Phosphorylation of the spindle checkpoint protein Mad2 regulates its conformational transition

Soonjoung Kim, Hongbin Sun, Haydn L. Ball, Katja Wassmann, Xuelian Luo, Hongtao Yu

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

Regulated conformational changes of proteins are critical for cellular signal transduction. The spindle checkpoint protein Mad2 is an unusual protein with two native folds: the latent open conformer (O-Mad2) and the activated closed conformer (C-Mad2). During mitosis, cytosolic O-Mad2 binds to the Mad1-Mad2 core complex at unattached kinetochores and undergoes conformational activation to become C-Mad2. C-Mad2 binds to and inhibits Cdc20, an activator of APC/C, to prevent precocious anaphase onset. Here, we show that the conformational transition of Mad2 is regulated by phos- phorylation of S195 in its C-terminal region. The phospho-mimicking Mad2S195D mutant and the phospho-S195 Mad2 protein obtained using intein-mediated semisynthesis do not form C-Mad2 on their own. Mad2S195D fails to bind to Cdc20, a low-affinity ligand, but still binds to high-affinity ligands, such as Mad1 and MBP1, forming ligand-bound C-Mad2. Overexpression of Mad2S195D in human cells causes checkpoint defects. Our results indicate that Mad2 phosphor- ylation inhibits its function through differentially regulating its binding to Mad1 and Cdc20 and establish that the conformational change of Mad2 is regulated by posttranslational mechanisms.

Original languageEnglish (US)
Pages (from-to)19772-19777
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume107
Issue number46
DOIs
StatePublished - Nov 16 2010

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Mad2 Proteins
Phosphorylation
Ligands
Inteins
Kinetochores
Anaphase
Mitosis
Signal Transduction
Proteins

Keywords

  • Conformational change
  • Mitosis
  • NMR
  • Protein structure
  • Spindle checkpoint

ASJC Scopus subject areas

  • General

Cite this

Phosphorylation of the spindle checkpoint protein Mad2 regulates its conformational transition. / Kim, Soonjoung; Sun, Hongbin; Ball, Haydn L.; Wassmann, Katja; Luo, Xuelian; Yu, Hongtao.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 107, No. 46, 16.11.2010, p. 19772-19777.

Research output: Contribution to journalArticle

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