40S ribosomes as substrate and had comparatively little S6 peptide kinase activity, was minimally affected by PTPase 1B. These observations suggest that at least two distinct "S6 kinases" are involved in ribosomal protein S6 phosphorylation in vivo and that the activation pathways for these enzymes differ in their sensitivity to PTPase 1B.
Cicirelli
M F
MF
Howard Hughes Medical Institute, University of Washington School of Medicine, Seattle 98195.
Tonks
N K
NK
Diltz
C D
CD
Weiel
J E
JE
Fischer
E H
EH
Krebs
E G
EG
D000818:Animals; D005260:Female; D066298:In Vitro Techniques; D007328:Insulin / Q000494:pharmacology*; D007700:Kinetics; D008845:Microinjections; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism*; D010749:Phosphoprotein Phosphatases / Q000008:administration & dosage / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases; D019893:Ribosomal Protein S6 Kinases; D014443:Tyrosine; D014982:Xenopus laevis
D007328:Insulin; D014443:Tyrosine; D011494:Protein Kinases; D019893:Ribosomal Protein S6 Kinases; D010749:Phosphoprotein Phosphatases; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases