The C-terminal domains of human neurofibromin and its budding yeast homologs Ira1 and Ira2 regulate the metaphase to anaphase transition

Cell Cycle. 2014;13(17):2780-9. doi: 10.4161/15384101.2015.945870.

Abstract

The human tumor suppressor neurofibromin contains a cysteine and serine-rich domain/Ras-GTPase activating protein domain (CSRD/RasGAP) and a C-terminal domain (CTD). Domain studies of neurofibromin suggest it has other functions in addition to being a RasGAP, but the mechanisms underlying its tumor suppressor activity are not well understood. The budding yeast Saccharomyces cerevisiae is a good model system for studying neurofibromin function because it possesses Ira1 and Ira2, which are homologous to human neurofibromin in both sequence and function. We found that overexpression of CTD or a neurofibromin CTD-homologous domain (CHD) of Ira1/2 in budding yeast delayed degradation of the securin protein Pds1, whereas overexpression of CSRD/RasGAP did not affect Pds1 degradation. We also found that when CTD or CHD was overexpressed, the number of cells in metaphase was higher than in the control. These results demonstrate that CTD and CHD function in the metaphase to anaphase transition. In addition, Δira1Δira2 cells bypassed mitotic arrest in response to spindle damage, indicating that Ira1 and Ira2 may be involved in the spindle assembly checkpoint (SAC). However, Δira1Δira2Δmad2 cells are more sensitive to spindle damage than Δmad2 or Δira1Δira2 cells are, suggesting that Ira1/2 and Mad2 function in different pathways. Overexpression of CTD but not CSRD/RasGAP partially rescued the hypersensitivity of Δira1Δira2Δmad2 cells to microtubule-destabilizing drugs, indicating a role for CTD in the SAC pathway. Taken together, independently of RasGAP activity, the C-terminal domains of neurofibromin, Ira1, and Ira2 regulate the metaphase to anaphase transition in a Mad2-independent fashion.

Keywords: APC, anaphase promoting complex; CHD, CTD-homologous domain; CSRD/RasGAP, cysteine and serine-rich domain/Ras-GTPase activating protein; CTD; CTD, C-terminal domain; Ira1; Ira2; NF1, neurofibromatosis type 1; Pds1; SAC, spindle assembly checkpoint; budding yeast; metaphase to anaphase transition; neurofibromin; spindle assembly checkpoint.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Anaphase*
  • GTPase-Activating Proteins / chemistry*
  • GTPase-Activating Proteins / metabolism*
  • Humans
  • M Phase Cell Cycle Checkpoints
  • Mad2 Proteins / metabolism
  • Metaphase*
  • Molecular Sequence Data
  • Neurofibromin 1 / chemistry*
  • Neurofibromin 1 / metabolism
  • Protein Structure, Tertiary
  • Saccharomyces cerevisiae / cytology
  • Saccharomyces cerevisiae / metabolism
  • Saccharomyces cerevisiae Proteins / chemistry*
  • Saccharomyces cerevisiae Proteins / metabolism*
  • Sequence Homology, Amino Acid*
  • Structure-Activity Relationship
  • ras GTPase-Activating Proteins / metabolism

Substances

  • GTPase-Activating Proteins
  • IRA1 protein, S cerevisiae
  • IRA2 protein, S cerevisiae
  • MAD2 protein, S cerevisiae
  • Mad2 Proteins
  • Neurofibromin 1
  • Saccharomyces cerevisiae Proteins
  • ras GTPase-Activating Proteins

Grant support

This work was supported by a grant from the 21C Frontier Microbial Genomics and Application Center Program, the Ministry of Education, Science and Technology of the Republic of Korea (11-2008-10-005-00). There is no conflict of interest on the contents of this manuscript.