Constricting Membranes at the Nano and Micro Scale

COMMENTARY

Constricting membranes at the nano and micro scale

Olivia L. Moorena and Dorothy A. Schaferb,1 aDepartment of Cell Biology, Washington University School of Medicine, St. Louis, MO 63110; and bDepartments of Biology and Cell Biology, University of Virginia, Charlottesville, VA 22904

ynamin GTPases regulate membrane traffic, in part, by constricting nanometer-sized membrane tubes to form vesi- clesD by fission. In this issue of PNAS, Chua et al. (1) report that dynamin2 remodels membranes on a much grander scale by influencing the contractile activity of actomyosin encircling epithelial cells at their apical cell–cell junctions. Using a simple epithelial cell system, Madin-Darby canine kidney (MDCK) epithelial cells, Chua et al. Fig. 1. Dynamin GTPase constricts membranes during endocytosis and at epithelial cell apical junctions. found that apical junctional complexes Dynamins promote ﬁssion of Ϸ50-nm-diameter membrane tubes to form clathrin-coated vesicles (on that contain the cytoskeletal linker pro- right). As described by Chua et al. (1), epithelial cells expressing the mutant dynamin2 protein, dynamin2- tein, ZO-1, and elements of the basolat- K44A (blue cell), become highly constricted, suggesting a new role for dynamin2 in regulating the eral F-actin cytoskeleton, were missing actomyosin cytoskeleton. or poorly organized in cells depleted of dynamin2. Notably, in MDCK cells expressing the GTP-binding-defective mu- portantly, apical constrictions induced tions of cortactin with actin filaments tant protein, dynamin2-K44A, polarized by dynamin2-K44A required dynamic were required for robust constriction, membrane domains defined by ZO-1- actin filaments and active myosin II, be- but binding to Arp2/3 complex was not. containing apical junctions formed, but cause constriction was inhibited by Because dynamin2 is a tetramer and oli- the circumferential actomyosin network agents that perturb filament assembly gomerizes (10, 11), complexes of dy- associated with junctional complexes and turnover or block myosin II ATPase namin2-K44A and cortactin could form was constricted Ͼ2-fold over that in activity. a multivalent scaffold capable of inter- cells expressing wild-type dynamin2 Contractile actomyosin networks at acting with multiple actin filaments. (Fig. 1). Together, these finding sug- epithelial apical junctional boundaries Both dynamin2-K44A and cortactin are gested that dynamin2 provides cues that maintain a dynamic, yet invincible, epi- enriched at apical junctions (1) where establish and maintain the polarized epithelial sheet. These networks drive mor- cortactin could interact with the junc- thelial phenotype. Moreover, they sug- phogenesis of sheets, heal their wounds, tional proteins, ZO-1 or p120-catenin gested that dynamin2 influences the orchestrate cell divisions, and extrude (12, 13). Thus, cortactin could bridge contractile activity of apical junctional apoptotic cells, while at the same time, actin filaments of actomyosin and actomyosin networks. preserving a cellular barrier that is the membrane-associated proteins enriched Because dynamins are well-known hallmark of epithelial tissues (6). Con- at apical junctions. Cortactin’s capacity regulators of endocytosis and membrane traction by these networks depends on to respond to and integrate a variety of recycling pathways (2), and establishing myosin motors that slide actin filaments cellular signals might further enhance and maintaining polarized membrane and actin polymers organized into ap- the actions of such a dynamin2– domains and apical junctions requires an propriate networks by filament cross- cortactin–F-actin scaffold (14). intricate balance of endocytosis and in- linking proteins (7, 8). Connections of A growing body of evidence impli- tracellular membrane traffic (3–5), it is the actin network to the plasma mem- cates dynamins as actin filament regula- not surprising that dynamin2 would be brane are also required to effect changes tors. At the biochemical level, dynamin involved in orchestrating epithelial cell in cell shape. In many systems, Rho- family proteins share many binding part- polarity. However, Chua et al. (1) focus family GTPases and their effectors regu- ners that associate either directly, like on the unexpected participation of dy- late assembly and activity of actomyosin cortactin, or indirectly with actin fila- namin2 in regulating contractility of the networks. ments via their regulators (15). Many apical junctional actomyosin network. The intriguing findings by Chua et al. dynamin-associated proteins intersect Extreme apical constriction was only (1) prompt several questions about pathways that promote de novo filament induced in MDCK cells when mutant mechanisms by which the large GTPase assembly via the Arp2/3 complex. At the dynamin2 proteins having low GTP- dynamin2 influences actomyosin con- cellular level, the functions of a variety binding affinity were expressed. Treat- tractility. Some insight comes from the of cellular structures that depend on ment with the dynamin inhibitor dyna- finding that the F-actin binding protein dynamic actin filaments are perturbed in sore or expressing mutant proteins that cortactin is involved in promoting in- cells expressing mutant dynamin pro- stabilize the GTP-bound form of dy- tense apical constrictions in partnership teins, most often dynamin2-K44A (15). namin2 did not promote apical constric- with dynamin2-K44A. Cortactin associ- tions. Although these reagents inhibit ates with dynamin2 via interactions of endocytic pathways, Chua et al. showed the cortactin Src homology 3 domain Author contributions: O.L.M. and D.A.S. wrote the paper. that excessive apical constriction did not and dynamin2’s proline-rich domain (9). The authors declare no conﬂict of interest. result when either clathrin-mediated or Apical constriction did not occur in cells See companion article on page 20770. caveolae-dependent endocytosis was expressing a mutant form of cortactin 1To whom correspondence should be addressed. E-mail: blocked using different strategies. Im- that does not bind dynamin2. Interac- [email protected].

