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Hid, Rpr and Grim negatively regulate DIAP1 levels through distinct mechanisms

Abstract

Inhibitor of apoptosis (IAP) proteins suppress apoptosis and inhibit caspases. Several IAPs also function as ubiquitin-protein ligases. Regulators of IAP auto-ubiquitination, and thus IAP levels, have yet to be identified. Here we show that Head involution defective (Hid), Reaper (Rpr) and Grim downregulate Drosophila melanogaster IAP1 (DIAP) protein levels. Hid stimulates DIAP1 polyubiquitination and degradation. In contrast to Hid, Rpr and Grim can downregulate DIAP1 through mechanisms that do not require DIAP1 function as a ubiquitin-protein ligase. Observations with Grim suggest that one mechanism by which these proteins produce a relative decrease in DIAP1 levels is to promote a general suppression of protein translation. These observations define two mechanisms through which DIAP1 ubiquitination controls cell death: first, increased ubiquitination promotes degradation directly; second, a decrease in global protein synthesis results in a differential loss of short-lived proteins such as DIAP1. Because loss of DIAP1 is sufficient to promote caspase activation, these mechanisms should promote apoptosis.

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Figure 1: Loss of DIAP1 results in Drice activation.
Figure 2: Expression of Hid or Rpr results in a loss of DIAP1 in Drosophila wing discs.
Figure 3: Expression of Hid, Rpr or Grim results in a loss of DIAP1 in the embryo.
Figure 4: Expression of Hid, Rpr or Grim does not influence DIAP1 transcript levels.
Figure 6: Hid stimulates DIAP1 polyubiquitination.
Figure 5: Hid-dependent downregulation of DIAP1 requires DIAP1 ubiquitin-protein ligase activity, but Rpr-dependent downregulation does not.
Figure 7: Hid induces a post-translational loss of DIAP1, whereas Grim causes a general suppression of translation.

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Acknowledgements

We thank members of the Deshaies lab for their assistance with initial ubiquitination assays and S. Kornbluth for providing the Rpr peptide, and for sharing unpublished observations. We also thank P.D. Zamore and members of his lab for providing the Drosophila embryo translation extract and translation protocols. We also thank G.M. Rubin and E. Kwan for the production of the anti-DIAP1 monoclonal antibody and K. White for th6 flies. This work was supported in part by a grant from the Deutsche Forschungsgemeinschaft (MU1168/4-1) to H.-A.J.M. and grants from the Burroughs Wellcome Fund (New Investigator awards in the Pharmacological Sciences), the Ellison Medical Foundation, and a National Institutes of Health grant GM057422-01 to B.A.H. S.J.Y. was supported by a Jane Coffin Childs Postdoctoral fellowship.

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Correspondence to H.-Arno J. Müller or Bruce A. Hay.

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Figure S1

DIAP1 and Dronc half-life in S2 cells. (PDF 73 kb)

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Yoo, S., Huh, J., Muro, I. et al. Hid, Rpr and Grim negatively regulate DIAP1 levels through distinct mechanisms. Nat Cell Biol 4, 416–424 (2002). https://doi.org/10.1038/ncb793

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