Role of WAVE3 as an of actin binding protein in the pathology of triple negative breast cancer.

Breast cancer, a prevalent global health concern, has sparked extensive research efforts, particularly focusing on triple negative breast cancer (TNBC), a subtype lacking estrogen receptor (ER), progesterone receptor, and epidermal growth factor receptor. TNBC's aggressive nature and resistance to hormone-based therapies heightens the risk of tumor progression and recurrence. Actin-binding proteins, specifically WAVE3 from the Wiskott-Aldrich syndrome protein (WASP) family, have emerged as major drivers in understanding TNBC biology. This review delves into the intricate molecular makeup of TNBC, shedding light on actin's fundamental role in cellular processes. Actin, a structural element in the cytoskeleton, regulates various cellular pathways essential for homeostasis. Its dynamic nature enables functions such as cell migration, motility, intracellular transport, cell division, and signal transduction. Actin-binding proteins, including WAVE3, play pivotal roles in these processes. WAVE3, a member of the WASP family, remains the focus of this review due to its potential involvement in TNBC progression. While actin-binding proteins are studied for their roles in healthy cellular cycles, their significance in TNBC remains underexplored. This review aims to discuss WAVE3's impact on TNBC, exploring its molecular makeup, functions, and significance in tumor progression. The intricate structure of WAVE3, featuring elements like the verprolin-cofilin-acidic domain and regulatory elements, plays a crucial role in regulating actin dynamics. Dysregulation of WAVE3 in TNBC has been linked to enhanced cell migration, invasion, extracellular matrix remodeling, epithelial-mesenchymal transition, tumor proliferation, and therapeutic resistance. Understanding the role of actin-binding proteins in cancer biology has potential clinical implications, making them potential prognostic biomarkers and promising therapeutic targets. The review emphasizes the need for further research into actin-binding proteins' clinical applications, diagnostic value, and therapeutic interventions. In conclusion, this comprehensive review explores the complex interplay between actin and actin-binding proteins, with special emphasis on WAVE3, in the context of TNBC. By unraveling the molecular intricacies, structural characteristics, and functional significance, the review paves the way for future research directions, clinical applications, and potential therapeutic strategies in the challenging landscape of TNBC.

Role of actin-binding proteins in cataract formation.

INTRODUCTION: Actin has been implicated in lens opacification; however, the specific actin-related pathways involved in cataracts remain unelucidated. In this study, actin-related proteome changes and signaling pathways involved in the development of cataracts were evaluated. METHODS: The anterior capsule and phacoemulsification (phaco) cassette contents were collected during cataract surgery from 11 patients with diabetic cataract (DC), 12 patients with age-related cataract (ARC), and seven patients with post-vitrectomy cataract (PVC). Untargeted, global identification and quantification of proteins was performed through liquid chromatography-mass spectrometry with the data-independent acquisition (DIA). RESULTS: In phaco cassette samples, proteins with significantly lower expression in ARC than in DC and PVC were involved in various pathways, including actin binding, actin cytoskeleton reorganization, actin filament capping, cortical actin cytoskeleton organization, and small GTPase-mediated signal transduction pathways. In anterior capsules, proteins with significantly lower expression in ARC than in DC and PVC were involved in actin binding and actin cytoskeleton reorganization pathways. CONCLUSION: Actin cytoskeleton and actin-binding proteins are involved in lens fiber elongation and differentiation. Rho GTPases contribute to actin cytoskeletal reorganization, and their inactivation is linked to abnormal lens fiber migration. These findings link actin binding to lens fiber integrity, lens opacification, and cataracts.

Bioinformatics Analysis of Actin Interactome: Characterization of the Nuclear and Cytoplasmic Actin-Binding Proteins.

Actin is present in the cytoplasm and nucleus of every eukaryotic cell. In the cytoplasm, framework and motor functions of actin are associated with its ability to polymerize to form F-actin. In the nucleus, globular actin plays a significant functional role. For a globular protein, actin has a uniquely large number of proteins with which it interacts. Bioinformatics analysis of the actin interactome showed that only a part of actin-binding proteins are both cytoplasmic and nuclear. There are proteins that interact only with cytoplasmic, or only with nuclear actin. The first pool includes proteins associated with the formation, regulation, and functioning of the actin cytoskeleton predominate, while nuclear actin-binding proteins are involved in the majority of key nuclear processes, from regulation of transcription to DNA damage response. Bioinformatics analysis of the structure of actin-binding proteins showed that these are mainly intrinsically disordered proteins, many of which are part of membrane-less organelles. Interestingly, although the number of intrinsically disordered actin-binding proteins in the nucleus is greater than in the cytoplasm, the drivers for the formation of the membrane-less organelles in the cytoplasm are significantly (four times) greater than in the nucleus.

The Critical Role of the Shroom Family Proteins in Morphogenesis, Organogenesis and Disease.

The Shroom (Shrm) family of actin-binding proteins has a unique and highly conserved Apx/Shrm Domain 2 (ASD2) motif. Shroom protein directs the subcellular localization of Rho-associated kinase (ROCK), which remodels the actomyosin cytoskeleton and changes cellular morphology via its ability to phosphorylate and activate non-muscle myosin II. Therefore, the Shrm-ROCK complex is critical for the cellular shape and the development of many tissues, including the neural tube, eye, intestines, heart, and vasculature system. Importantly, the structure and expression of Shrm proteins are also associated with neural tube defects, chronic kidney disease, metastasis of carcinoma, and X-link mental retardation. Therefore, a better understanding of Shrm-mediated signaling transduction pathways is essential for the development of new therapeutic strategies to minimize damage resulting in abnormal Shrm proteins. This paper provides a comprehensive overview of the various Shrm proteins and their roles in morphogenesis and disease.