www.pnas.org͞cgi͞doi͞10.1073͞pnas.0911630106 PNAS ͉ December 8, 2009 ͉ vol. 106 ͉ no. 49 ͉ 20559–20560 Downloaded by guest on September 29, 2021 Interestingly, a role for dynamin2 in reg- sient needs to shape cellular structures modeling complexes and activities. ulating microtubule dynamics was re- or pull them together. Tension gener- Alternatively, or in addition, local recently reported (16), suggesting that ated by actomyosin networks may be modeling of membranes in response to dynamin’s influence on cytoskeletal pro- transient or temporally regulated to ef- dynamin2 GTPase-dependent lipid in- cesses may be more wide-ranging than fect local changes in cell shape or me- teractions could indirectly influence ac- the actin filament cytoskeleton. chanics. For example, pulsed cycles of tomyosin networks via effects on actin In the end, Chua et al. (1) propose contraction and retraction within indi- regulators whose activities are modu- that a complex of GTP-depleted dy- vidual cells of epithelial sheets were lated directly by lipids (23, 24). namin2 and cortactin act together to recently implicated in driving morphoge- Mature epithelial apical junctions are promote formation of robust actomyosin netic movements in Drosophila embryos highly dynamic structures, in part, be- networks that tightly constrict epithelial (19, 20). Could cycles of GTP hydrolysis cause intense vesicle traffic maintains cell–cell junctions. Complementary bio- junctional integrity and the polarized chemical studies of dynamin2, cortactin, epithelial state. If dynamin2 influences and actin filaments in vitro show that both endocytic traffic and actomyosin, complexes of nucleotide-free dynamin2 dynamin2 inﬂuences the then these dual roles must be coordi- and cortactin cross-link actin filaments contractile activity of nated in space and time. Chua et al. (1) in tight bundles (17). Because efficient propose that coordinating the endocytic contractility depends on both myosin II apical junctional and cytoskeletal activities of dynamin2 motor activity and the extent of filament might depend on dynamin2’s bound cross-linking within the actin network, actomyosin networks. guanine nucleotide. They suggest that one hypothesis to explain how dynamin2 nucleotide-specific interactions of dy- promotes apical constriction is that namin2 and either endocytic or cy- dynamin2-K44A and cortactin cross-link by dynamin2 temporally regulate acto- toskeletal effectors tips dynamin2 junctional actin filaments and, thereby, myosin networks? Consistent with its function toward either endocytosis or tune the actomyosin machinery in favor ability to couple GTP hydrolysis with actomyosin-based contractility. However, of a highly constricted state. Prominent constriction and fission of lipid tubules mechanisms by which dynamin2 juggles actomyosin-containing stress fibers were in vitro (21, 22), GTP hydrolysis by dy- its membrane remodeling and cytoskel- also assembled in fibroblasts expressing namin2 actively remodeled cross-linked etal activities are likely more complex. dynamin1-K44A (18), consistent with actin filaments in vitro (17). Filament Membrane remodeling by dynamins may this hypothesis. In contrast, actomyosin remodeling by dynamin2 GTPase activ- be intricately coordinated with its activi- arrays were poorly organized in osteo- ity changed the spatial organization of ties that influence actin filaments. De- sarcoma cells depleted of dynamin2 cross-linked actin filaments, exposed cades of effort aimed at unraveling the (17). Thus, a role for dynamin2 in or- polymerization-competent filament ends, mechanisms by which dynamin promotes chestrating actomyosin filament net- and influenced the binding of other fila- clathrin-mediated endocytosis will most works is emerging. Exactly where and on ment regulators, including the actin de- certainly provide directions for unraveling which pool of actin filaments dynamin2 polymerization factor cofilin (17). Thus, how dynamin carries out cytoskeletal- and cortactin exert their effects on acto- cycles of GTP hydrolysis by filament- related functions. myosin networks are questions for associated dynamin2 could broadly in- future studies. fluence the architecture, dynamics, and ACKNOWLEDGMENTS. This work is supported by It is also worth considering whether mechanical properties of actin networks. National Institutes of Health/National Institute of General Medical Sciences Grant R01-GM067222 dynamin2 GTPase activity might gener- The large cohort of dynamin2 actin- (to D.A.S.) and Ruth L. Kirschstein National Re- ally influence actomyosin-based forces associated partners has the potential to search Service Award Grant F32-GM083538 that ebb and flow depending on tran- comprise a versatile set of filament re- (to O.L.M.).

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