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A Yeast 2-Hybrid Screen in Batch to Compare Protein Interactions
Published on: June 6, 2018
Proteomic and yeast 2-hybrid screens to identify PTEN binding partners
Priyanka Tibarewal1, Laura Spinelli2, Helene Maccario3
1Institute of Biological Chemistry, Biophysics and Bioengineering, Heriot Watt University, Edinburgh, UK; School of Life Sciences, University of Dundee, Dundee, UK; UCL Cancer Centre, University College London, London, UK.
Abstract:
PTEN is a phosphoinositide lipid phosphatase and an important tumour suppressor protein. PTEN function is reduced or lost in around a third of all human cancers through diverse mechanisms, from gene deletion to changes in the function of proteins which regulate PTEN through direct protein binding. Here we present data from SILAC (Stable Isotope Labelling by Amino acids in Cell culture) proteomic screens to identify proteins which bind to PTEN. These experiments using untransformed epithelial cells and glioma cells identified several novel candidate proteins in addition to many previously identified PTEN binding partners and many proteins which are recognised as common false positives using these methods. From subsequent co-expression pull-down experiments we provide further evidence supporting the physical interaction of PTEN with MMP1, Myosin 18A and SHROOM3. We also performed yeast two-hybrid screens which identify the previously recognised PTEN binding partner MSP58 in addition to the nuclear import export receptor TNPO3. These experiments identify several novel candidate binding partners of PTEN and provide further data addressing the set of proteins that interact with this important tumour suppressor.
Insights
Researchers identified novel proteins interacting with PTEN, a key tumor suppressor. These findings advance our understanding of PTEN regulation and its role in cancer by revealing new binding partners.
Area of Science:
- Molecular Biology
- Biochemistry
- Oncology
Background:
- PTEN (Phosphatase and tensin homolog) is a crucial tumor suppressor protein.
- Loss or reduction of PTEN function occurs in approximately one-third of human cancers via various mechanisms.
- Understanding PTEN's regulatory network is vital for cancer research.
Purpose of the Study:
- To identify novel protein-protein interactions involving PTEN.
- To expand the known interactome of PTEN.
- To uncover new regulatory mechanisms of this tumor suppressor.
Main Methods:
- Utilized Stable Isotope Labeling by Amino acids in Cell culture (SILAC) proteomic screening.
- Performed co-expression pull-down experiments.
- Conducted yeast two-hybrid screens.
Main Results:
- Identified several novel candidate PTEN-binding proteins.
- Confirmed physical interactions between PTEN and MMP1, Myosin 18A, and SHROOM3.
- Discovered interactions with MSP58 and the nuclear import-export receptor TNPO3.
Conclusions:
- This study identifies novel PTEN-interacting proteins, including MMP1, Myosin 18A, SHROOM3, MSP58, and TNPO3.
- These findings contribute to a comprehensive understanding of PTEN's regulatory network.
- The identified interactions offer potential new avenues for cancer therapy research.
