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PMID: 11909965 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Fission yeast Mad3p is required for Mad2p to inhibit the anaphase-promoting complex and localizes to kinetochores in a Bub1p-, Bub3p-, and Mph1p-dependent manner.

Molecular and cellular biology ·Vol. 22 ·No. 8 ·2002-04-00 ·Pages 2728-42

Millband DN, Hardwick KG

Abstract

The spindle checkpoint delays the metaphase-to-anaphase transition in response to spindle and kinetochore defects. Genetic screens in budding yeast identified the Mad and Bub proteins as key components of this conserved regulatory pathway. Here we present the fission yeast homologue of Mad3p. Cells devoid of mad3(+) are unable to arrest their cell cycle in the presence of microtubule defects. Mad3p coimmunoprecipitates Bub3p, Mad2p, and the spindle checkpoint effector Slp1/Cdc20p. We demonstrate that Mad3p function is required for the overexpression of Mad2p to result in a metaphase arrest. Mad1p, Bub1p, and Bub3p are not required for this arrest. Thus, Mad3p appears to have a crucial role in transducing the inhibitory "wait anaphase" signal to the anaphase-promoting complex (APC). Mad3-green fluorescent protein (GFP) is recruited to unattached kinetochores early in mitosis and accumulates there upon prolonged checkpoint activation. For the first time, we have systematically studied the dependency of Mad3/BubR1 protein recruitment to kinetochores. We find Mad3-GFP kinetochore localization to be dependent upon Bub1p, Bub3p, and the Mph1p kinase, but not upon Mad1p or Mad2p. We discuss the implications of these findings in the context of our current understanding of spindle checkpoint function.

MeSH Terms
Amino Acid Sequence Anaphase Calcium-Binding Proteins/genetics,metabolism Carrier Proteins/metabolism Cell Cycle Proteins/genetics,metabolism Fungal Proteins/genetics,metabolism Genes, Fungal Kinetochores/metabolism Mad2 Proteins Models, Biological Molecular Sequence Data Nuclear Proteins Protein Kinases/metabolism Protein Serine-Threonine Kinases Recombinant Fusion Proteins/metabolism Schizosaccharomyces/cytology,genetics,metabolism Schizosaccharomyces pombe Proteins Sequence Homology, Amino Acid Signal Transduction Spindle Apparatus/metabolism
Chemicals
Calcium-Binding Proteins Carrier Proteins Cell Cycle Proteins Fungal Proteins Mad2 Proteins Nuclear Proteins Recombinant Fusion Proteins Schizosaccharomyces pombe Proteins mad2 protein, S pombe mad3 protein, S pombe Protein Kinases Bub1 spindle checkpoint protein Protein Serine-Threonine Kinases
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Millband David N
Wellcome Trust Centre for Cell Biology, Institute of Cell and Molecular Biology, University of Edinburgh, Edinburgh, Scotland EH9 3JR, United Kingdom.
Hardwick Kevin G
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/MCB.22.8.2728-2742.2002
Published
2002-04-00
Pages
2728-42
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC133725
Subset
IM
External Links
PubMed source DOI: 10.1128/MCB.22.8.2728-2742.2002 PMC Full Text View journal details
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