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Radiolabeling and Quantification of Cellular Levels of Phosphoinositides by High Performance Liquid Chromatography-coupled Flow Scintillation
Published on: January 6, 2016
Fab1p PtdIns(3)P 5-kinase function essential for protein sorting in the multivesicular body
1Division of Cellular and Molecular Medicine and Howard Hughes Medical Institute, University of California, San Diego, School of Medicine, La Jolla 92093-0668, USA.
Proper sorting of cell surface receptors into multivesicular bodies (MVBs) is crucial for their lysosomal degradation. This study reveals phosphoinositide signaling, specifically PtdIns(3,5)P2, is essential for cargo-selective MVB sorting in yeast.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Signal-transducing cell surface receptors require sorting into multivesicular bodies (MVBs) for down-regulation and lysosomal degradation.
- Yeast mutants with defects in vacuolar/lysosomal transport accumulate MVBs, indicating a role for these pathways in MVB processing.
Purpose of the Study:
- To investigate the molecular mechanisms governing cargo-selective sorting into MVBs.
- To elucidate the role of phosphoinositide signaling in vacuolar/lysosomal protein transport.
Main Methods:
- Analysis of yeast mutants defective in vacuolar transport.
- Characterization of carboxypeptidase S and Ste2p localization within MVBs.
- Assessing the requirement of VPS gene products and Fab1p kinase activity for protein sorting.
Main Results:
- Carboxypeptidase S and Ste2p are targeted to the vacuole lumen, dependent on specific VPS gene products.
- The PtdIns(3)P 5-kinase activity of Fab1p, producing PtdIns(3,5)P2, is essential for cargo-selective sorting into the vacuole lumen.
- Distinct phosphoinositide signaling events regulate vacuolar/lysosomal protein transport.
Conclusions:
- Phosphoinositide signaling plays a critical role at multiple stages of vacuolar/lysosomal protein transport.
- PtdIns(3,5)P2 synthesis is directly coupled to the regulation of MVB sorting.
- This study highlights the importance of MVB sorting for receptor regulation and cellular homeostasis.
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