Molecular analysis of kinetochore architecture in fission yeast

Abstract

Kinetochore composition and structure are critical for understanding how kinetochores of different types perform similar functions in chromosome segregation. We used affinity purification to investigate the kinetochore composition and assembly in Schizosaccharomyces pombe. We identified a conserved DASH complex that functions to ensure precise chromosome segregation. Unlike DASH in budding yeast that is localized onto kinetochores throughout the cell cycle, SpDASH is localized onto kinetochores only in mitosis. We also identified two independent groups of kinetochore components, one of which, the Sim4 complex, contains several novel Fta proteins in addition to known kinetochore components. DASH is likely to be associated with the Sim4 complex via Dad1 protein. The other group, Ndc80-MIND-Spc7 complex, contains the conserved Ndc80 and MIND complexes and Spc7 protein.We propose that fission yeast kinetochore is comprised of at least two major structural motifs that are biochemically separable. Our results suggest a high degree of conservation between the kinetochores of budding yeast and fission yeast even though many individual protein subunits do not have a high degree of sequence similarity.

Reconstruction of the Kinetochore during Meiosis in Fission Yeast Schizosaccharomyces pombe

“…NMS group proteins reassemble to the kinetochore during prophase and toward metaphase in meiosis, and subsequently DASH group proteins localize at the centromere during chromosome segregation. These groupings are generally consistent with the complex structures revealed by genetic interactions and proteomic analyses (De Wulf et al, 2003;Cheeseman et al, 2004;Obuse et al, 2004;Liu et al, 2005).…”

“…Centromere localization limited to the period of chromosome segregation supports their role in spindle attachment. Taken together, the behavior of these groups was generally consistent with the subcomplex structures that have been identified from genetic interaction and biochemical purification studies (Hayashi et al, 2004;Obuse et al, 2004;Liu et al, 2005).…”

“…Subcomplex structures of the S. pombe kinetochore, similar to that of S. cerevisiae and humans, have been reported (Hayashi et al, 2004;Obuse et al, 2004;Liu et al, 2005). The S. pombe kinetochore contains the Ndc80 complex (Ndc80, Nuf2, Spc24, and Spc25), which is highly conserved in many organisms from yeasts to humans (Nabetani et al, 2001;Wigge and Kilmartin, 2001).…”

“…These proteins include Sim4 and Mis15, which have been reported to depend on the Mis6 protein for their centromere localization (Takahashi et al, 2000;Pidoux et al, 2003;Hayashi et al, 2004). Thus, it is likely that these proteins compose the Mis6 complex (also called the Sim4 complex in Liu et al, 2005), which corresponds to the S. cerevisiae COMA and Ctf19 complexes. Ten S. pombe proteins, which are conserved in the S. cerevisiae DASH complex, were purified as a complex by biochemical purification .…”

A non-ring-like form of the Dam1 complex modulates microtubule dynamics in fission yeast

“…20). Similar to ScDam1, SpDam1 is localized at kinetochores as well as on the spindle and the intranuclear MT (20)(21)(22), and functions in coupling kinetochores to MTs (22)(23)(24). Using quantitative fluorescence microscopy, it was determined that on average only seven to eight copies of SpDam1 per kinetochore were found (25).…”