Limits...
CSC-1: a subunit of the Aurora B kinase complex that binds to the survivin-like protein BIR-1 and the incenp-like protein ICP-1.

Romano A, Guse A, Krascenicova I, Schnabel H, Schnabel R, Glotzer M - J. Cell Biol. (2003)

Bottom Line: CSC-1 associates with ICP-1, BIR-1, and AIR-2 in vivo.ICP-1 dramatically stimulates AIR-2 kinase activity.This activity is not stimulated by CSC-1/BIR-1, suggesting that these two subunits function as targeting subunits for AIR-2 kinase.

View Article: PubMed Central - PubMed

Affiliation: Research Institute of Molecular Pathology, Dr. Bohrgasse 7, Vienna A-1030, Austria.

ABSTRACT
The Aurora B kinase complex is a critical regulator of chromosome segregation and cytokinesis. In Caenorhabditis elegans, AIR-2 (Aurora B) function requires ICP-1 (Incenp) and BIR-1 (Survivin). In various systems, Aurora B binds to orthologues of these proteins. Through genetic analysis, we have identified a new subunit of the Aurora B kinase complex, CSC-1. C. elegans embryos depleted of CSC-1, AIR-2, ICP-1, or BIR-1 have identical phenotypes. CSC-1, BIR-1, and ICP-1 are interdependent for their localization, and all are required for AIR-2 localization. In vitro, CSC-1 binds directly to BIR-1. The CSC-1/BIR-1 complex, but not the individual subunits, associates with ICP-1. CSC-1 associates with ICP-1, BIR-1, and AIR-2 in vivo. ICP-1 dramatically stimulates AIR-2 kinase activity. This activity is not stimulated by CSC-1/BIR-1, suggesting that these two subunits function as targeting subunits for AIR-2 kinase.

Show MeSH
CSC-1 associates with ABI complex members in vivo and in vitro. (A) Whole cell extracts were prepared from wild-type embryos and immunoprecipitated with anti–CSC-1 antibodies. Western blots of the immunoprecipitates reveal that BIR-1 and ICP-1 associate with CSC-1. The coprecipitation of ICP-1 and BIR-1 is specific because it required anti–CSC-1 antibodies (lane 3) and embryonic lysates (lane 4). (B) CSC-1 binds to BIR-1 in vitro, and this interaction requires Zinc binding by BIR-1. Zinc chelation prevents the CSC-1/BIR-1 interaction. Mutation of one of the cysteine residues of BIR-1 (C83) predicted to coordinate zinc abolishes the interaction with CSC-1. CSC-1 and BIR-1 were translated in vitro in the presence of [35S]methionine and mixed in the presence of the indicated compounds. Immunoprecipitations were run on SDS-PAGE and the radioactive products detected with a phosphoimager. Luciferase (Luc) was included as a nonspecific control for the binding reactions.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2172917&req=5

fig4: CSC-1 associates with ABI complex members in vivo and in vitro. (A) Whole cell extracts were prepared from wild-type embryos and immunoprecipitated with anti–CSC-1 antibodies. Western blots of the immunoprecipitates reveal that BIR-1 and ICP-1 associate with CSC-1. The coprecipitation of ICP-1 and BIR-1 is specific because it required anti–CSC-1 antibodies (lane 3) and embryonic lysates (lane 4). (B) CSC-1 binds to BIR-1 in vitro, and this interaction requires Zinc binding by BIR-1. Zinc chelation prevents the CSC-1/BIR-1 interaction. Mutation of one of the cysteine residues of BIR-1 (C83) predicted to coordinate zinc abolishes the interaction with CSC-1. CSC-1 and BIR-1 were translated in vitro in the presence of [35S]methionine and mixed in the presence of the indicated compounds. Immunoprecipitations were run on SDS-PAGE and the radioactive products detected with a phosphoimager. Luciferase (Luc) was included as a nonspecific control for the binding reactions.

Mentions: As CSC-1, BIR-1, and ICP-1 are interdependent for their localization, these proteins likely form a protein complex in vivo. To examine whether such a complex exists, we used Western blotting to determine if anti–CSC-1 immunoprecipitates (IPs) contain ABI complex members. Indeed, both BIR-1 and ICP-1 associate with CSC-1 in C. elegans embryo extracts (Fig. 4 A). We conclude that in vivo, CSC-1 is a member of the ABI complex.


