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Bag1–Hsp70 mediates a physiological stress signalling pathway that regulates Raf-1/ERK and cell growth

Abstract

Survival after stress requires the precise orchestration of cell-signalling events to ensure that biosynthetic processes are alerted and cell survival pathways are initiated. Here we show that Bag1, a co-chaperone for heat-shock protein 70 (Hsp70), coordinates signals for cell growth in response to cell stress, by downregulating the activity of Raf-1 kinase. Raf-1 and Hsp70 compete for binding to Bag1, such that Bag1 binds to and activates Raf-1, subsequently activating the downstream extracellular signal-related kinases (ERKs). When levels of Hsp70 are elevated after heat shock, or in cells conditionally overexpressing Hsp70, Bag1–Raf-1 is displaced by Bag1–Hsp70, and DNA synthesis is arrested. Mutants Bag1C204A and Bag1E208A, which cannot bind Hsp70, constitutively activate Raf-1/ERK kinases but are unaffected by Hsp70; consequently neither Bag1–Raf-1 nor DNA synthesis is negatively affected during heat shock. Likewise, mutants Hsp70F245S, Hsp70R262W and Hsp70L282R, which retain chaperone activity but do not bind to Bag1, fail to repress Bag1 activation of Raf-1/ERK kinase. We propose that Bag1 functions in the heat-shock response to coordinate cell growth signals and mitogenesis, and that Hsp70 functions as a sensor in stress signalling.

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Figure 1: Identification of the Bag1 regions required for interaction with Hsp70 and Raf-1.
Figure 2: Raf-1 and Hsp70 compete for binding to Bag1.
Figure 3: Bag1 activates Raf-1 independent of Ras.
Figure 4: In vivo effect of Bag1 on the Raf-1/ERK kinase pathway, and negative effect of Hsp70 on Bag1 activity.
Figure 5: Characterization of Hsp70 mutants defective in Bag1 interaction.
Figure 6: Expression of Bag1 mutants defective in Hsp70 binding interferes with the arrest of DNA synthesis by heat shock.

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Acknowledgements

The authors thank Ellen Nollen for providing advice and the OT-Bag1 cell line, Dr. John Erikson (University of Turku) for valuable discussions, support from the Carol and Martin Gollub Foundation, and a grant to R.I.M. from National Institutes of General Medical Sciences, members of the laboratory for their comments on the manuscript, and many colleagues for generously providing key reagents.

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Correspondence to Richard I. Morimoto.

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Song, J., Takeda, M. & Morimoto, R. Bag1–Hsp70 mediates a physiological stress signalling pathway that regulates Raf-1/ERK and cell growth. Nat Cell Biol 3, 276–282 (2001). https://doi.org/10.1038/35060068

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