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The CTLH ubiquitin ligase substrates ZMYND19 and MKLN1 negatively regulate mTORC1 at the lysosomal membrane

Yin Wang1,2, Yifei Liao1,2, Yizhe Sun1,2

  • 1Division of Infectious Diseases, Department of Medicine, Brigham and Women's Hospital, Boston, MA, USA.

Nature Communications
|November 28, 2025
PubMed

Insights

The carboxy terminal to LisH (CTLH) E3 ligase complex inhibits phosphoinositide 3 kinase/mammalian target of rapamycin (PI3K/mTOR) pathway signaling in Epstein-Barr virus-associated gastric cancer. This discovery offers new therapeutic targets for treating this cancer.

Area of Science:

  • Oncology
  • Molecular Biology
  • Biochemistry

Background:

  • Epstein-Barr virus-associated gastric carcinoma (EBVaGC) often features mutations activating the phosphoinositide 3 kinase (PI3K) pathway.
  • Dysregulation of the PI3K/mammalian target of rapamycin (mTOR) pathway is a key driver of EBVaGC proliferation.

Purpose of the Study:

  • To identify genetic modifiers that synergistically inhibit EBVaGC proliferation when combined with a PI3K antagonist.
  • To elucidate the role of the carboxy terminal to LisH (CTLH) E3 ligase complex in PI3K/mTOR pathway regulation within EBVaGC.

Main Methods:

  • Genome-wide CRISPR/Cas9 screening in EBVaGC cells treated with the PI3K antagonist alpelisib.
  • Analysis of CTLH E3 ligase subunits, substrates (MKLN1, ZMYND19), and their interaction with mTORC1.
  • Investigation of mTORC1 signaling components, including Rheb, S6, and 4E-BP1.

Main Results:

  • Multiple CTLH E3 ligase subunits, including MAEA, were identified as synergistic hits in the CRISPR/Cas9 screen.
  • MAEA knockout led to accumulation of CTLH substrates MKLN1 and ZMYND19.
  • MKLN1 and ZMYND19 inhibit mTORC1 activity by blocking interactions with Rheb and downstream substrates S6 and 4E-BP1, independent of lysosomal recruitment.

Conclusions:

  • The CTLH E3 ligase complex acts as a negative regulator of mTORC1 signaling in EBVaGC.
  • CTLH modulates mTORC1 activity at the lysosomal membrane through substrate interactions and the ubiquitin-proteasome system.
  • Targeting the CTLH complex presents a potential therapeutic strategy for EBVaGC.

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