The WASP and WAVE family proteins

A review of the cytoskeleton-organizing WASP and WAVE family proteins

α-Actinin-4 Enhances Colorectal Cancer Cell Invasion by Suppressing Focal Adhesion Maturation

α-Actinins (ACTNs) are known to crosslink actin filaments at focal adhesions in migrating cells. Among the four isoforms of mammalian ACTNs, ACTN1 and ACTN4 are ubiquitously expressed. Recently, ACTN4 was reported to enhance cancer cell motility, invasion, and metastasis. However, the mechanism by which ACTN4 drives these malignant phenotypes remains unclear. Here, we show that ACTN4, but not ACTN1, induces the formation of immature focal adhesions in DLD-1 cells, leading to the rapid turnover of focal adhesions. Interestingly, zyxin (ZYX) assembly to focal adhesions was markedly decreased in ACTN4-expressing DLD-1 cells, while the recruitment of paxillin (PAX) occurred normally. On the other hand, in ACTN1-expressing DLD-1 cells, PAX and ZYX were normally recruited to focal adhesions, suggesting that ACTN4 specifically impairs focal adhesion maturation by inhibiting the recruitment of ZYX to focal complexes. Using purified recombinant proteins, we found that ZYX binding to ACTN4 was defective under conditions where ZYX binding to ACTN1 was observed. Furthermore, Matrigel invasion of SW480 cells that express high endogenous levels of ACTN4 protein was inhibited by ectopic expression of ACTN1. Altogether, our results suggest that ZYX defective binding to ACTN4, which occupies focal adhesions instead of ACTN1, induces the formation of immature focal adhesions, resulting in the enhancement of cell motility and invasion

Optimization of WAVE2 complex–induced actin polymerization by membrane-bound IRSp53, PIP3, and Rac

WAVE2 activates the actin-related protein (Arp) 2/3 complex for Rac-induced actin polymerization during lamellipodium formation and exists as a large WAVE2 protein complex with Sra1/PIR121, Nap1, Abi1, and HSPC300. IRSp53 binds to both Rac and Cdc42 and is proposed to link Rac to WAVE2. We found that the knockdown of IRSp53 by RNA interference decreased lamellipodium formation without a decrease in the amount of WAVE2 complex. Localization of WAVE2 at the cell periphery was retained in IRSp53 knockdown cells. Moreover, activated Cdc42 but not Rac weakened the association between WAVE2 and IRSp53. When we measured Arp2/3 activation in vitro, the WAVE2 complex isolated from the membrane fraction of cells was fully active in an IRSp53-dependent manner but WAVE2 isolated from the cytosol was not. Purified WAVE2 and purified WAVE2 complex were activated by IRSp53 in a Rac-dependent manner with PIP3-containing liposomes. Therefore, IRSp53 optimizes the activity of the WAVE2 complex in the presence of activated Rac and PIP3

DA-Raf1, a competent intrinsic dominant-negative antagonist of the Ras–ERK pathway, is required for myogenic differentiation

Ras activates Raf, leading to the extracellular-regulated kinase (ERK)–mitogen-activated protein kinase pathway, which is involved in a variety of cellular, physiological, and pathological responses. Thus, regulators of this Ras–Raf interaction play crucial roles in these responses. In this study, we report a novel regulator of the Ras–Raf interaction named DA-Raf1. DA-Raf1 is a splicing isoform of A-Raf with a wider tissue distribution than A-Raf. It contains the Ras-binding domain but lacks the kinase domain, which is responsible for activation of the ERK pathway. As inferred from its structure, DA-Raf1 bound to activated Ras as well as M-Ras and interfered with the ERK pathway. The Ras–ERK pathway is essential for the negative regulation of myogenic differentiation induced by growth factors. DA-Raf1 served as a positive regulator of myogenic differentiation by inducing cell cycle arrest, the expression of myogenin and other muscle-specific proteins, and myotube formation. These results imply that DA-Raf1 is the first identified competent, intrinsic, dominant-negative antagonist of the Ras–ERK pathway

Phospholipase Cδ4 is required for Ca2+ mobilization essential for acrosome reaction in sperm

Zona pellucida (ZP)–induced acrosome reaction in sperm is a required step for mammalian fertilization. However, the precise mechanism of the acrosome reaction remains unclear. We previously reported that PLCδ4 is involved in the ZP-induced acrosome reaction in mouse sperm. Here we have monitored Ca2+ responses in single sperm, and we report that the [Ca2+]i increase in response to ZP, which is essential for driving the acrosome reaction in vivo, is absent in PLCδ4−/− sperm. Progesterone, another physiological inducer of the acrosome reaction, failed to induce sustained [Ca2+]i increases in PLCδ4−/− sperm, and consequently the acrosome reaction was partially inhibited. In addition, we observed oscillatory [Ca2+]i increases in wild-type sperm in response to these acrosome inducers. Calcium imaging studies revealed that the [Ca2+]i increases induced by exposure to ZP and progesterone started at different sites within the sperm head, indicating that these agonists induce the acrosome reaction via different Ca2+ mechanisms. Furthermore, store-operated channel (SOC) activity was severely impaired in PLCδ4−/− sperm. These results indicate that PLCδ4 is an important enzyme for intracellular [Ca2+]i mobilization in the ZP-induced acrosome reaction and for sustained [Ca2+]i increases through SOC induced by ZP and progesterone in sperm

Phosphoinositide-binding interface proteins involved in shaping cell membranes

The mechanism by which cell and cell membrane shapes are created has long been a subject of great interest. Among the phosphoinositide-binding proteins, a group of proteins that can change the shape of membranes, in addition to the phosphoinositide-binding ability, has been found. These proteins, which contain membrane-deforming domains such as the BAR, EFC/F-BAR, and the IMD/I-BAR domains, led to inward-invaginated tubes or outward protrusions of the membrane, resulting in a variety of membrane shapes. Furthermore, these proteins not only bind to phosphoinositide, but also to the N-WASP/WAVE complex and the actin polymerization machinery, which generates a driving force to shape the membranes

SH3YL1 regulates dorsal ruffle formation by a novel phosphoinositide-binding domain

The newly identified SYLF lipid-binding domain of SH3YL1 mediates phosphoinositide binding during dorsal membrane morphogenesis