Abstract
We are much better at taking cells apart than putting them together. Reconstitution of biological processes from component molecules has been a powerful but difficult approach to studying functional organization in biology. Recently, the convergence of biochemical and cell biological advances with new experimental and computational tools is providing the opportunity to reconstitute increasingly complex processes. We predict that this bottom-up strategy will uncover basic processes that guide cellular assembly, advancing both basic and applied sciences.
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References
Groves, J. T. Bending mechanics and molecular organization in biological membranes. Annu. Rev. Phys. Chem. 58, 697–717 (2007).
Garcia-Saez, A. J., Chiantia, S. & Schwille, P. Effect of line tension on the lateral organization of lipid membranes. J. Biol. Chem. 282, 33537–33544 (2007).
Karsenti, E. Self-organization in cell biology: a brief history. Nature Rev. Mol. Cell Biol. 9, 255–262 (2008).
Kron, S. J. & Spudich, J. A. Fluorescent actin filaments move on myosin fixed to a glass surface. Proc. Natl Acad. Sci. USA 83, 6272–6276 (1986).
Hawking, S. W. The Universe in a Nutshell (Bantam, New York, 2001).
Kritikou, E. To see them contract for the first time. Nature Rev. Mol. Cell Biol. 9, S6 (2008).
Watson, J. D. & Crick, F. H. Molecular structure of nucleic acids; a structure for deoxyribose nucleic acid. Nature 171, 737–738 (1953).
Kornberg, A. Biologic synthesis of deoxyribonucleic acid. Science 131, 1503–1508 (1960).
Lehman, I. R., Bessman, M. J., Simms, E. S. & Kornberg, A. Enzymatic synthesis of deoxyribonucleic acid. I. Preparation of substrates and partial purification of an enzyme from Escherichia coli. J. Biol. Chem. 233, 163–170 (1958).
Schekman, R. Protein localization and membrane traffic in yeast. Annu. Rev. Cell Biol. 1, 115–143 (1985).
Orci, L., Glick, B. S. & Rothman, J. E. A new type of coated vesicular carrier that appears not to contain clathrin: its possible role in protein transport within the Golgi stack. Cell 46, 171–184 (1986).
Barlowe, C. et al. COPII: a membrane coat formed by Sec proteins that drive vesicle budding from the endoplasmic reticulum. Cell 77, 895–907 (1994).
Higgins, N. P., Peebles, C. L., Sugino, A. & Cozzarelli, N. R. Purification of subunits of Escherichia coli DNA gyrase and reconstitution of enzymatic activity. Proc. Natl Acad. Sci. USA 75, 1773–1777 (1978).
Gorlich, D., Hartmann, E., Prehn, S. & Rapoport, T. A. A protein of the endoplasmic reticulum involved early in polypeptide translocation. Nature 357, 47–52 (1992).
Nakajima, M. et al. Reconstitution of circadian oscillation of cyanobacterial KaiC phosphorylation in vitro. Science 308, 414–415 (2005).
Surrey, T., Nedelec, F., Leibler, S. & Karsenti, E. Physical properties determining self-organization of motors and microtubules. Science 292, 1167–1171 (2001).
Haviv, L. et al. Reconstitution of the transition from lamellipodium to filopodium in a membrane-free system. Proc. Natl Acad. Sci. USA 103, 4906–4911 (2006).
Karsenti, E., Nedelec, F. & Surrey, T. Modelling microtubule patterns. Nature Cell Biol. 8, 1204–1211 (2006).
Pollard, T. D. The cytoskeleton, cellular motility and the reductionist agenda. Nature 422, 741–745 (2003).
Heald, R. et al. Self-organization of microtubules into bipolar spindles around artificial chromosomes in Xenopus egg extracts. Nature 382, 420–425 (1996).
Pistor, S., Chakraborty, T., Niebuhr, K., Domann, E. & Wehland, J. The ActA protein of Listeria monocytogenes acts as a nucleator inducing reorganization of the actin cytoskeleton. EMBO J. 13, 758–763 (1994).
Loisel, T. P., Boujemaa, R., Pantaloni, D. & Carlier, M. F. Reconstitution of actin-based motility of Listeria and Shigella using pure proteins. Nature 401, 613–616 (1999).
Akin, O. & Mullins, R. D. Capping protein increases the rate of actin-based motility by promoting filament nucleation by the Arp2/3 complex. Cell 133, 841–851 (2008).
