The RZZ complex requires the N-terminus of KNL1 to mediate optimal Mad1 kinetochore localization in human cells

Open Biol. 2015 Nov;5(11):150160. doi: 10.1098/rsob.150160.

Abstract

The spindle assembly checkpoint is a surveillance mechanism that blocks anaphase onset until all chromosomes are properly attached to microtubules of the mitotic spindle. Checkpoint activity requires kinetochore localization of Mad1/Mad2 to inhibit activation of the anaphase promoting complex/cyclosome in the presence of unattached kinetochores. In budding yeast and Caenorhabditis elegans, Bub1, recruited to kinetochores through KNL1, recruits Mad1/Mad2 by direct linkage with Mad1. However, in human cells it is not yet established which kinetochore protein(s) function as the Mad1/Mad2 receptor. Both Bub1 and the RZZ complex have been implicated in Mad1/Mad2 kinetochore recruitment; however, their specific roles remain unclear. Here, we investigate the contributions of Bub1, RZZ and KNL1 to Mad1/Mad2 kinetochore recruitment. We find that the RZZ complex localizes to the N-terminus of KNL1, downstream of Bub1, to mediate robust Mad1/Mad2 kinetochore localization. Our data also point to the existence of a KNL1-, Bub1-independent mechanism for RZZ and Mad1/Mad2 kinetochore recruitment. Based on our results, we propose that in humans, the primary mediator for Mad1/Mad2 kinetochore localization is the RZZ complex.

Keywords: Bub1; KNL1; Mad1; RZZ complex; kinetochore; spindle checkpoint.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Cell Cycle Proteins / metabolism*
  • HeLa Cells
  • Humans
  • Kinetochores / metabolism*
  • Mad2 Proteins / metabolism
  • Microtubule-Associated Proteins / chemistry
  • Microtubule-Associated Proteins / metabolism*
  • Nuclear Proteins / metabolism*
  • Protein Binding
  • Protein Serine-Threonine Kinases / metabolism
  • Protein Structure, Tertiary
  • Protein Transport

Substances

  • Cell Cycle Proteins
  • Knl1 protein, human
  • MAD1L1 protein, human
  • MAD2L1 protein, human
  • Mad2 Proteins
  • Microtubule-Associated Proteins
  • Nuclear Proteins
  • BUB1 protein, human
  • Protein Serine-Threonine Kinases