Effects of FK506-binding Protein 12 and FK506 on Autophosphorylation of Epidermal Growth Factor Receptor

Marco Lopez-Ilasaca,Cordelia Schiene,Gerhard Küllertz,Thomas Tradler,Gunter Fischer,Reinhard Wetzker

doi:10.1074/jbc.273.16.9430

Abstract

FK506-binding proteins and cyclophilins are intracellular proteins that express peptidylproline cis-trans-isomerase (PPIase) activity. The effects of FK506-binding protein 12 (FKBP12) and the cyclophilins 18 and 23 on autophosphorylation of the epidermal growth factor (EGF) receptor prepared from plasma membranes of the human epidermoid cell line A431 have been investigated. Whereas FKBP12 inhibited EGF receptor tyrosine kinase activity in a concentration-dependent manner, the cyclophilins did not affect autophosphorylation. In contrast to the wild-type enzyme, several variants of FKBP12 with greatly reduced PPIase activity were unable to suppress EGF receptor tyrosine kinase significantly. Pervanadate an inhibitor of protein tyrosine phosphatases, abolished the effect of FKBP12 on EGF receptor autophosphorylation. Finally, FK506 and rapamycin, which are known to block the PPIase activity of FKBP12, induced a significant stimulation of EGF receptor autophosphorylation in intact A431 cells suggesting suppression of EGF receptor autophosphorylation by intracellular FKBP12 in vivo. Taken together the data point to an inhibitory function of FKBP12 in EGF receptor signaling, possibly induced by stimulation of a protein tyrosine phosphatase coupled to the EGF receptor. Both PPIase activity and substrate specificity of FKBP12 seem to be indispensable for this effect.

Highlights

Peptidylproline cis-trans-isomerases (PPIases1; EC number 5.2.1.8) are ubiquitous and abundant enzymes conserved from prokaryotes to eukaryotes
FK506-binding protein 12 (FKBP12) Inhibits Autophosphorylation of epidermal growth factor (EGF) Receptor— The effect of different peptidylproline cis-transisomerase (PPIase) on EGF receptor tyrosine kinase prepared from A431 cells has been investigated
Changing the ATP concentration from 1 to 100 ␮M did not affect the ratio of the EGF receptor autophosphorylation in the absence of to that in the presence of 1.5 ␮M FKBP12

Summary

Introduction

Peptidylproline cis-trans-isomerases (PPIases; EC number 5.2.1.8) are ubiquitous and abundant enzymes conserved from prokaryotes to eukaryotes. In human T cells the cytosolic proteins Cyp and FKBP12, which are the archetypal PPIases, were shown to represent the primary targets of the immunosuppressive drugs cyclosporin A (CsA), rapamycin, and FK506. These microbial products were identified as tightly binding inhibitors of the enzymatic activity of their respective family of PPIases (6 – 8). FKBP binds to and modulates the function of calcium release channels [26, 27] and has been found complexed to the type I receptors of transforming growth factor-␤ [28] These pieces of evidence point to a more widespread role of FKBPs in intracellular signal transduction. We show that FKBP12 inhibits EGF receptor autophosphorylation in vitro and present evi-

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