The Faculty


Mechanisms of actin assembly: Real-time observation of recapitulated cytoskeleton structures by single filament microscopy


B.A., Ohio Wesleyan University, 1995
Ph.D., Purdue University, 2001

Research Summary

The focus of our research group is to determine the fundamental biochemical mechanisms that govern how cells coordinate actin filament assembly for a wide range of tasks (Figure 1). Actin filament assembly is required for processes that are necessary for proper development such as division, establishing polarity and migration, yet when unregulated can lead to devastating defects.

Mechanistically diverse nucleating proteins are required to initiate actin polymerization at the right time and place. The actin-related protein Arp2/3 complex initiates the assembly of short-branched filaments that are ideal for pushing on cell membranes and lipids (Figure 1). Recently the paradigm that the Arp2/3 complex is the principle actin filament nucleator in cells was radically shifted. Formins were identified as a second major nucleating factor that participate in a growing list of cellular processes that require long-straight bundled actin filaments including division, polarization and motility. However, many of the molecular details of how formins mediate actin filament assembly remain unclear, and it is not understood how cells separately utilize the Arp2/3 complex and multiple formin isoforms to accomplish diverse fundamental tasks (Figure 1).

Elucidating the mechanisms of complex actin-based processes requires the rigorous characterization of the participating proteins both in and out of live cells. Therefore we are combining in vitro approaches with the genetically tractable fission yeast to utilize diverse techniques from genetics, live cell fluorescence microscopy, biophysics, biochemistry, and innovative single actin filament imaging assays. Utilizing fission yeast allows rapid progress, and given the evolutionary conservation between the participating set of actin-binding proteins in fission yeast and mammals, our studies should pertain directly to healthy and diseased mammalian cells (Figure 1).


Figure 1. The Arp2/3 complex and Formin drive actin filament assembly for diverse cellular processes in both animal and fission yeast cells. (A) Extra- and intra-cellular signals activate nucleating proteins that rapidly assemble actin filaments. In animal cells Formins assemble actin filaments for division (Formin 1) and filopodia in migrating cells (Formin 2). The Arp2/3 complex makes filaments for the lamellipodia of migrating cells. (B) In fission yeast Formins nucleate actin filament assembly for division (Cdc12p) and polarized growth (For3p), whereas the Arp2/3 complex is required for motile endocytic actin patches.

Selected Publications

Anderson, C.A., Kovar, D.R., Gardel, M.L., and Winkelman, J.D. (2021). LIM domain proteins in cell mechanobiology. Cytoskeleton, 1-9. 

Winkelman, J.D.*, Anderson, C.A.*, Suarez, C., Kovar, D.R., and Gardel, M.L. (2020). Evolutionarily diverse LIM domain-containing proteins bind stressed actin filaments through a conserved mechanism. Proceedings of the National Academy of Sciences of the United States of America, 117(41), 25532– 25542. 

Mancl, J. M., Suarez, C., Liang, W. G., Kovar, D. R., and Tang, W.-J. (2020) Pseudomonas aeruginosa exoenzyme Y directly bundles actin filaments. J. Biol. Chem.

Ajeti, V., Tabatabai, A. P., Fleszar, A. J., Staddon, M. F., Seara, D. S., Suarez, C., Yousafzai, M. S., Bi, D., Kovar, D. R., Banerjee, S., and Murrell, M. P. (2019) Wound healing coordinates actin architectures to regulate mechanical work. Nature Physics. 

Christensen, J. R., Craig, E. W., Glista, M. J., Mueller, D. M., Li, Y., Sees, J. A., Huang, S., Suarez, C., Mets, L. J., Kovar, D. R., and Avasthi, P. (2019) Chlamydomonas reinhardtii formin FOR1 and profilin PRF1 are optimized for acute rapid actin filament assembly. MBoC. 

Freedman SL, Suarez C, Winkelman JD, Kovar DR, Voth GA, Dinner AR, Hocky GM (2019). Mechanical and kinetic factors drive sorting of F-actin cross-linkers on bundles. PNAS 

Christensen JR*, Homa KE*, Morganthaler AN, Brown RR, Suarez C, Harker AJ, O’Connell ME, Kovar DR (2019). Cooperation between tropomyosin and α-actinin inhibits fimbrin association with actin filament networks in fission yeast. Elife 

Harker AJ, Katkar HH, Bidone TC, Aydin F, Voth GA, Applewhite DA, Kovar DR (2019). Ena/VASP processive elongation is modulated by avidity on actin filaments bundled by the filopodia crosslinker fascin. Mol Biol Cell. (PubMed)

McCall PM, Srivastava S, Perry SL, Kovar DR, Gardel ML, Tirrell MV (2018). Partitioning and enhanced self-assembly of actin in polypeptide coacervates. Biophys J. 114(7):1636-1645. 

Zimmermann D, Homa KE, Hocky GM, Pollard LW, De La Cruz EM, Voth GA, Trybus KM, Kovar DR (2017). Mechanoregulated inhibition of formin facilitates contractile actomyosin ring assembly. Nat Commun. 26;8(1):703. 

Suarez C, McCall PM, Gardel ML, Kovar DR (2017). When Is “Enough” Enough? Cell Syst. 24;4(5):480-482.

Burke TA, Harker AJ, Dominguez R, Kovar DR (2017). The bacterial virulence factors VopL and VopF nucleate actin from the pointed end. J Cell Biol. 216(5):1267-1276.

Christensen JR, Hocky GM, Homa KE, Morganthaler AN, Hitchock-DeGregori SE, Voth GA, Kovar DR (2017). Competition between Tropomyosin, Fimbrin, and ADF/Cofilin drives their sorting to distinct actin filament networks. Elife. (10;6). 

Suarez C, Kovar DR (2016). Internetwork competition for monomers governs actin cytoskeleton organization. Nat Rev Mol Cell Biol. (12) 799-810. 

Winkelman JD, Suarez C, Hocky GM, Harker AJ, Morganthaler AN, Christensen JR, Voth GA, Bartles JR, Kovar DR (2016). Fascin- and α-Actinin-Bundled Networks Contain Intrinsic Structural Features that Drive Protein Sorting. Curr Biol. 26(20):2697-2706. 

Li Y, Christensen JR, Homa KE, Hocky GM, Fok A, Sees JA, Voth GA, Kovar DR. (2016). The F-actin bundler α-actinin Ain1 is tailored for ring assembly and constriction during cytokinesis in fission yeast. Mol Biol Cell. 1;27(11):1821-33. 

Alberico EO, Zhu ZC, Wu YF, Gardner MK, Kovar DR, Goodson HV. (2016). Interactions between the Microtubule Binding Protein EB1 and F-Actin. J Mol Biol. 428(6):1304-14. 

Zimmermann D, Morganthaler AN, Kovar DR, Suarez C. (2016). In Vitro Biochemical Characterization of Cytokinesis Actin-Binding Proteins. Methods Mol Biol. 1369:151-79. 

Heisler DB, Kudryashova E, Grinevich DO, Suarez C, Winkelman JD, Birukov KG, Kotha SR, Parinandi NL, Vavylonis D, Kovar DR, Kudryashov DS  (2015). ACTIN-DIRECTED TOXIN. ACD toxin-produced actin oligomers poison formin-controlled actin polymerization. Science. 349(6247):535-9.