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In a typical Power Device on the 0.16μm node, the isolation module is one of the most critical steps. The trench to be filled in those devices is rather deep and needs a considerable amount of a suitable dielectric material. The choice of dielectric in the present paper is falling on the SubAtmosphericUndopedSilicaGlass (SAUSG oxide) [1].

Filopodia explore the environment, sensing soluble and mechanical cues during directional motility and tissue morphogenesis. How filopodia are initiated and spatially restricted to specific sites on the plasma membrane is still unclear. Here, we show that the membrane deforming and curvature sensing IRSp53 (Insulin Receptor Substrate of 53 kDa) protein slows down actin filament barbed end growth. This inhibition is relieved by CDC42 and counteracted by VASP, which also binds to IRSp53. The VASP:IRSp53 interaction is regulated by activated CDC42 and promotes high-density clustering of VASP, which is required for processive actin filament elongation. The interaction also mediates VASP recruitment to liposomes. In cells, IRSp53 and VASP accumulate at discrete foci at the leading edge, where filopodia are initiated. Genetic removal of IRSp53 impairs the formation of VASP foci, filopodia and chemotactic motility, while IRSp53 null mice display defective wound healing. Thus, IRSp53 dampens barbed end growth. CDC42 activation inhibits this activity and promotes IRSp53-dependent recruitment and clustering of VASP to drive actin assembly. These events result in spatial restriction of VASP filament elongation for initiation of filopodia during cell migration, invasion, and tissue repair.

The interest towards Copper RDL (Re-Distribution Layer) is due to some advantages related to this approach. First of all it is cheaper than conventional Damascene approach; moreover it allows thicknesses as high as 10µm or more whereas with Damascene architecture Cu thickness is limited to

Mdm2 is the major negative regulator of p53 tumor-suppressor activity. This oncoprotein is overexpressed in many human tumors that retain the wild-type p53 allele. As such, targeted inhibition of Mdm2 is being considered as a therapeutic anticancer strategy. The N-terminal hydrophobic pocket of Mdm2 binds to p53 and thereby inhibits the transcription of p53 target genes. Additionally, the C-terminus of Mdm2 contains a RING domain with intrinsic ubiquitin E3 ligase activity. By recruiting E2 ubiquitin-conjugating enzyme(s), Mdm2 acts as a molecular scaffold to facilitate p53 ubiquitination and proteasome-dependent degradation. Mdmx (Mdm4), an Mdm2 homolog, also has a RING domain and hetero-oligomerizes with Mdm2 to stimulate its E3 ligase activity. Recent studies have shown that C-terminal residues adjacent to the RING domain of both Mdm2 and Mdmx contribute to Mdm2 E3 ligase activity. However, the molecular mechanisms mediating this process remain unclear, and the biological consequences of inhibiting Mdm2/Mdmx co-operation or blocking Mdm2 ligase function are relatively unexplored. This study presents biochemical and cell biological data that further elucidate the mechanisms by which Mdm2 and Mdmx co-operate to regulate p53 level and activity. We use chemical and genetic approaches to demonstrate that functional inhibition of Mdm2 ubiquitin ligase activity is insufficient for p53 activation. This unexpected result suggests that concomitant treatment with Mdm2/Mdmx antagonists may be needed to achieve therapeutic benefit.

Actin-crosslinking proteins organize actin into highly dynamic and architecturally diverse subcellular scaffolds that orchestrate a variety of mechanical processes, including lamellipodial and filopodial protrusions in motile cells. How signalling pathways control and coordinate the activity of these crosslinkers is poorly defined. IRSp53, a multi-domain protein that can associate with the Rho-GTPases Rac and Cdc42, participates in these processes mainly through its amino-terminal IMD (IRSp53 and MIM domain). The isolated IMD has actin-bundling activity in vitro and is sufficient to induce filopodia in vivo. However, the manner of regulation of this activity in the full-length protein remains largely unknown. Eps8 is involved in actin dynamics through its actin barbed-ends capping activity and its ability to modulate Rac activity. Moreover, Eps8 binds to IRSp53. Here, we describe a novel actin crosslinking activity of Eps8. Additionally, Eps8 activates and synergizes with IRSp53 in mediating actin bundling in vitro, enhancing IRSp53-dependent membrane extensions in vivo. Cdc42 binds to and controls the cellular distribution of the IRSp53-Eps8 complex, supporting the existence of a Cdc42-IRSp53-Eps8 signalling pathway. Consistently, Cdc42-induced filopodia are inhibited following individual removal of either IRSp53 or Eps8. Collectively, these results support a model whereby the synergic bundling activity of the IRSp53-Eps8 complex, regulated by Cdc42, contributes to the generation of actin bundles, thus promoting filopodial protrusions.

