The C-terminus of Bfa1p in budding yeast is essential to induce mitotic arrest in response to diverse checkpoint-activating signals

Junwon Kim, John Jeong, Kiwon Song

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

During mitosis, genomic integrity is maintained by the proper coordination of anaphase entry and mitotic exit through mitotic checkpoints. In budding yeast, exit from mitosis is triggered by the activation of the small GTPase Tem1p. Bfa1p in association with Bub2p negatively regulates Tem1p in response to spindle damage, spindle misorientation, and DNA damage, resulting in cell cycle arrest. To delineate the Bfa1p domains that respond to distinct checkpoint signals, we constructed 13 Bfa1 deletion mutants. The C-terminal 184 amino acids of Bfa1p (Bfa1-D8391-574) contained the entire capacity of Bfa1p to generate mitotic arrest in response to spindle damage, spindle misorientation, and DNA damage. This domain was also enough to interact with the mitotic exit network proteins Tem1p, Bub2p, and Cdc5p, and to localize to the spindle pole body (SPB). Over-expression of Bfa1-D8391-574 induced late anaphase arrest as efficient as the full-length Bfa1p in a Bub2p-dependent manner. In contrast, the N-terminal portion of Bfa1p (Bfa1-D161-376) could not localize to SPB and did not block mitotic exit in response to diverse checkpoint signals. Bfa1-D161-376 interacted with Tem1p but not with Bub2p and its over-expression partially arrested cells in mitosis in the absence of Bub2p. By random mutagenesis of Bfa1-D8391-574 with hydroxylamine, we isolated a point mutant of D8, D8E438K, which interacts with both Tem1p and Bub2p but cannot respond to checkpoint signals. This mutant also showed reduced efficiency in the localization to SPB. Taken together, our study demonstrated that various checkpoint signals are transmitted to the C-terminal domain of Bfa1 (Bfal-D8391-574) and that Bfa1p localization to SPB is necessary but not sufficient to regulate mitotic exit in response to various checkpoint signals.

Original languageEnglish
Pages (from-to)399-418
Number of pages20
JournalGenes to Cells
Volume9
Issue number5
DOIs
Publication statusPublished - 2004 May 1

Fingerprint

Spindle Pole Bodies
Saccharomycetales
Mitosis
Anaphase
DNA Damage
M Phase Cell Cycle Checkpoints
Hydroxylamine
Monomeric GTP-Binding Proteins
Cell Cycle Checkpoints
Mutagenesis
Amino Acids
Proteins

All Science Journal Classification (ASJC) codes

  • Genetics
  • Cell Biology

Cite this

@article{a742ccbb62c84ae29a2a18936a26a069,
title = "The C-terminus of Bfa1p in budding yeast is essential to induce mitotic arrest in response to diverse checkpoint-activating signals",
abstract = "During mitosis, genomic integrity is maintained by the proper coordination of anaphase entry and mitotic exit through mitotic checkpoints. In budding yeast, exit from mitosis is triggered by the activation of the small GTPase Tem1p. Bfa1p in association with Bub2p negatively regulates Tem1p in response to spindle damage, spindle misorientation, and DNA damage, resulting in cell cycle arrest. To delineate the Bfa1p domains that respond to distinct checkpoint signals, we constructed 13 Bfa1 deletion mutants. The C-terminal 184 amino acids of Bfa1p (Bfa1-D8391-574) contained the entire capacity of Bfa1p to generate mitotic arrest in response to spindle damage, spindle misorientation, and DNA damage. This domain was also enough to interact with the mitotic exit network proteins Tem1p, Bub2p, and Cdc5p, and to localize to the spindle pole body (SPB). Over-expression of Bfa1-D8391-574 induced late anaphase arrest as efficient as the full-length Bfa1p in a Bub2p-dependent manner. In contrast, the N-terminal portion of Bfa1p (Bfa1-D161-376) could not localize to SPB and did not block mitotic exit in response to diverse checkpoint signals. Bfa1-D161-376 interacted with Tem1p but not with Bub2p and its over-expression partially arrested cells in mitosis in the absence of Bub2p. By random mutagenesis of Bfa1-D8391-574 with hydroxylamine, we isolated a point mutant of D8, D8E438K, which interacts with both Tem1p and Bub2p but cannot respond to checkpoint signals. This mutant also showed reduced efficiency in the localization to SPB. Taken together, our study demonstrated that various checkpoint signals are transmitted to the C-terminal domain of Bfa1 (Bfal-D8391-574) and that Bfa1p localization to SPB is necessary but not sufficient to regulate mitotic exit in response to various checkpoint signals.",
author = "Junwon Kim and John Jeong and Kiwon Song",
year = "2004",
month = "5",
day = "1",
doi = "10.1111/j.1356-9597.2004.00731.x",
language = "English",
volume = "9",
pages = "399--418",
journal = "Genes to Cells",
issn = "1356-9597",
publisher = "Wiley-Blackwell",
number = "5",

}

The C-terminus of Bfa1p in budding yeast is essential to induce mitotic arrest in response to diverse checkpoint-activating signals. / Kim, Junwon; Jeong, John; Song, Kiwon.

