The intercalated disc protein, mXinα, is capable of interacting with β-catenin and bundling actin filaments

Sunju Choi, Elisabeth A. Gustafson-Wagner, Qinchuan Wang, Shannon M. Harlan, Haley W. Sinn, Jenny L C Lin, Jim J C Lin

Research output: Contribution to journalArticle

Abstract

Targeted deletion of mXinα results in cardiac hypertrophy and cardiomyopathy with conduction defects (Gustafson-Wagner, E., Sinn, H. W., Chen, Y.-L., Wang, D.-Z., Reiter, R. S., Lin, J. L.-C., Yang, B., Williamson, R. A., Chen, J. N., Lin, C.-I., and Lin, J. J.-C. (2007) Am. J. Physiol. 293, H2680-H2692). To understand the underlying mechanisms leading to such cardiac defects, the functional domains of mXinα and its interacting proteins were investigated. Interaction studies using co-immunoprecipitation, pull-down, and yeast two-hybrid assays revealed that mXinα directly interacts with β-catenin. The β-catenin-binding site on mXinα was mapped to amino acids 535-636, which overlaps with the known actin-binding domains composed of the Xin repeats. The overlapping nature of these domains provides insight into the molecular mechanism for mXinα localization and function. Purified recombinant glutathione S-transferase- or His-tagged mXinα proteins are capable of binding and bundling actin filaments, as determined by co-sedimentation and electron microscopic studies. The binding to actin was saturated at an approximate stoichiometry of nine actin monomers to one mXinα. A stronger interaction was observed between mXinα C-terminal deletion and actin as compared with the interaction between full-length mXinα and actin. Furthermore, force expression of green fluorescent protein fused to an mXinα C-terminal deletion in cultured cells showed greater stress fiber localization compared with force-expressed GFP-mXinα. These results suggest a model whereby the C terminus of mXinα may prevent the full-length molecule from binding to actin, until the β-catenin- binding domain is occupied by β-catenin. The binding of mXinα to β-catenin at the adherens junction would then facilitate actin binding. In support of this model, we found that the actin binding and bundling activity of mXinα was enhanced in the presence of β-catenin.

Original languageEnglish (US)
Pages (from-to)36024-36036
Number of pages13
JournalJournal of Biological Chemistry
Volume282
Issue number49
DOIs
StatePublished - Dec 7 2007
Externally publishedYes

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Catenins
Actin Cytoskeleton
Actins
Proteins
Adherens Junctions
Stress Fibers
Two-Hybrid System Techniques
Defects
Cardiomegaly
Green Fluorescent Proteins
Glutathione Transferase
Cardiomyopathies
Immunoprecipitation
Protein Binding
Sedimentation
Stoichiometry
Cultured Cells
Yeast
Assays
Binding Sites

ASJC Scopus subject areas

  • Biochemistry

Cite this

Choi, S., Gustafson-Wagner, E. A., Wang, Q., Harlan, S. M., Sinn, H. W., Lin, J. L. C., & Lin, J. J. C. (2007). The intercalated disc protein, mXinα, is capable of interacting with β-catenin and bundling actin filaments. Journal of Biological Chemistry, 282(49), 36024-36036. https://doi.org/10.1074/jbc.M707639200

The intercalated disc protein, mXinα, is capable of interacting with β-catenin and bundling actin filaments. / Choi, Sunju; Gustafson-Wagner, Elisabeth A.; Wang, Qinchuan; Harlan, Shannon M.; Sinn, Haley W.; Lin, Jenny L C; Lin, Jim J C.

In: Journal of Biological Chemistry, Vol. 282, No. 49, 07.12.2007, p. 36024-36036.

Research output: Contribution to journalArticle

Choi, Sunju ; Gustafson-Wagner, Elisabeth A. ; Wang, Qinchuan ; Harlan, Shannon M. ; Sinn, Haley W. ; Lin, Jenny L C ; Lin, Jim J C. / The intercalated disc protein, mXinα, is capable of interacting with β-catenin and bundling actin filaments. In: Journal of Biological Chemistry. 2007 ; Vol. 282, No. 49. pp. 36024-36036.
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abstract = "Targeted deletion of mXinα results in cardiac hypertrophy and cardiomyopathy with conduction defects (Gustafson-Wagner, E., Sinn, H. W., Chen, Y.-L., Wang, D.-Z., Reiter, R. S., Lin, J. L.-C., Yang, B., Williamson, R. A., Chen, J. N., Lin, C.-I., and Lin, J. J.-C. (2007) Am. J. Physiol. 293, H2680-H2692). To understand the underlying mechanisms leading to such cardiac defects, the functional domains of mXinα and its interacting proteins were investigated. Interaction studies using co-immunoprecipitation, pull-down, and yeast two-hybrid assays revealed that mXinα directly interacts with β-catenin. The β-catenin-binding site on mXinα was mapped to amino acids 535-636, which overlaps with the known actin-binding domains composed of the Xin repeats. The overlapping nature of these domains provides insight into the molecular mechanism for mXinα localization and function. Purified recombinant glutathione S-transferase- or His-tagged mXinα proteins are capable of binding and bundling actin filaments, as determined by co-sedimentation and electron microscopic studies. The binding to actin was saturated at an approximate stoichiometry of nine actin monomers to one mXinα. A stronger interaction was observed between mXinα C-terminal deletion and actin as compared with the interaction between full-length mXinα and actin. Furthermore, force expression of green fluorescent protein fused to an mXinα C-terminal deletion in cultured cells showed greater stress fiber localization compared with force-expressed GFP-mXinα. These results suggest a model whereby the C terminus of mXinα may prevent the full-length molecule from binding to actin, until the β-catenin- binding domain is occupied by β-catenin. The binding of mXinα to β-catenin at the adherens junction would then facilitate actin binding. In support of this model, we found that the actin binding and bundling activity of mXinα was enhanced in the presence of β-catenin.",
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AU - Gustafson-Wagner, Elisabeth A.

AU - Wang, Qinchuan

AU - Harlan, Shannon M.

AU - Sinn, Haley W.

AU - Lin, Jenny L C

AU - Lin, Jim J C

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