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Actin microfilament aggregation induced by withaferin A is mediated by annexin II

Nature Chemical Biology volume 2, pages 33–38 (2006)Cite this article

Abstract

The actin cytoskeleton supports diverse cellular processes such as endocytosis, oriented growth, adhesion and migration1. The dynamic nature of the cytoskeleton, however, has made it difficult to define the roles of the many accessory molecules that modulate actin organization, especially the multifunctional adapter protein annexin II (refs. 2,3). We now report that the compound withaferin A (1) can alter cytoskeletal architecture in a previously unknown manner by covalently binding annexin II and stimulating its basal F-actin cross-linking activity. Drug-mediated disruption of F-actin organization is dependent on annexin II expression by cells and markedly limits their migratory and invasive capabilities at subcytotoxic concentrations. Given the extensive ethnobotanical history of withaferin-containing plant preparations in the treatment of cancer and inflammatory and neurological disorders, we suggest that annexin II represents a feasible, previously unexploited target for therapeutic intervention by small-molecule drugs4.

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Figure 1: Stable and specific binding of annexin II by withaferin.
Figure 2: Molecular modeling of the withaferin binding site in human annexin II.
Figure 3: Effect of withaferin on AIIt function in vitro.
Figure 4: Withaferin disrupts the actin cytoskeleton in an annexin II–dependent manner.
Figure 5: Effects of withaferin on intact cells.

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Acknowledgements

We thank V. Gerke and J. Ayala-Sanmartin for their gifts of recombinant annexin II and p11 expression plasmids; D.M. Waisman for human recombinant AIIt; G. Tsaprailis and C. Payne for suggestions; and the University of Arizona Electron Microscopy Core; the SWEHSC Proteomics Core and C. McLellan for technical assistance. The assistance of K. Wijeratne in obtaining physical and spectral data for biotinylated withaferin A is also appreciated. This work was supported by US National Institutes of Health (NIH) grants R21-CA091056 (L.W.) and R01-CA090265 (A.A.L.G.). Fellowship support was provided to R.F. by the Tee Up for Tots Foundation. Support for the Southwest Environmental Health Sciences Center was provided by National Institute of Environmental Health Sciences grant ES06694 and NIH/National Cancer Institute grant CA023074-26.

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Authors and Affiliations

  1. Cancer Biology Interdisciplinary Graduate Program University of Arizona Health Sciences Center, 1501 North Campbell Avenue, Tucson, 85724, Arizona, USA

    Ryan R Falsey, Nikhil Shirahatti, Daruka Mahadevan, A A Leslie Gunatilaka & Luke Whitesell

  2. Steele Memorial Children's Research Center, University of Arizona Health Sciences Center, 1501 North Campbell Avenue, Tucson, 85724, Arizona, USA

    Ryan R Falsey, Marilyn T Marron & Luke Whitesell

  3. Southwest Center for Natural Products Research, University of Arizona, 250 East Valencia Road, Tucson, 85706, Arizona, USA

    G M Kamal B Gunaherath & A A Leslie Gunatilaka

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Correspondence to Luke Whitesell.

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Supplementary information

Supplementary Fig. 1

Effect of withaferin (WA) on cell morphology and survival. (PDF 2456 kb)

Supplementary Fig. 2

Design and synthesis of biotinyl-withaferin (B-WA) for solid phase immobilization. (PDF 582 kb)

Supplementary Fig. 3

Identification of annexin II (AII) as a target of withaferin (WA). (PDF 469 kb)

Supplementary Fig. 4

Withaferin (WA) binds to AIIt but not other annexin family members. (PDF 2241 kb)

Supplementary Fig. 5

5 Mass spectroscopy identification of withaferin (WA) adduct site in annexin II (AII). (PDF 467 kb)

Supplementary Fig. 6

Enforced expression of AIIt in HepG2 cells. (PDF 496 kb)

Supplementary Methods (PDF 70 kb)

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Falsey, R., Marron, M., Gunaherath, G. et al. Actin microfilament aggregation induced by withaferin A is mediated by annexin II. Nat Chem Biol 2, 33–38 (2006). https://doi.org/10.1038/nchembio755

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