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The type I TGF-β receptor engages TRAF6 to activate TAK1 in a receptor kinase-independent manner

Abstract

Transforming growth factor-β (TGF-β) is a multifunctional cytokine that regulates embryonic development and tissue homeostasis; however, aberrations of its activity occur in cancer1,2. TGF-β signals through its Type II and Type I receptors (TβRII and TβRI) causing phosphorylation of Smad proteins3,4. TGF-β-associated kinase 1 (TAK1), a member of the mitogen-activated protein kinase kinase kinase (MAPKKK) family, was originally identified as an effector of TGF-β-induced p38 activation5. However, the molecular mechanisms for its activation are unknown. Here we report that the ubiquitin ligase (E3) TRAF6 interacts with a consensus motif present in TβRI. The TβRI–TRAF6 interaction is required for TGF-β-induced autoubiquitylation of TRAF6 and subsequent activation of the TAK1–p38/JNK pathway, which leads to apoptosis. TβRI kinase activity is required for activation of the canonical Smad pathway, whereas E3 activity of TRAF6 regulates the activation of TAK1 in a receptor kinase-independent manner. Intriguingly, TGF-β-induced TRAF6-mediated Lys 63-linked polyubiquitylation of TAK1 Lys 34 correlates with TAK1 activation. Our data show that TGF-β specifically activates TAK1 through interaction of TβRI with TRAF6, whereas activation of Smad2 is not dependent on TRAF6.

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Figure 1: TGF-β induces Lys 63-linked polyubiquitylation and activation of TAK1 as shown by in vivo ubiquitylation assays.
Figure 2: Endogenous TAK1 interacts with the TGF-β type I receptor (ALK5).
Figure 3: Interaction between TRAF6 and ALK5 is required for TGF-β-induced activation of TAK1 and p38 MAPK.
Figure 4: The E3 ubiquitin ligase activity of TRAF6 is required for TGF-β-induced activation of TAK1 and p38.
Figure 5: E3 activity of TRAF6 is required for TGF-β-induced activation of p38/JNK and apoptosis but not for activation of Smad2.

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Acknowledgements

We are grateful to P. ten Dijke, K. Miyazono, I. Dikic, K. Matsumoto, J. Ninomiya-Tsuji, Z.J. Chen and V.M. Dixit for providing expression vectors and reagents, and J. Inoue for his gift of wild-type and TRAF6−/− MEFs. We are grateful to U. Engström who synthesized the p-TAK1 peptide used for generation of rabbit anti-serum. We thank J. Ericsson for his valuable advice on how to perform in vitro ubiquitylation assays, and A. Moustakas and A. Morén for their advice on how to perform in vivo ubiquitylation assays. This work was supported in part by grants from the Swedish Medical Research Council, the Swedish Cancer Society, Wenner-Grenska Society and the Torsten and Ragnar Söderbergs Foundation (M.L.).

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A.S., N.T., S.G., A.M., V.v.B., N.S., S.Z. and M.L. performed the experiments; C.-H.H. and M.L. planned the project; A.S., N.T., C.-H.H. and M.L. prepared the manuscript.

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Correspondence to Maréne Landström.

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The authors declare no competing financial interests.

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Supplementary Figures S1, S2, S3, S4, S5 and S6 (PDF 904 kb)

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Sorrentino, A., Thakur, N., Grimsby, S. et al. The type I TGF-β receptor engages TRAF6 to activate TAK1 in a receptor kinase-independent manner. Nat Cell Biol 10, 1199–1207 (2008). https://doi.org/10.1038/ncb1780

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