The Src family kinase Yes triggers hyperosmotic activation of the epidermal growth factor receptor and CD95.

Roland Reinehr,Dieter Haüssinger,Andrea Höngen,Stephan Becker

doi:10.1074/jbc.m401519200

Abstract

Hyperosmotic exposure of rat hepatocytes triggers epidermal growth factor receptor (EGFR) activation, which results in an activation of the CD95 system and sensitizes the cells toward apoptosis (Reinehr, R., Schliess, F., and Haüssinger, D. (2003) FASEB J. 17, 731-733). The mechanisms underlying the hyperosmotic EGFR activation were studied. Hyperosmotic exposure (405 mosm) resulted in a rapid activation of the Src kinase family members Yes, Fyn, and Lck. Hyperosmotic Yes, but not Fyn activation, was antioxidant-sensitive and was followed by a rapid Yes/EGFR association. PP-2 abolished the hyperosmotic activation of Fyn and Lck but not activation of Yes and EGFR and their association. However, these latter processes were prevented in the presence of SU6656. SU6656 and antioxidants, but not PP-2 and AG1478, also inhibited the hyperosmotic JNK activation. Cyclic AMP had no effect on hyperosmotic Yes and JNK activation but prevented EGFR/Yes association and EGFR activation in an H89-sensitive way. When the hyperosmolarity-induced Yes-EGFR protein complex started to disappear after 30 min, an association between EGFR and CD95 became apparent, which was followed by CD95 tyrosine phosphorylation and activation. SU6656 but not PP-2 also inhibited EGFR/CD95 association, CD95 tyrosine phosphorylation, CD95 membrane trafficking, and death-inducing signaling complex (DISC) formation. EGFR knockdown had no effect on hyperosmotic Yes activation but prevented CD95 tyrosine phosphorylation, membrane targeting, and DISC formation. Hyperosmotic EGFR and CD95 activation was also largely blunted following Yes knockdown. The data suggest that hyperosmotic signaling triggers an oxidative stress-dependent Yes activation, which is followed by JNK and EGFR activation and subsequent activation of the CD95 system. However, the functional relevance of hyperosmolarity-induced Fyn and Lck activation remains to be elucidated.

Highlights

Changes in liver cell hydration are important regulators of hepatic metabolism, gene expression, and transport across the plasma membrane through activation of osmosensing and osmosignaling pathways [1,2,3,4,5,6]
Apart from effects on metabolism and gene expression [1,2,3,4,5,6], hyperosmotic hepatocyte shrinkage triggers a rapid translocation of intracellular CD95 to the plasma membrane, which is accompanied by DISC1 formation and sensitizes hepatocytes toward CD95 ligand (CD95L)-induced apoptosis [9, 10]
In line with previous studies on inhibitor sensitivities of Src kinase family members, hyperosmotic Yes activation was largely abolished in the presence of SU6656 [22] but was insensitive to inhibition by PP-2 [23], herbimycin A [24], or emodin [25] (Fig. 1A)

Summary

Introduction

Changes in liver cell hydration are important regulators of hepatic metabolism, gene expression, and transport across the plasma membrane through activation of osmosensing and osmosignaling pathways [1,2,3,4,5,6]. Hyperosmotic Activation of the EGFR Is Triggered by Src Family Kinase Yes in Rat Hepatocytes—Hyperosmotic exposure of 24-h-cultured rat hepatocytes was shown to induce within 1 min a genistein-sensitive activation of the EGFR in rat hepatocytes [10] and Huh7 cells [21] (see Fig. 1D).

Results

Conclusion