Protein interactions regulate the systems-level behavior of cells; thus, deciphering the structure and dynamics of protein interaction networks in their cellular context is a central goal in biology. We have performed a genome-wide in vivo screen for protein-protein interactions in Saccharomyces cerevisiae by means of a protein-fragment complementation assay (PCA). We identified 2770 interactions among 1124 endogenously expressed proteins. Comparison with previous studies confirmed known interactions, but most were not known, revealing a previously unexplored subspace of the yeast protein interactome. The PCA detected structural and topological relationships between proteins, providing an 8-nanometer-resolution map of dynamically interacting complexes in vivo and extended networks that provide insights into fundamental cellular processes, including cell polarization and autophagy, pathways that are evolutionarily conserved and central to both development and human health.T he elucidation of protein-protein interaction networks (PINs, or interactomes) holds the promise of answering fundamental questions about how the biochemical machinery of cells organizes matter, information, and energy transformations to perform specific functions (1). An essential and rarely addressed question is whether protein complexes and PINs that are reconstructed or reconstituted in vitro or removed from the normal context in which they are expressed reflect their organization in living cells. For eukaryotes, the test bed for large-scale analysis of PINs is the yeast Saccharomyces cerevisiae, where several PIN analyses have been performed using yeast two-hybrid screens (Y2H) (2-4) or tandem affinity purification followed by massspectrometric analyses (TAP-MSs) (5-8). Each approach captures specific features of protein interactions; two-hybrid methods are best at measuring direct binary interactions between pairs of proteins, whereas affinity purification techniques best capture stable protein complexes. However, neither approach measures interactions between proteins in their natural cellular context, and are not easily amenable to studying protein complexes that are transiently associated or dynamic under different conditions, that do not survive in vitro purification, or that cannot be transported to the nucleus and form interactions in the absence of other
Phenotypes associated with genetic variants can be altered by interactions with other genetic variants (GxG), with the environment (GxE), or both (GxGxE). Yeast genetic interactions have been mapped on a global scale, but the environmental influence on the plasticity of genetic networks has not been examined systematically. To assess environmental rewiring of genetic networks, we examined 14 diverse conditions and scored 30,000 functionally representative yeast gene pairs for dynamic, differential interactions. Different conditions revealed novel differential interactions, which often uncovered functional connections between distantly related gene pairs. However, the majority of observed genetic interactions remained unchanged in different conditions, suggesting that the global yeast genetic interaction network is robust to environmental perturbation and captures the fundamental functional architecture of a eukaryotic cell.
The rate of cell-cycle progression must be tuned in response to nutrient levels to ensure that sufficient materials are synthesized to generate viable daughters. We report that accumulation of the yeast M phase B-cyclin CLB2 mRNA depends on assembly and activation of the heterogeneous nuclear RNA-binding protein (hnRNP) arginine methyltransferase Hmt1, which is promoted by the kinase Dbf2 and countered by the PP2A phosphatase Pph22. Activated Hmt1 methylates hnRNPs, which in turn stabilize CLB2 transcripts. Dbf2 activation of Hmt1 is highly cooperative, producing a sharp increase in CLB2, whereas Pph22 dephosphorylation is graded such that small changes in PP2A activity can cause large shifts in Dbf2-mediated Hmt1 activity. Starvation and rapamycin inhibition of TOR activate Pph22, causing a depletion of CLB2 and delay of M phase. We propose a general model wherein changes to Pph22 activity modulate cyclin mRNA stability to tune cell-cycle progression to environmental conditions.
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