The RING-type E3 ligase RNF186 ubiquitinates Sestrin-2 and thereby controls nutrient sensing

Travis B. Lear; Karina C. Lockwood; Yurong Ouyang; John W. Evankovich; Mads B. Larsen; Bo Lin; Yuan Liu; Bill B. Chen

doi:10.1074/jbc.AC119.010671

The RING-type E3 ligase RNF186 ubiquitinates Sestrin-2 and thereby controls nutrient sensing

^‡Department of Environmental and Occupational Health, School of Public Health, University of Pittsburgh, Pittsburgh, Pennsylvania 15261
^§Aging Institute, Department of Medicine, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania 15213
^¶Acute Lung Injury Center of Excellence, Division of Pulmonary, Allergy, and Critical Care Medicine, Department of Medicine, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania 15213
^‖Vascular Medicine Institute, Department of Medicine, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania 15213

↵1 To whom correspondence may be addressed: Bridgeside Point 1, 100 Technology Dr., Pittsburgh, PA 15219. Tel.: 412-624-2664; E-mail: liuy13{at}upmc.edu.
↵2 To whom correspondence may be addressed: Bridgeside Point 1, 100 Technology Dr., Pittsburgh, PA 15219. Tel.: 412-624-2664; E-mail: chenb{at}upmc.edu.

Edited by George N. DeMartino

Abstract

Nutrient sensing is a critical cellular process controlling metabolism and signaling. mTOR complex 1 (mTORC1) is the primary signaling hub for nutrient sensing and, when activated, stimulates anabolic processes while decreasing autophagic flux. mTORC1 receives nutrient status signals from intracellular amino acid sensors. One of these sensors, Sestrin-2, functions as an intracellular sensor of cytosolic leucine and inhibitor of mTORC1 activity. Genetic studies of Sestrin-2 have confirmed its critical role in regulating mTORC1 activity, especially in the case of leucine starvation. Sestrin-2 is known to be transcriptionally controlled by several mechanisms; however, the post-translational proteolytic regulation of Sestrin-2 remains unclear. Here, we explored how Sestrin-2 is regulated through the ubiquitin proteasome system. Using an unbiased screening approach of an siRNA library targeting ubiquitin E3 ligases, we identified a RING-type E3 ligase, ring finger protein 186 (RNF186), that critically mediates the Sestrin-2 ubiquitination and degradation. We observed that RNF186 and Sestrin-2 bind each other through distinct C-terminal motifs and that Lys-13 in Sestrin-2 is a putative ubiquitin acceptor site. RNF186 knockdown increased Sestrin-2 protein levels and decreased mTORC1 activation. These results reveal a new mechanism of E3 ligase control of mTORC1 activity through the RNF186-Sestrin-2 axis, suggesting that RNF186 inhibition may be a potential strategy to increase levels of the mTORC1 inhibitor Sestrin-2.

Footnotes

This work was supported by the University of Pittsburgh Aging Institute seed fund (to B. B. C and Y. L.) as well as NIDDK, National Institutes of Health (NIH), Grant 1R01DK119627 (to Y. L.) and NHLBI, NIH, Grants 5F31HL143843–02 (to T. B. L.), 1K08HL144820 (to J. W. E.), 5R01HL14277702 (to Y. L.), and 5R35HL139860–02 and 5R01HL133184 (to B. B. C.). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.