Novel insulin-elicited phosphoproteins in adipocytes

Cell Signal. 2005 Jan;17(1):59-66. doi: 10.1016/j.cellsig.2004.05.013.

Abstract

Akt is a key insulin-activated protein kinase. We searched for Akt substrates in 3T3-L1 adipocytes by means of immunoprecipitation with an Akt phosphomotif-specific antibody (PAS antibody). Four insulin-elicited phosphoproteins were isolated and identified by mass spectrometry. The identity of each protein was established by isolating the protein from lysates of untreated and insulin-treated adipocytes with an antibody specific for the protein and showing that the PAS antibody reacted only with the protein in the immunoprecipitate from insulin-treated cells. These proteins have sizes of 47, 75, 105, and 250 kDa on SDS PAGE, and have been designated pp47, 75, 105, and 250. The effect of inhibitors on the phosphorylation of the proteins, the identified sites of phosphorylation, and in vitro phosphorylation by recombinant Akt further indicated that pp47, 105, and 250 are likely to be Akt substrates, whereas pp75 may not be. pp47 and 105 are novel proteins with no known or predicted function. pp75 was previously found as a protein that associated with the colony-stimulating factor receptor, designated as Fms-interacting protein. pp250 is a novel protein with a predicted GTPase activating protein (GAP) domain for Rheb and/or Rap at its carboxy terminus. The subcellular and tissue distributions of the four proteins were determined.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • 3T3 Cells
  • Adipocytes / drug effects
  • Adipocytes / metabolism*
  • Amino Acid Sequence
  • Animals
  • Binding Sites
  • Carboxylic Ester Hydrolases
  • Insulin / pharmacology*
  • Membrane Proteins
  • Mice
  • Molecular Sequence Data
  • Peptide Fragments / chemistry
  • Phosphoproteins / chemistry
  • Phosphoproteins / drug effects
  • Phosphoproteins / metabolism*
  • Phosphorylation
  • Protein-Serine-Threonine Kinases / metabolism
  • Proto-Oncogene Proteins / metabolism
  • Proto-Oncogene Proteins c-akt

Substances

  • Insulin
  • Membrane Proteins
  • Peptide Fragments
  • Phosphoproteins
  • Proto-Oncogene Proteins
  • Protein-Serine-Threonine Kinases
  • Proto-Oncogene Proteins c-akt
  • Abhd15 protein, mouse
  • Carboxylic Ester Hydrolases