Vignjevic, D. et al. Formation of filopodia-like bundles in vitro from a dendritic network. J. Cell Biol. 160, 951–962 (2003).
Brieher, W. M., Coughlin, M. & Mitchison, T. J. Fascin-mediated propulsion of Listeria monocytogenes independent of frequent nucleation by the Arp2/3 complex. J. Cell Biol. 165, 233–242 (2004).
Shih, Y. L. & Rothfield, L. The bacterial cytoskeleton. Microbiol. Mol. Biol. Rev. 70, 729–754 (2006).
Garner, E. C., Campbell, C. S., Weibel, D. B. & Mullins, R. D. Reconstitution of DNA segregation driven by assembly of a prokaryotic actin homolog. Science 315, 1270–1274 (2007).
Garner, E. C., Campbell, C. S. & Mullins, R. D. Dynamic instability in a DNA-segregating prokaryotic actin homolog. Science 306, 1021–1025 (2004).
Sackmann, E. & Tanaka, M. Supported membranes on soft polymer cushions: fabrication, characterization and applications. Trends Biotechnol. 18, 58–64 (2000).
Groves, J. T., Wulfing, C. & Boxer, S. G. Electrical manipulation of glycan-phosphatidyl inositol-tethered proteins in planar supported bilayers. Biophys. J. 71, 2716–2723 (1996).
Grakoui, A. et al. The immunological synapse: a molecular machine controlling T cell activation. Science 285, 221–227 (1999).
Pautot, S., Lee, H., Isacoff, E. Y. & Groves, J. T. Neuronal synapse interaction reconstituted between live cells and supported lipid bilayers. Nature Chem. Biol. 1, 283–289 (2005).
Loose, M., Fischer-Friedrich, E., Ries, J., Kruse, K. & Schwille, P. Spatial regulators for bacterial cell division self-organize into surface waves in vitro. Science 320, 789–792 (2008).
Co, C., Wong, D. T., Gierke, S., Chang, V. & Taunton, J. Mechanism of actin network attachment to moving membranes: barbed end capture by N-WASP WH2 domains. Cell 128, 901–913 (2007).
Pucadyil, T. J. & Schmid, S. L. Real-time visualization of dynamin-catalyzed membrane fission and vesicle release. Cell 135, 1263–1275 (2008).
Takei, K. et al. Generation of coated intermediates of clathrin-mediated endocytosis on protein-free liposomes. Cell 94, 131–141 (1998).
Matsuoka, K. et al. COPII-coated vesicle formation reconstituted with purified coat proteins and chemically defined liposomes. Cell 93, 263–275 (1998).
Palade, G. E. The organization of living matter. Proc. Natl Acad. Sci. USA 52, 613–634 (1964).
Roux., A., Uyhazi, K., Frost, A. & De Camilli, P. GTP-dependent twisting of dynamin implicates constriction and tension in membrane fission. Nature 441, 528–531 (2006).
Koster, G., VanDuijn, M., Hofs, B. & Dogterom, M. Membrane tube formation from giant vesicles by dynamic association of motor proteins. Proc. Natl Acad. Sci. USA 100, 15583–15588 (2003).
Giardini, P. A., Fletcher, D. A. & Theriot, J. A. Compression forces generated by actin comet tails on lipid vesicles. Proc. Natl Acad. Sci. USA 100, 6493–6498 (2003).
Upadhyaya, A. & van Oudenaarden, A. Biomimetic systems for studying actin-based motility. Curr. Biol. 13, R734–R744 (2003).
Cortese, J. D., Schwab, B., Frieden, C. & Elson, E. L. Actin polymerization induces a shape change in actin-containing vesicles. Proc. Natl Acad. Sci. USA 86, 5773–5777 (1989).
Fygenson, D. K., Marko, J. F. & Libchaber, A. Mechanics of microtubule-based membrane extension. Phys. Rev. Lett. 79, 4497 (1997).
Pontani, L. L. et al. Reconstitution of an actin cortex inside a liposome. Biophys. J. 96, 192–198 (2009).
Liu, A. P. & Fletcher, D. A. Actin polymerization serves as a membrane domain switch in model lipid bilayers. Biophys. J. 91, 4064–4070 (2006).
Holowka, D., Sheets, E. D. & Baird, B. Interactions between FcɛRI and lipid raft components are regulated by the actin cytoskeleton. J. Cell Sci. 113, 1009–1019 (2000).
Liu, A. P. et al. Membrane-induced bundling of actin filaments. Nature Phys. 4, 789–793 (2008).
