Fission yeast Opy1 is an endogenous PI(4,5)P2 sensor that binds to the phosphatidylinositol 4-phosphate 5-kinase Its3.

Phosphoinositides (PIPs) are a dynamic family of lipids that execute diverse roles in cell biology. PIP levels are regulated by numerous enzymes, but our understanding of how these enzymes are controlled in space and time is incomplete. One role of the PIP phosphatidylinositol (4,5)-bisphosphate [PI(4,5)P2] is to anchor the cytokinetic ring (CR) to the plasma membrane (PM) in Schizosaccharomyces pombe While examining potential PI(4,5)P2-binding proteins for roles in CR anchoring, we identified the dual pleckstrin homology (PH) domain-containing protein Opy1. Although related proteins are implicated in PIP regulation, we found no role for S. pombe Opy1 in CR anchoring, which would be expected if it modulated PM PI(4,5)P2 levels. Our data indicate that although Opy1 senses PM PI(4,5)P2 levels and binds to the phosphatidylinositol 4-phosphate 5-kinase (PI5-kinase) Its3, Opy1 does not regulate Its3 kinase activity or PM PI(4,5)P2 levels, a striking difference from its Saccharomyces cerevisiae homolog. However, overexpression of Opy1 resulted in cytokinesis defects, as might be expected if it sequestered PI(4,5)P2 Our results highlight the evolutionary divergence of dual PH domain-containing proteins and the need for caution when interpreting results based on their overexpression.This article has an associated First Person interview with the first author of the paper.

Dyskerin, tRNA genes, and condensin tether pericentric chromatin to the spindle axis in mitosis.

Condensin is enriched in the pericentromere of budding yeast chromosomes where it is constrained to the spindle axis in metaphase. Pericentric condensin contributes to chromatin compaction, resistance to microtubule-based spindle forces, and spindle length and variance regulation. Condensin is clustered along the spindle axis in a heterogeneous fashion. We demonstrate that pericentric enrichment of condensin is mediated by interactions with transfer ribonucleic acid (tRNA) genes and their regulatory factors. This recruitment is important for generating axial tension on the pericentromere and coordinating movement between pericentromeres from different chromosomes. The interaction between condensin and tRNA genes in the pericentromere reveals a feature of yeast centromeres that has profound implications for the function and evolution of mitotic segregation mechanisms.

Individual pericentromeres display coordinated motion and stretching in the yeast spindle.

The mitotic segregation apparatus composed of microtubules and chromatin functions to faithfully partition a duplicated genome into two daughter cells. Microtubules exert extensional pulling force on sister chromatids toward opposite poles, whereas pericentric chromatin resists with contractile springlike properties. Tension generated from these opposing forces silences the spindle checkpoint to ensure accurate chromosome segregation. It is unknown how the cell senses tension across multiple microtubule attachment sites, considering the stochastic dynamics of microtubule growth and shortening. In budding yeast, there is one microtubule attachment site per chromosome. By labeling several chromosomes, we find that pericentromeres display coordinated motion and stretching in metaphase. The pericentromeres of different chromosomes exhibit physical linkage dependent on centromere function and structural maintenance of chromosomes complexes. Coordinated motion is dependent on condensin and the kinesin motor Cin8, whereas coordinated stretching is dependent on pericentric cohesin and Cin8. Linking of pericentric chromatin through cohesin, condensin, and kinetochore microtubules functions to coordinate dynamics across multiple attachment sites.