Proteomics of isolated mitotic chromosomes identifies the kinetochore protein Ska3/Rama1

S. Ohta, J.-C. Bukowski-Wills, L. Wood, F. de Lima Alves, Z. Chen, J. Rappsilber, W.C. Earnshaw

Research output: Contribution to journalArticlepeer-review

Abstract / Description of output

Despite many decades of study, mitotic chromosomes remain poorly characterized with respect to their structure and composition. Here, we have purified mitotic chromosomes from nocodazole-treated chicken DT40 cells. These chromosomes have a 0.7:1:1 ratio of nonhistone proteins to histones to DNA. They also contain a significant content of RNAs that have yet to be characterized. Overall, the isolated chromosomes contained >4000 polypeptides, >500 of which are either novel or uncharacterized. Elsewhere, we have developed an approach for comparing the results of multiple proteomics experiments. As a validation of this approach, one of 13 novel centromere proteins identified was found to occur in a complex with the previously described proteins Ska1 and Ska2. This novel protein, now known as Ska3/Rama1, occupies a unique domain in the outer kinetochore and was revealed by RNA interference (RNAi) experiments to be essential for cell cycle progression in human cells. The approach presented here offers a powerful way to define the functional proteome of complex organelles and structures whose composition is not simple or fixed.
Original languageEnglish
Pages (from-to)433-438
Number of pages6
JournalCold Spring Harbor Symposia on Quantitative Biology
Publication statusPublished - 1 Jan 2010

Keywords / Materials (for Non-textual outputs)

  • Animals
  • Cell Line
  • Chickens
  • DNA
  • Histones
  • Humans
  • Kinetochores
  • Microtubule-Associated Proteins
  • Mitosis
  • Protein Binding
  • Proteome
  • Proteomics


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