Pronk, S., Geissler, P. L. & Fletcher, D. A. Limits of filopodium stability. Phys. Rev. Lett. 100, 258102 (2008).
Osawa, M., Anderson, D. E. & Erickson, H. P. Reconstitution of contractile FtsZ rings in liposomes. Science 320, 792–794 (2008).
Doh, J. & Irvine, D. J. Immunological synapse arrays: patterned protein surfaces that modulate immunological synapse structure formation in T cells. Proc. Natl Acad. Sci. USA 103, 5700–5705 (2006).
Cosentino Lagomarsino, M. et al. Microtubule organization in three-dimensional confined geometries: evaluating the role of elasticity through a combined in vitro and modeling approach. Biophys. J. 92, 1046–1057 (2007).
Fernandez-Suarez, M. & Ting, A. Y. Fluorescent probes for super-resolution imaging in living cells. Nature Rev. Mol. Cell Biol. 9, 929–943 (2008).
Alberts, J. B. & Odell, G. M. In silico reconstitution of Listeria propulsion exhibits nano-saltation. PLoS Biol. 2, e412 (2004).
Pautot, S., Frisken, B. J. & Weitz, D. A. Engineering asymmetric vesicles. Proc. Natl Acad. Sci. USA 100, 10718–10721 (2003).
Noireaux, V. & Libchaber, A. A vesicle bioreactor as a step toward an artificial cell assembly. Proc. Natl Acad. Sci. USA 101, 17669–17674 (2004).
Okushima, S., Nisisako, T., Torii, T. & Higuchi, T. Controlled production of monodisperse double emulsions by two-step droplet breakup in microfluidic devices. Langmuir 20, 9905–9908 (2004).
Utada, A. S. et al. Monodisperse double emulsions generated from a microcapillary device. Science 308, 537–541 (2005).
Shum, H. C., Kim, J. W. & Weitz, D. A. Microfluidic fabrication of monodisperse biocompatible and biodegradable polymersomes with controlled permeability. J. Am. Chem. Soc. 130, 9543–9549 (2008).
Funakoshi, K., Suzuki, H. & Takeuchi, S. Formation of giant lipid vesicle-like compartments from a planar lipid membrane by a pulsed jet flow. J. Am. Chem. Soc. 129, 12608–12609 (2007).
Stachowiak, J. C. et al. Unilamellar vesicle formation and encapsulation by microfluidic jetting. Proc. Natl Acad. Sci. USA 105, 4697–4702 (2008).
Mossman, K. D., Campi, G., Groves, J. T. & Dustin, M. L. Altered TCR signaling from geometrically repatterned immunological synapses. Science 310, 1191–1193 (2005).
Theriot, J. A., Mitchison, T. J., Tilney, L. G. & Portnoy, D. A. The rate of actin-based motility of intracellular Listeria monocytogenes equals the rate of actin polymerization. Nature 357, 257–260 (1992).
Cameron, L. A., Footer, M. J., van Oudenaarden, A. & Theriot, J. A. Motility of ActA protein-coated microspheres driven by actin polymerization. Proc. Natl Acad. Sci. USA 96, 4908–4913 (1999).
Gaetz, J., Gueroui, Z., Libchaber, A. & Kapoor, T. M. Examining how the spatial organization of chromatin signals influences metaphase spindle assembly. Nature Cell Biol. 8, 924–932 (2006).
Roux., A. et al. A minimal system allowing tubulation with molecular motors pulling on giant liposomes. Proc. Natl Acad. Sci. USA 99, 5394–5399 (2002).
Wollert, T., Wunder, C., Lippincott-Schwartz, J. & Hurley, J. H. Membrane scission by the ESCRT-III complex. Nature 458, 172–177 (2009).
Acknowledgements
We apologize to those colleagues whose original and important work could not be cited owing to space limitations. We thank D. Richmond, J. Stachowiak and the rest of the Fletcher laboratory, as well as T. Pucadyil, for helpful discussion. A.P.L. is supported by the Natural Sciences and Engineering Research Council of Canada. D.A.F. is funded by National Institutes of Health R01 grants and a Nanomedicine Development Centre grant.
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Liu, A., Fletcher, D. Biology under construction: in vitro reconstitution of cellular function. Nat Rev Mol Cell Biol 10, 644–650 (2009). https://doi.org/10.1038/nrm2746
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DOI: https://doi.org/10.1038/nrm2746
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