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PMID: 7593191 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Mad1p, a phosphoprotein component of the spindle assembly checkpoint in budding yeast.

The Journal of cell biology ·Vol. 131 ·No. 3 ·1995-11-00 ·Pages 709-20

Hardwick KG, Murray AW

Abstract

The spindle assembly checkpoint prevents cells from initiating anaphase until the spindle has been fully assembled. We previously isolated mitotic arrest deficient (mad) mutants that inactivate this checkpoint and thus increase the sensitivity of cells to benomyl, a drug that interferes with mitotic spindle assembly by depolymerizing microtubules. We have cloned the MAD1 gene and show that when it is disrupted yeast cells have the same phenotype as the previously isolated mad1 mutants: they fail to delay the metaphase to anaphase transition in response to microtubule depolymerization. MAD1 is predicted to encode a 90-kD coiled-coil protein. Anti-Mad1p antibodies give a novel punctate nuclear staining pattern and cell fractionation reveals that the bulk of Mad1p is soluble. Mad1p becomes hyperphosphorylated when wild-type cells are arrested in mitosis by benomyl treatment, or by placing a cold sensitive tubulin mutant at the restrictive temperature. This modification does not occur in G1-arrested cells treated with benomyl or in cells arrested in mitosis by defects in the mitotic cyclin proteolysis machinery, suggesting that Mad1p hyperphosphorylation is a step in the activation of the spindle assembly checkpoint. Analysis of Mad1p phosphorylation in other spindle assembly checkpoint mutants reveals that this response to microtubule-disrupting agents is defective in some (mad2, bub1, and bub3) but not all (mad3, bub2) mutant strains. We discuss the possible functions of Mad1p at this cell cycle checkpoint.

MeSH Terms
Amino Acid Sequence Base Sequence Carrier Proteins Cell Cycle Proteins Cell Nucleus/chemistry Fungal Proteins/physiology Molecular Sequence Data Mutation/physiology Nuclear Proteins/genetics,metabolism Phosphoproteins/genetics,metabolism,physiology Phosphorylation Repressor Proteins Spindle Apparatus/chemistry,physiology Yeasts/genetics
Chemicals
Carrier Proteins Cell Cycle Proteins Fungal Proteins MAD1L1 protein, human Nuclear Proteins Phosphoproteins Repressor Proteins
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Hardwick K G
Department of Physiology, University of California, San Francisco 94143-0444, USA.
Murray A W
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Article Info
Journal
The Journal of cell biology
Abbr.
J Cell Biol
ISSN
0021-9525
DOI
10.1083/jcb.131.3.709
Published
1995-11-00
Pages
709-20
Language
English
Region
United States
NLM ID
0375356
PMCID
PMC2120625
Subset
IM
Databases
GENBANK
U14632
External Links
PubMed source DOI: 10.1083/jcb.131.3.709 PMC Full Text View journal details
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