Assembling the Protein Architecture of the Budding Yeast Kinetochore-Microtubule Attachment using FRET
| Autor: | Ajit P. Joglekar, Isabella Theresa Felzer-Kim, Kaushik Gurunathan, Pavithra Aravamudhan |
|---|---|
| Rok vydání: | 2014 |
| Předmět: |
Chromosome movement
Saccharomyces cerevisiae Proteins Microtubule-associated protein Cell Cycle Proteins Biology Microtubules Models Biological Article General Biochemistry Genetics and Molecular Biology Microtubule Fluorescence Resonance Energy Transfer Kinetochores Cell Cycle Protein Agricultural and Biological Sciences(all) Kinetochore Biochemistry Genetics and Molecular Biology(all) Nuclear Proteins biology.organism_classification Cell biology NDC80 Förster resonance energy transfer Saccharomycetales Multiprotein Complexes General Agricultural and Biological Sciences Microtubule-Associated Proteins |
| Zdroj: | Current Biology. 24(13):1437-1446 |
| ISSN: | 0960-9822 |
| DOI: | 10.1016/j.cub.2014.05.014 |
| Popis: | Summary Background The kinetochore is a multiprotein machine that couples chromosome movement to microtubule (MT) polymerization and depolymerization. It uses numerous copies of at least three MT-binding proteins to generate bidirectional movement. The nanoscale organization of these proteins within the kinetochore plays an important role in shaping the mechanisms that drive persistent, bidirectional movement of the kinetochore. Results We used fluorescence resonance energy transfer (FRET) between genetically encoded fluorescent proteins fused to kinetochore subunits to reconstruct the nanoscale organization of the budding yeast kinetochore. We performed >60 FRET and high-resolution colocalization measurements involving the essential MT-binding kinetochore components: Ndc80, Dam1, Spc105, and Stu2. These measurements reveal that neighboring Ndc80 complexes within the kinetochore are narrowly distributed along the length of the MT. Dam1 complex molecules are concentrated near the MT-binding domains of Ndc80. Stu2 localizes in high abundance within a narrowly defined territory within the kinetochore centered ∼20 nm on the centromeric side of the Dam1 complex. Conclusions Our data show that the MT attachment site of the budding yeast kinetochore is well organized. Ndc80, Dam1, and Stu2 are all narrowly distributed about their average positions along the kinetochore-MT axis. The relative organization of these components, their narrow distributions, and their known MT-binding properties together elucidate how their combined actions generate persistent, bidirectional kinetochore movement coupled to MT polymerization and depolymerization. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|