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O-GlcNAcylation of the intellectual disability protein DDX3X exerts proteostatic cell cycle control

Conor W Mitchell1,2, Huijie Yuan1,2, Marie Sønderstrup-Jensen1

  • 1Department of Molecular Biology and Genetics, Aarhus Universitet, Aarhus, Denmark.

Open Biology
|July 1, 2025
PubMed

Insights

O-GlcNAcylation of DDX3X at Ser584 stabilizes the protein, preventing its degradation. This proteostatic mechanism is crucial for regulating cell cycle entry into S phase by controlling cyclin E1 expression.

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Biochemistry

Background:

  • O-GlcNAcylation is a critical post-translational modification regulating cellular processes.
  • Cell cycle progression is sensitive to O-GlcNAc homeostasis, but regulatory mechanisms are unclear.
  • DDX3X, an RNA helicase linked to microcephaly, plays a role in cell cycle control.

Purpose of the Study:

  • To investigate the role of O-GlcNAcylation in regulating DDX3X stability and cell cycle progression.
  • To identify specific O-GlcNAcylation sites on DDX3X and their functional consequences.
  • To elucidate the proteostatic mechanisms linking O-GlcNAc modification to cell cycle regulation.

Main Methods:

  • Site-specific mutagenesis to alter O-GlcNAcylation at Ser584 of DDX3X.
  • Proteasomal degradation assays to assess DDX3X stability.
  • Western blotting to quantify protein levels of DDX3X and cyclin E1.
  • Flow cytometry to analyze cell cycle progression.

Main Results:

  • Loss of O-GlcNAcylation at Ser584 of DDX3X led to its increased degradation by the proteasome.
  • Reduced DDX3X stability resulted in decreased expression of its target gene, cyclin E1.
  • Impaired O-GlcNAcylation of DDX3X caused a delay in cell cycle progression from G1 to S phase.

Conclusions:

  • Ser584 O-GlcNAcylation of DDX3X acts as a proteostatic mechanism controlling its stability.
  • This modification is essential for regulating S-phase entry by modulating DDX3X levels and cyclin E1 expression.
  • A single O-GlcNAc site on DDX3X significantly impacts protein stability and cell cycle dynamics.
Keywords:
O-GlcNAccell cyclecell signalling

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