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Src activates retrograde membrane traffic through phosphorylation of GBF1

Joanne Chia1, Shyi-Chyi Wang1,2, Sheena Wee1

  • 1Institute of Molecular and Cell Biology, Singapore, Singapore.

Elife
|December 6, 2021
PubMed

Insights

Src tyrosine kinase controls cancer-critical protein glycosylation by relocating GALNTs enzymes. It phosphorylates GBF1, promoting its binding to Arf1, which drives this essential transport event.

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Biochemistry

Background:

  • Src tyrosine kinase regulates protein glycosylation, a process critical for cancer progression.
  • The relocation of GALNTs (GalNAc-transferases) from the Golgi to the ER is essential for this glycosylation control.
  • The underlying mechanism by which Src induces this specific Golgi-to-ER trafficking remains elusive.

Purpose of the Study:

  • To elucidate the molecular mechanism by which Src tyrosine kinase mediates the relocation of GALNTs enzymes.
  • To investigate the role of the Guanine nucleotide Exchange Factor (GEF) GBF1 and small GTPase Arf1 in Src-induced protein trafficking.

Main Methods:

  • Utilized Src kinase activity assays and phosphoproteomic analysis to identify phosphorylation sites on GBF1.
  • Employed molecular modeling to predict the structural impact of phosphorylation on GBF1-Arf1 interactions.
  • Generated and tested GBF1 mutants (defective and phosphomimetic) to assess their role in transport carrier formation and GALNTs relocation.

Main Results:

  • Src induces the formation of tubular transport carriers containing GALNTs enzymes.
  • Src phosphorylates GBF1 on 10 tyrosine residues, including Y876 and Y898 within the Sec7 GEF domain.
  • Phosphorylation at Y876 and Y898 promotes GBF1 binding to Arf1, leading to carrier formation and GALNTs relocation.
  • Mutant and phosphomimetic GBF1 analyses confirmed the functional importance of these phosphorylation-dependent rearrangements.

Conclusions:

  • Src tyrosine kinase promotes GALNTs relocation by enhancing the binding of GBF1 to Arf1 through specific tyrosine phosphorylations.
  • This phosphorylation-dependent regulation of GEF-GTPase interaction represents a potential conserved mechanism for controlling intracellular transport.
  • Understanding this pathway offers new insights into the regulation of protein glycosylation in cancer.
Keywords:
Arf1GBF1Srccell biologygolgihumanmembrane trafficsignalling

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