RT Journal Article
SR Electronic
T1 Visual detection of binary, ternary, and quaternary protein-protein interactions in fission yeast by Pil1 co-tethering assay
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2021.04.13.439745
DO 10.1101/2021.04.13.439745
A1 Zhong-Qiu Yu
A1 Xiao-Man Liu
A1 Dan Zhao
A1 Dan-Dan Xu
A1 Li-Lin Du
YR 2021
UL http://biorxiv.org/content/early/2021/04/14/2021.04.13.439745.abstract
AB Protein-protein interactions are vital for executing nearly all cellular processes. To facilitate the detection of protein-protein interactions in living cells of the fission yeast Schizosaccharomyces pombe, here we present an efficient and convenient method termed the Pil1 co-tethering assay. In its basic form, we tether a bait protein to mCherry-tagged Pil1, which forms cortical filamentary structures, and examine whether a GFP-tagged prey protein colocalizes with the bait. We demonstrate that this assay is capable of detecting pairwise protein-protein interactions of cytosolic proteins, transmembrane proteins, and nuclear proteins. Furthermore, we show that this assay can be used for detecting not only binary protein-protein interactions, but also ternary and quaternary protein-protein interactions. Using this assay, we systematically characterized the protein-protein interactions in the Atg1 complex and in the phosphatidylinositol 3-kinase (PtdIns3K) complexes and found that Atg38 is incorporated into the PtdIns3K complex I via an Atg38-Vps34 interaction. Our data show that this assay is a useful and versatile tool and should be added to the routine toolbox of fission yeast researchers.Competing Interest StatementThe authors have declared no competing interest.Y2Hyeast two-hybridFRETfluorescence resonance energy transferBiFCbimolecular fluorescence complementationAIMAtg8-family-interacting motifNLSnuclear localization signal