An aspirational goal in cell biology is de novo synthesis of whole cells, with the expectation that this will reveal principles of spatiotemporal organization. Natural cells are defined by their boundaries, usually lipid bilayer plasma membranes stabilized by external cell walls or cytoskeletal cortices comprising actin. Typically, researchers approach cell synthesis by preparing protein mixtures in bulk, then partitioning them into cell-sized containers using water-in-oil emulsions, giant unilamellar vesicles, or microfabricated chambers. On page 631 of this issue, Cheng and Ferrell (1) describe an alternative approach to cell synthesis, whereby a bulk preparation of concentrated extract from frog eggs self-organizes into regularly spaced, cell-like units.
The resulting preparations resemble natural syncytia, which are large cells in which many nuclei share a common cytoplasm. Like natural syncytia, they are organized by microtubule asters, which are cytoskeletal structures that usually emanate from a centrosome (see the figure). Partitioning of frog egg cytoplasm into cell-like units by microtubule asters was reported previously (2, 3). However, Cheng and Ferrell found that self-organization can occur in the absence of centrosomes and that mitotic division of units can happen when they contain DNA and centrosomes.
Syncytial organization is characteristic of certain protist and fungal species (4). It has been most studied in early fruit fly (Drosophila melanogaster) embryos, for which the first 13 divisions are syncytial. The cell-like units in syncytial D. melanogaster embryos are composed of a nucleus with an associated centrosome, microtubule aster, and organelles. They are called energids and behave in many ways like membrane-delimited cells (5, 6). How energids self-organize, divide, and space out, all while sharing a common cytoplasm, remains unclear.
Cheng and Ferrell reported mitotic division of their cell-like units when nuclei and centrosomes were present, in resemblance to natural syncytia. Similar syncytial divisions were reported in droplets extracted from syncytial D. melanogaster embryos with a micropipette (7), but the frog extract system can be biochemically manipulated. Frog egg extracts have long been used to study cell cycle oscillations driven by cyclin B synthesis and destruction (8). Cycles can continue for many hours if oxygen and carbon dioxide exchange are facilitated by partitioning the extract into cell-like droplets (9). It is exciting to now see this biochemical oscillator drive syncytial mitosis. In fertilized frog eggs, each cell cycle is accompanied by cell division. However, syncytial division can occur when cell division (specifically cytokinesis, when duplicated cells are separated) is blocked (10), so the mechanisms that organize normal cleavage divisions can also generate syncytia, as seen in Cheng and Ferrell’s extract system.
What molecular mechanisms are likely to organize and partition asters in synthetic syncytia and their natural counterparts? In physical systems, assembly of discrete particles or droplets out of a bulk phase can be driven by a simple condensation or aggregation process. It is possible that the syncytial organization observed by Cheng and Ferrell is also driven by some bulk condensation or aggregation process. Egg extract contains high concentrations of endoplasmic reticulum, mitochondria, ribosomes, and glycogen, any of which might spontaneously aggregate. Using chemical inhibitors, Cheng and Ferrell found that emergence of syncytium-like organization depended on microtubules and the minus-end–directed motor protein dynein, pointing to a role of microtubule-based mechanisms rather than bulk aggregation.
Cheng and Ferrell observed microtubule aster formation with or without centrosomes. In frog eggs, asters are nucleated by centrosomes, and their growth to hundreds of microns is further facilitated by microtubule-stimulated nucleation distant from the centrosome (11). Radial asters can also self-assemble by dynein-dependent mechanisms in the absence of centrosomes. These partially understood mechanisms are thought to involve inward transport of microtubule minus ends, and/or microtubule nucleating complexes, by dynein that can be part of a multiprotein complex (12) or attached to vesicles (13).
CHENG AND FERRELL OBSERVED SYNCITIUM-LIKE ORGANIZATION OF FROG EGG EXTRACT, POTENTIALLY PROVIDING AN ALTERNATIVE METHOD OF CELL SYNTHESIS.
GRAPHIC: K. HOLOSKI/SCIENCE
Syncytia also require boundaries to delimit asters and space them out. These boundary-formation mechanisms need only operate directly on microtubules, because organelles are entrained to microtubules by dynein-mediated transport. One known boundary-forming mechanism involves pruning of antiparallel microtubules by the combined action of a microtubule cross-linker [protein regulator of cytokinesis 1 (PRC1)] and a kinesin motor that caps plus ends, KIF4A (14). Another involves inhibition of plus-end growth by aurora kinase B (3), which is transported to aster boundaries by kinesins as part of the chromosomal passenger complex (2). These mechanisms cooperate to partition the microtubule cytoskeleton during cytokinesis and may be involved in the mitotic division of cell-like units observed by Cheng and Ferrell.
Synthetic approaches test hypotheses for cell organization in a manner that is orthogonal to genetic perturbation. Frog egg extract provides a versatile system for cell synthesis, whether in bulk to form syncytia (1, 3) or in droplets to mimic individual cells (9). Individual proteins can be removed or added, and knowledge of the quantitative proteome facilitates systems-level analysis (15). One important caveat is that the frog egg is not an ordinary cell, especially in its huge spatial dimension, and egg organization mechanisms may differ in interesting ways from those in small cells. Cell biologists are urged to consider syncytia in research as well as physically discrete cells. Understanding how energids, microtubule asters, or equivalent organization units assemble, divide, and space out in a shared cytoplasm is likely to reveal organization principles that are relevant to all cells.
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