There is a body of literature that describes the geometry and the physics of filopodia using either stochastic models or partial differential equations and elasticity and coarse-grained theory. Comparatively, there is a paucity of models focusing on the regulation of the network of proteins that control the formation of different actin structures. Using a combination of in-vivo and in-vitro experiments together with a system of ordinary differential equations, we focused on a small number of well-characterized, interacting molecules involved in actin-dependent filopodia formation: the actin remodeler Eps8, whose capping and bundling activities are a function of its ligands, Abi-1 and IRSp53, respectively; VASP and Capping Protein (CP), which exert antagonistic functions in controlling filament elongation. The model emphasizes the essential role of complexes that contain the membrane deforming protein IRSp53, in the process of filopodia initiation. This model accurately accounted for all observations, including a seemingly paradoxical result whereby genetic removal of Eps8 reduced filopodia in HeLa, but increased them in hippocampal neurons, and generated quantitative predictions, which were experimentally verified. The model further permitted us to explain how filopodia are generated in different cellular contexts, depending on the dynamic interaction established by Eps8, IRSp53 and VASP with actin filaments, thus revealing an unexpected plasticity of the signaling network that governs the multifunctional activities of its components in the formation of filopodia., Author Summary Cells move and interact with the environment by forming migratory structures composed of self organized polymers of actin. These protrusions can be flat and short surfaces, the lamellipodia, or adopt an elongated, finger-like shape called filopodia. In this article, we analyze the ‘computation’ performed by cells when they opt to form filopodia. We focus our attention on some initiators of filopodia that play an essential role due to their interaction with the cell membrane. We analyze the formation of these filopodia initiators in different genotypes, thus providing a way to rationalize the behaviors of different cells in terms of tendency to form filopodia. Our results, based on the combination of experimental and computational approaches, suggest that cells have developed molecular networks that are extremely flexible in their capability to follow the path leading to filopodia formation. In this sense the role of an element of the network, Eps8, is paradigmatic, as this protein can both induce or inhibit the formation of filopodia depending on the cellular context.

Endocytosis and recycling are emerging as essential components of the wiring enabling cells to perceive extracellular signals and resolve them in a temporally and spatially controlled fashion, directly influencing not only the duration and intensity of the signaling output, but also its correct location. One process, which requires the precise resolution of spatial information, is actin-based cell motility. This is achieved by coordinating membrane traffic, cell substrate adhesion, and actin remodeling in order to generate propulsive forces responsible for the formation of a diverse set of polarized migratory protrusions, the first steps of cell locomotion. Here, we will discuss how prototypical endocytic molecules control actin dynamics, frequently by linking the core machinery of actin polymerization to the plasma membrane. We will further discuss how endocytosis and recycling ensure spatial restriction of signaling to actin dynamics, thus enabling cells to migrate in response to different extracellular stimuli and in diverse microenvironments adopting diverse motile strategies, which have important implications in relevant physiological and pathological processes, first and foremost cancer cell invasion and dissemination.

The unusual dual functions of the actin-binding protein EPS8 as an actin capping and actin bundling factor are mapped to distinct structural features of the protein and to distinct physiological activities in vivo., Actin capping and cross-linking proteins regulate the dynamics and architectures of different cellular protrusions. Eps8 is the founding member of a unique family of capping proteins capable of side-binding and bundling actin filaments. However, the structural basis through which Eps8 exerts these functions remains elusive. Here, we combined biochemical, molecular, and genetic approaches with electron microscopy and image analysis to dissect the molecular mechanism responsible for the distinct activities of Eps8. We propose that bundling activity of Eps8 is mainly mediated by a compact four helix bundle, which is contacting three actin subunits along the filament. The capping activity is mainly mediated by a amphipathic helix that binds within the hydrophobic pocket at the barbed ends of actin blocking further addition of actin monomers. Single-point mutagenesis validated these modes of binding, permitting us to dissect Eps8 capping from bundling activity in vitro. We further showed that the capping and bundling activities of Eps8 can be fully dissected in vivo, demonstrating the physiological relevance of the identified Eps8 structural/functional modules. Eps8 controls actin-based motility through its capping activity, while, as a bundler, is essential for proper intestinal morphogenesis of developing Caenorhabditis elegans., Author Summary One of the key components of the cytoskeleton of cells is actin, which allows cells to move. Actin-based motility is involved in many biological processes, such as intestinal development, intracellular trafficking and cell migration. Actin monomers are individual building blocks that can be linked together to form actin filaments. Numerous actin-binding proteins are involved in controlling the higher order architecture and dynamics of these actin filaments within cells. For example, actin capping proteins regulate actin dynamics by controlling the number of growing filament ends, and actin cross-linking or bundling proteins determine how to organize these filaments into higher order structures. The protein Eps8 is capable of capping as well as bundling actin filaments. However, the structural basis of this dual role of Eps8 remains unknown. In this study, we use a combination of techniques to unravel the molecular and structural basis of Eps8 interactions with actin filaments. We show that distinct structural modules of Eps8 are responsible for capping versus bundling activity, and we determine the contributions of these modules in vitro and in vivo. At the functional level, we find that Eps8 regulates actin-based motility and cellular trafficking through its capping activity, whereas Eps8-mediated bundling is essential for intestinal morphogenesis.