Cellular Control of Cortical Actin Nucleation

Abstract

SummaryThe contractile actin cortex is a thin layer of actin, myosin, and actin-binding proteins that subtends the membrane of animal cells. The cortex is the main determinant of cell shape and plays a fundamental role in cell division [1–3], migration [4], and tissue morphogenesis [5]. For example, cortex contractility plays a crucial role in amoeboid migration of metastatic cells [6] and during division, where its misregulation can lead to aneuploidy [7]. Despite its importance, our knowledge of the cortex is poor, and even the proteins nucleating it remain unknown, though a number of candidates have been proposed based on indirect evidence [8–15]. Here, we used two independent approaches to identify cortical actin nucleators: a proteomic analysis using cortex-rich isolated blebs, and a localization/small hairpin RNA (shRNA) screen searching for phenotypes with a weakened cortex or altered contractility. This unbiased study revealed that two proteins generated the majority of cortical actin: the formin mDia1 and the Arp2/3 complex. Each nucleator contributed a similar amount of F-actin to the cortex but had very different accumulation kinetics. Electron microscopy examination revealed that each nucleator affected cortical network architecture differently. mDia1 depletion led to failure in division, but Arp2/3 depletion did not. Interestingly, despite not affecting division on its own, Arp2/3 inhibition potentiated the effect of mDia1 depletion. Our findings indicate that the bulk of the actin cortex is nucleated by mDia1 and Arp2/3 and suggest a mechanism for rapid fine-tuning of cortex structure and mechanics by adjusting the relative contribution of each nucleator.

Membrane bleb: A seesaw game of two small GTPases

“…In addition to the activation of ezrin, active RhoA also activates another RhoA effector protein, formins, which are essential for the regrowth of actin filaments at the retracting membrane. 18,19 In the expanding phase of membrane blebs, we found that Rnd3 is recruited to the expanding plasma membrane. The membrane localization of Rnd3 was gradually lost when reassembly of actin filaments started (Fig.…”

“…Subsequently, Stastna et al and Bovellan et al demonstrated that formin family proteins, key regulators of actin nucleation, are recruited to the membrane blebs and essential for the reassembly of actin filaments. 18,19 In addition, Logue et al showed that Eps8, a plus end capping protein, is recruited to the plasma membrane on a similar timescale as ezrin and essential for the formation of membrane blebs. 20 Thus, important regulators of actin cortex during membrane blebs were identified; however how these proteins are coordinated during reassembly of actin filaments in membrane blebs has remained unclear.…”

Cytoskeletal control of B cell responses to antigens

“…The cortical cytoskeleton is generated by both ARP2/3 and formin activity at the plasma membrane, creating meshwork of short filaments [12][13][14] . The meshwork is approximately 100 nm thick, has a pore size of about 50 nm and attaches to the plasma membrane via ezrin, moesin and class I myosins [15][16][17] .…”

Actomyosin dynamics drive local membrane component organization in an in vitro active composite layer

“…The membrane models mentioned above have by-and-large neglected this active nature of the actin cortex where actin filaments are being continuously polymerized and depolymerized (18-21), in addition to being persistently acted upon by a variety of myosin motors (22-24) that consume ATP and exert contractile stresses on cortical actin filaments, continually remodeling the architecture of the cortex (4, 21, 25). These active processes in turn can generate tangential stresses and currents on the cell surface, which could drive the dynamics and local composition of membrane components at different scales (22,(26)(27)(28)(29).Actin polymerization is proposed to be driven at the membrane by two nucleators, the Arp2/3 complex, which creates a densely branched network, as well as formins that nucleate filaments (18,21,30). A number of myosin motors are also associated with the juxtamembranous actin cortex, of which nonmuscle myosin II is the major component in remodeling the cortex and creating actin flows (4,23,25,26,31,32).…”

“…The membrane models mentioned above have by-and-large neglected this active nature of the actin cortex where actin filaments are being continuously polymerized and depolymerized (18)(19)(20)(21), in addition to being persistently acted upon by a variety of myosin motors (22)(23)(24) that consume ATP and exert contractile stresses on cortical actin filaments, continually remodeling the architecture of the cortex (4,21,25). These active processes in turn can generate tangential stresses and currents on the cell surface, which could drive the dynamics and local composition of membrane components at different scales (22,(26)(27)(28)(29).…”