title: Discovering protein-protein interactions within the programmed cell death network using a protein-fragment complementation screen.
abstract: Apoptosis and autophagy are distinct biological processes, each driven by a different set of protein-protein interactions, with significant crosstalk via direct interactions among apoptotic and autophagic proteins. To measure the global profile of these interactions, we adapted the Gaussia luciferase protein-fragment complementation assay (GLuc PCA), which monitors binding between proteins fused to complementary fragments of a luciferase reporter. A library encompassing 63 apoptotic and autophagic proteins was constructed for the analysis of ∼3,600 protein-pair combinations. This generated a detailed landscape of the apoptotic and autophagic modules and points of interface between them, identifying 46 previously unknown interactions. One of these interactions, between DAPK2, a Ser/Thr kinase that promotes autophagy, and 14-3-3τ, was further investigated. We mapped the region responsible for 14-3-3τ binding and proved that this interaction inhibits DAPK2 dimerization and activity. This proof of concept underscores the power of the GLuc PCA platform for the discovery of biochemical pathways within the cell death network.
authors: Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors: Yuval
authors: Y
authors: Gilad
authors: Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors: Ruth
authors: R
authors: Shiloh
authors: Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors: Yaara
authors: Y
authors: Ber
authors: Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors: Shani
authors: S
authors: Bialik
authors: Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel. Electronic address: adi.kimchi@weizmann.ac.il.
authors: Adi
authors: A
authors: Kimchi
mesh_terms: D048948:14-3-3 Proteins / Q000378:metabolism; D017209:Apoptosis / Q000235:genetics*; D001343:Autophagy / Q000235:genetics*; D064550:Death-Associated Protein Kinases / Q000378:metabolism; D053263:Gene Regulatory Networks; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D011485:Protein Binding
chemical_list: D048948:14-3-3 Proteins; C579104:DAPK2 protein, human; D064550:Death-Associated Protein Kinases