Interaction of the insulin receptor with the receptor-like protein tyrosine phosphatases PTPalpha and PTPepsilon in living cells.
The interactions between the insulin receptor and the two highly homologous receptor-like protein tyrosine phosphatases (PTPase) PTPalpha and PTPepsilon were studied in living cells by using bioluminescence resonance energy transfer. In human embryonic kidney 293 cells expressing the insulin receptor fused to luciferase and substrate-trapping mutants of PTPalpha or PTPepsilon fused to the fluorescent protein Topaz, insulin induces an increase in resonance energy transfer that could be followed in real time in living cells. Insulin effect could be detected at very early time points and was maximal less than 2 min after insulin addition. Bioluminescence resonance energy-transfer saturation experiments indicate that insulin does not stimulate the recruitment of protein tyrosine phosphatase molecules to the insulin receptor but rather induces conformational changes within preassociated insulin receptor/protein tyrosine phosphatase complexes. Physical preassociation of the insulin receptor with these protein tyrosine phosphatases at the plasma membrane, in the absence of insulin, was also demonstrated by chemical cross-linking with a non-cell-permeable agent. These data provide the first evidence that PTPalpha and PTPepsilon associate with the insulin receptor in the basal state and suggest that these protein tyrosine phosphatases may constitute important negative regulators of the insulin receptor tyrosine kinase activity by acting rapidly at the plasma membrane level.
Lacasa
Danièle
D
Department of Cell Biology, Institut Cochin, CNRS/UMR 8104, INSERM U567, Université Paris V, 22 Rue Méchain, 75014 Paris, France.
Boute
Nicolas
N
Issad
Tarik
T
D002478:Cells, Cultured; D006801:Humans; D007328:Insulin / Q000494:pharmacology; D008163:Luminescent Measurements; D011487:Protein Conformation; D017027:Protein Tyrosine Phosphatases / Q000737:chemistry / Q000378:metabolism*; D011972:Receptor, Insulin / Q000737:chemistry / Q000378:metabolism*; D054630:Receptor-Like Protein Tyrosine Phosphatases, Class 4
D007328:Insulin; D011972:Receptor, Insulin; C516841:PTPRA protein, human; D017027:Protein Tyrosine Phosphatases; D054630:Receptor-Like Protein Tyrosine Phosphatases, Class 4