Ubiquitylation of the ER-Shaping Protein Lunapark via the CRL3KLHL12 Ubiquitin Ligase Complex

Cell Rep. 2020 May 19;31(7):107664. doi: 10.1016/j.celrep.2020.107664.

Abstract

Cullin-RING ligases (CRLs) control key cellular processes by promoting ubiquitylation of a multitude of soluble cytosolic and nuclear proteins. Subsets of CRL complexes are recruited and activated locally at cellular membranes; however, few CRL functions and substrates at these distinct cellular compartments are known. Here, we use a proteomic screen to identify proteins that are ubiquitylated at cellular membranes and found that Lunapark, an endoplasmic reticulum (ER)-shaping protein localized to ER three-way junctions, is ubiquitylated by the CRL3KLHL12 ubiquitin ligase. We demonstrate that Lunapark interacts with mechanistic target of rapamycin complex-1 (mTORC1), a central cellular regulator that coordinates growth and metabolism with environmental conditions. We show that mTORC1 binds Lunapark specifically at three-way junctions, and lysosomes, where mTORC1 is activated, make contact with three-way junctions where Lunapark resides. Inhibition of Lunapark ubiquitylation results in neurodevelopmental defects indicating that KLHL12-dependent ubiquitylation of Lunapark is required for normal growth and development.

Keywords: ER three-way junction; Lunapark; cullin-RING ligases; endoplasmic reticulum; lysosome; mTORC1; ubiquitin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptor Proteins, Signal Transducing / metabolism*
  • Animals
  • Animals, Genetically Modified
  • Cell Line, Tumor
  • Cullin Proteins / metabolism*
  • Endoplasmic Reticulum / metabolism*
  • HEK293 Cells
  • Humans
  • Mechanistic Target of Rapamycin Complex 1 / metabolism
  • Membrane Proteins / metabolism*
  • Ubiquitin-Protein Ligases / metabolism*
  • Ubiquitination
  • Zebrafish

Substances

  • Adaptor Proteins, Signal Transducing
  • Cullin Proteins
  • KLHL12 protein, human
  • LNPK protein, human
  • Membrane Proteins
  • Ubiquitin-Protein Ligases
  • Mechanistic Target of Rapamycin Complex 1