CSC-1: a subunit of the Aurora B kinase complex that binds to the survivin-like protein BIR-1 and the incenp-like protein ICP-1.

Romano A, Guse A, Krascenicova I, Schnabel H, Schnabel R, Glotzer M - J. Cell Biol. (2003)

CSC-1 associates with ABI complex members in vivo and in vitro. (A) Whole cell extracts were prepared from wild-type embryos and immunoprecipitated with anti–CSC-1 antibodies. Western blots of the immunoprecipitates reveal that BIR-1 and ICP-1 associate with CSC-1. The coprecipitation of ICP-1 and BIR-1 is specific because it required anti–CSC-1 antibodies (lane 3) and embryonic lysates (lane 4). (B) CSC-1 binds to BIR-1 in vitro, and this interaction requires Zinc binding by BIR-1. Zinc chelation prevents the CSC-1/BIR-1 interaction. Mutation of one of the cysteine residues of BIR-1 (C83) predicted to coordinate zinc abolishes the interaction with CSC-1. CSC-1 and BIR-1 were translated in vitro in the presence of [35S]methionine and mixed in the presence of the indicated compounds. Immunoprecipitations were run on SDS-PAGE and the radioactive products detected with a phosphoimager. Luciferase (Luc) was included as a nonspecific control for the binding reactions.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172917&req=5

fig4: CSC-1 associates with ABI complex members in vivo and in vitro. (A) Whole cell extracts were prepared from wild-type embryos and immunoprecipitated with anti–CSC-1 antibodies. Western blots of the immunoprecipitates reveal that BIR-1 and ICP-1 associate with CSC-1. The coprecipitation of ICP-1 and BIR-1 is specific because it required anti–CSC-1 antibodies (lane 3) and embryonic lysates (lane 4). (B) CSC-1 binds to BIR-1 in vitro, and this interaction requires Zinc binding by BIR-1. Zinc chelation prevents the CSC-1/BIR-1 interaction. Mutation of one of the cysteine residues of BIR-1 (C83) predicted to coordinate zinc abolishes the interaction with CSC-1. CSC-1 and BIR-1 were translated in vitro in the presence of [35S]methionine and mixed in the presence of the indicated compounds. Immunoprecipitations were run on SDS-PAGE and the radioactive products detected with a phosphoimager. Luciferase (Luc) was included as a nonspecific control for the binding reactions.
Mentions: As CSC-1, BIR-1, and ICP-1 are interdependent for their localization, these proteins likely form a protein complex in vivo. To examine whether such a complex exists, we used Western blotting to determine if anti–CSC-1 immunoprecipitates (IPs) contain ABI complex members. Indeed, both BIR-1 and ICP-1 associate with CSC-1 in C. elegans embryo extracts (Fig. 4 A). We conclude that in vivo, CSC-1 is a member of the ABI complex.

Bottom Line: CSC-1 associates with ICP-1, BIR-1, and AIR-2 in vivo.ICP-1 dramatically stimulates AIR-2 kinase activity.This activity is not stimulated by CSC-1/BIR-1, suggesting that these two subunits function as targeting subunits for AIR-2 kinase.

View Article: PubMed Central - PubMed

Affiliation: Research Institute of Molecular Pathology, Dr. Bohrgasse 7, Vienna A-1030, Austria.

ABSTRACT
The Aurora B kinase complex is a critical regulator of chromosome segregation and cytokinesis. In Caenorhabditis elegans, AIR-2 (Aurora B) function requires ICP-1 (Incenp) and BIR-1 (Survivin). In various systems, Aurora B binds to orthologues of these proteins. Through genetic analysis, we have identified a new subunit of the Aurora B kinase complex, CSC-1. C. elegans embryos depleted of CSC-1, AIR-2, ICP-1, or BIR-1 have identical phenotypes. CSC-1, BIR-1, and ICP-1 are interdependent for their localization, and all are required for AIR-2 localization. In vitro, CSC-1 binds directly to BIR-1. The CSC-1/BIR-1 complex, but not the individual subunits, associates with ICP-1. CSC-1 associates with ICP-1, BIR-1, and AIR-2 in vivo. ICP-1 dramatically stimulates AIR-2 kinase activity. This activity is not stimulated by CSC-1/BIR-1, suggesting that these two subunits function as targeting subunits for AIR-2 kinase.

Show MeSH