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Updated: May 11, 2026

Imaging ATG9A, a Multi-Spanning Membrane Protein
Published on: June 16, 2023
Glycoprotein biosynthesis in a eukaryote lacking the membrane protein Rft1
Jennifer Jelk1, Ningguo Gao, Mauro Serricchio
1Department of Biochemistry, Weill Cornell Medical College, New York, New York 10065, USA.
Rft1 protein is not an M5-DLO flippase but may act as a chaperone for mature dolichol-linked oligosaccharide (mDLO) synthesis. This finding clarifies Rft1
Area of Science:
- Molecular Biology
- Cell Biology
- Glycoscience
Background:
- Mature dolichol-linked oligosaccharides (mDLOs) are crucial for eukaryotic protein N-glycosylation.
- The lipid intermediate Man5GlcNAc2-PP-dolichol (M5-DLO) must flip to the endoplasmic reticulum lumen for mDLO synthesis.
- The protein Rft1's role in this process has been debated, with yeast studies suggesting it's a flippase, but biochemical data are conflicting.
Purpose of the Study:
- To elucidate the precise function of the endoplasmic reticulum membrane protein Rft1 in mDLO biosynthesis.
- To investigate the in vivo role of Rft1 by studying its absence in an organism amenable to null mutant generation.
Main Methods:
- Generation and analysis of Rft1-null procyclic trypanosomes (TbRft1-null).
- Assessment of cell growth, steady-state levels of M5-DLO and mDLO.
- Analysis of N-glycosylation patterns in wild-type and mutant cells.
Main Results:
- TbRft1-null trypanosomes exhibited near-normal growth and maintained significant N-glycosylation.
- Mutant cells displayed 30-100-fold elevated steady-state levels of M5-DLO compared to wild-type.
- All N-glycans in mutant cells were derived from mDLO, indicating the excess M5-DLO was not utilized for glycosylation.
Conclusions:
- Rft1 does not function as the M5-DLO flippase.
- Rft1 likely facilitates the conversion of M5-DLO to mDLO through an alternative mechanism, potentially acting as an M5-DLO chaperone.
- This study redefines the role of Rft1 in the essential pathway of N-glycosylation.
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