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Modified Yeast-Two-Hybrid System to Identify Proteins Interacting with the Growth Factor Progranulin
Published on: January 17, 2012
Identification of Novel Interacting Proteins of FUZ and GPR161
Gabriella Salazar1, Anna Battenhouse2, Andrew J Kim1
1Department of Pediatrics, Dell Pediatric Research Institute, Dell Medical School, The University of Texas at Austin, Austin, Texas, USA.
Abstract:
Protein-protein interactions are central to the dynamic regulation of signaling pathways and provide critical insight into the cellular mechanisms underlying human disease. Our previous study demonstrated biochemical and genetic interactions between FUZ and GPR161 in sonic hedgehog signaling during spinal neural tube development. In this study, we sought to identify novel interacting proteins of FUZ and GPR161 to further characterize their biochemical and functional relationships. Using affinity-based liquid chromatography-tandem mass spectrometry of immunoprecipitated complexes from cells overexpressing FUZ and GPR161, we identified 159 shared co-interacting proteins, along with 289 proteins exclusive to FUZ and 617 proteins exclusive to GPR161. Gene Ontology (GO) analysis of the co-interactome revealed significant enrichment in proteasomal catabolic processes and intracellular trafficking pathways. GO analysis of the FUZ-specific interactome showed enrichment in cell cycle progression, mitochondrial membrane, and RNA metabolism, whereas the GPR161-specific interactome was enriched in receptor complex and endoplasmic reticulum-Golgi transport pathways. These findings were supported by STRING network analysis. Among the prioritized candidates, FKBP8 was validated as a binding partner of both FUZ and GPR161. Collectively, our proteomic study defines the protein interaction networks of FUZ and GPR161, highlighting their distinct and cooperative roles in multiple cellular processes and providing a foundation for future functional studies.
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