In: Genes to Cells, Vol. 9, No. 5, 01.05.2004, p. 399-418.

Research output: Contribution to journalArticle

TY - JOUR

T1 - The C-terminus of Bfa1p in budding yeast is essential to induce mitotic arrest in response to diverse checkpoint-activating signals

AU - Kim, Junwon

AU - Jeong, John

AU - Song, Kiwon

PY - 2004/5/1

Y1 - 2004/5/1

N2 - During mitosis, genomic integrity is maintained by the proper coordination of anaphase entry and mitotic exit through mitotic checkpoints. In budding yeast, exit from mitosis is triggered by the activation of the small GTPase Tem1p. Bfa1p in association with Bub2p negatively regulates Tem1p in response to spindle damage, spindle misorientation, and DNA damage, resulting in cell cycle arrest. To delineate the Bfa1p domains that respond to distinct checkpoint signals, we constructed 13 Bfa1 deletion mutants. The C-terminal 184 amino acids of Bfa1p (Bfa1-D8391-574) contained the entire capacity of Bfa1p to generate mitotic arrest in response to spindle damage, spindle misorientation, and DNA damage. This domain was also enough to interact with the mitotic exit network proteins Tem1p, Bub2p, and Cdc5p, and to localize to the spindle pole body (SPB). Over-expression of Bfa1-D8391-574 induced late anaphase arrest as efficient as the full-length Bfa1p in a Bub2p-dependent manner. In contrast, the N-terminal portion of Bfa1p (Bfa1-D161-376) could not localize to SPB and did not block mitotic exit in response to diverse checkpoint signals. Bfa1-D161-376 interacted with Tem1p but not with Bub2p and its over-expression partially arrested cells in mitosis in the absence of Bub2p. By random mutagenesis of Bfa1-D8391-574 with hydroxylamine, we isolated a point mutant of D8, D8E438K, which interacts with both Tem1p and Bub2p but cannot respond to checkpoint signals. This mutant also showed reduced efficiency in the localization to SPB. Taken together, our study demonstrated that various checkpoint signals are transmitted to the C-terminal domain of Bfa1 (Bfal-D8391-574) and that Bfa1p localization to SPB is necessary but not sufficient to regulate mitotic exit in response to various checkpoint signals.

AB - During mitosis, genomic integrity is maintained by the proper coordination of anaphase entry and mitotic exit through mitotic checkpoints. In budding yeast, exit from mitosis is triggered by the activation of the small GTPase Tem1p. Bfa1p in association with Bub2p negatively regulates Tem1p in response to spindle damage, spindle misorientation, and DNA damage, resulting in cell cycle arrest. To delineate the Bfa1p domains that respond to distinct checkpoint signals, we constructed 13 Bfa1 deletion mutants. The C-terminal 184 amino acids of Bfa1p (Bfa1-D8391-574) contained the entire capacity of Bfa1p to generate mitotic arrest in response to spindle damage, spindle misorientation, and DNA damage. This domain was also enough to interact with the mitotic exit network proteins Tem1p, Bub2p, and Cdc5p, and to localize to the spindle pole body (SPB). Over-expression of Bfa1-D8391-574 induced late anaphase arrest as efficient as the full-length Bfa1p in a Bub2p-dependent manner. In contrast, the N-terminal portion of Bfa1p (Bfa1-D161-376) could not localize to SPB and did not block mitotic exit in response to diverse checkpoint signals. Bfa1-D161-376 interacted with Tem1p but not with Bub2p and its over-expression partially arrested cells in mitosis in the absence of Bub2p. By random mutagenesis of Bfa1-D8391-574 with hydroxylamine, we isolated a point mutant of D8, D8E438K, which interacts with both Tem1p and Bub2p but cannot respond to checkpoint signals. This mutant also showed reduced efficiency in the localization to SPB. Taken together, our study demonstrated that various checkpoint signals are transmitted to the C-terminal domain of Bfa1 (Bfal-D8391-574) and that Bfa1p localization to SPB is necessary but not sufficient to regulate mitotic exit in response to various checkpoint signals.

UR - http://www.scopus.com/inward/record.url?scp=2942668035&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=2942668035&partnerID=8YFLogxK

U2 - 10.1111/j.1356-9597.2004.00731.x

DO - 10.1111/j.1356-9597.2004.00731.x

M3 - Article

C2 - 15147270

AN - SCOPUS:2942668035

VL - 9

SP - 399

EP - 418

JO - Genes to Cells

JF - Genes to Cells

SN - 1356-9597

IS - 5

ER -