A SPOPL/Cullin-3 ubiquitin ligase complex regulates endocytic trafficking by targeting EPS15 at endosomes

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Abstract

Cullin-3 (CUL3)-based ubiquitin ligases regulate endosome maturation and trafficking of endocytic cargo to lysosomes in mammalian cells. Here, we report that these functions depend on SPOPL, a substrate-specific CUL3 adaptor. We find that SPOPL associates with endosomes and is required for both the formation of multivesicular bodies (MVBs) and the endocytic host cell entry of influenza A virus. In SPOPL-depleted cells, endosomes are enlarged and fail to acquire intraluminal vesicles (ILVs). We identify a critical substrate ubiquitinated by CUL3-SPOPL as EPS15, an endocytic adaptor that also associates with the ESCRT-0 complex members HRS and STAM on endosomes. Indeed, EPS15 is ubiquitinated in a SPOPL-dependent manner, and accumulates with HRS in cells lacking SPOPL. Together, our data indicates that a CUL3-SPOPL E3 ubiquitin ligase complex regulates endocytic trafficking and MVB formation by ubiquitinating and degrading EPS15 at endosomes, thereby influencing influenza A virus infection as well as degradation of EGFR and other EPS15 targets.

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Michaela Gschweitl

Institute of Biochemistry, Department of Biology, Eidgenössische Technische Hochschule Zürich, Zürich, Switzerland

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The authors declare that no competing interests exist.
Anna Ulbricht

Institute of Biochemistry, Department of Biology, Eidgenössische Technische Hochschule Zürich, Zurich, Switzerland

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The authors declare that no competing interests exist.
Christopher A Barnes

Novo Nordisk Research Center, Seattle, United States

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Radoslav I Enchev

Institute of Biochemistry, Department of Biology, Eidgenössische Technische Hochschule Zürich, Zurich, Switzerland

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The authors declare that no competing interests exist.
Ingrid Stoffel-Studer

Institute of Biochemistry, Department of Biology, Eidgenössische Technische Hochschule Zürich, Zurich, Switzerland

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Nathalie Meyer-Schaller

Department of Biomedicine, University of Basel, Basel, Switzerland

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Jatta Huotari

nspm ltd, Meggen, Switzerland

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Yohei Yamauchi

Institute of Molecular Life Sciences, University of Zurich, Zurich, Switzerland

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Urs F Greber

Institute of Molecular Life Sciences, University of Zurich, Zurich, Switzerland

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The authors declare that no competing interests exist.
Ari Helenius

Institute of Biochemistry, Department of Biology, Eidgenössische Technische Hochschule Zürich, Zurich, Switzerland

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The authors declare that no competing interests exist.
Matthias Peter

Institute of Biochemistry, Department of Biology, Eidgenössische Technische Hochschule Zürich, Zurich, Switzerland

For correspondence
matthias.peter@bc.biol.ethz.ch

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The authors declare that no competing interests exist.

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