Structural and functional mechanisms of actin isoforms.

Actin is a highly conserved and fundamental protein in eukaryotes and participates in a broad spectrum of cellular functions. Cells maintain a conserved ratio of actin isoforms, with muscle and non-muscle actins representing the main actin isoforms in muscle and non-muscle cells, respectively. Actin isoforms have specific and redundant functional roles and display different biochemistries, cellular localization, and interactions with myosins and actin-binding proteins. Understanding the specific roles of actin isoforms from the structural and functional perspective is crucial for elucidating the intricacies of cytoskeletal dynamics and regulation and their implications in health and disease. Here, we review how the structure contributes to the functional mechanisms of actin isoforms with a special emphasis on the questions of how post-translational modifications and disease-linked mutations affect actin isoforms biochemistry, function, and interaction with actin-binding proteins and myosin motors.

Structural insights into the molecular mechanism of phytoplasma immunodominant membrane protein.

Immunodominant membrane protein (IMP) is a prevalent membrane protein in phytoplasma and has been confirmed to be an F-actin-binding protein. However, the intricate molecular mechanisms that govern the function of IMP require further elucidation. In this study, the X-ray crystallographic structure of IMP was determined and insights into its interaction with plant actin are provided. A comparative analysis with other proteins demonstrates that IMP shares structural homology with talin rod domain-containing protein 1 (TLNRD1), which also functions as an F-actin-binding protein. Subsequent molecular-docking studies of IMP and F-actin reveal that they possess complementary surfaces, suggesting a stable interaction. The low potential energy and high confidence score of the IMP-F-actin binding model indicate stable binding. Additionally, by employing immunoprecipitation and mass spectrometry, it was discovered that IMP serves as an interaction partner for the phytoplasmal effector causing phyllody 1 (PHYL1). It was then shown that both IMP and PHYL1 are highly expressed in the S2 stage of peanut witches' broom phytoplasma-infected Catharanthus roseus. The association between IMP and PHYL1 is substantiated through in vivo immunoprecipitation, an in vitro cross-linking assay and molecular-docking analysis. Collectively, these findings expand the current understanding of IMP interactions and enhance the comprehension of the interaction of IMP with plant F-actin. They also unveil a novel interaction pathway that may influence phytoplasma pathogenicity and host plant responses related to PHYL1. This discovery could pave the way for the development of new strategies to overcome phytoplasma-related plant diseases.

Arp2/3 complex- and formin-mediated actin cytoskeleton networks facilitate actin binding protein sorting in fission yeast.

While it is well-established that F-actin networks with specific organizations and dynamics are tightly regulated by distinct sets of associated actin-binding proteins (ABPs), how ABPs self-sort to particular F-actin networks remains largely unclear. We report that actin assembly factors Arp2/3 complex and formin Cdc12 tune the association of ABPs fimbrin Fim1 and tropomyosin Cdc8 to different F-actin networks in fission yeast. Genetic and pharmacological disruption of F-actin networks revealed that Fim1 is preferentially directed to Arp2/3-complex mediated actin patches, whereas Cdc8 is preferentially targeted to formin Cdc12-mediated filaments in the contractile ring. To investigate the role of Arp2/3 complex- and formin Cdc12-mediated actin assembly, we used four-color TIRF microscopy to observe the in vitro reconstitution of ABP sorting with purified proteins. Fim1 or Cdc8 alone bind similarly well to filaments assembled by either assembly factor. However, in 'competition' reactions containing both actin assembly factors and both ABPs, ∼2.0-fold more Fim1 and ∼3.5-fold more Cdc8 accumulates on Arp2/3 complex branch points and formin Cdc12-assembled actin filaments, respectively. These findings indicate that F-actin assembly factors Arp2/3 complex and formin Cdc12 help facilitate the recruitment of specific ABPs, thereby tuning ABP sorting and subsequently establishing the identity of F-actin networks in fission yeast.

Emerging roles of deubiquitinating enzymes in actin cytoskeleton and tumor metastasis.

BACKGROUND: Metastasis accounts for the majority of cancer-related deaths. Actin dynamics and actin-based cell migration and invasion are important factors in cancer metastasis. Metastasis is characterized by actin polymerization and depolymerization, which are precisely regulated by molecular changes involving a plethora of actin regulators, including actin-binding proteins (ABPs) and signalling pathways, that enable cancer cell dissemination from the primary tumour. Research on deubiquitinating enzymes (DUBs) has revealed their vital roles in actin dynamics and actin-based migration and invasion during cancer metastasis. CONCLUSION: Here, we review how DUBs drive tumour metastasis by participating in actin rearrangement and actin-based migration and invasion. We summarize the well-characterized and essential actin cytoskeleton signalling molecules related to DUBs, including Rho GTPases, Src kinases, and ABPs such as cofilin and cortactin. Other DUBs that modulate actin-based migration signalling pathways are also discussed. Finally, we discuss and address therapeutic opportunities and ongoing challenges related to DUBs with respect to actin dynamics.

Changes of Ex Vivo Cervical Epithelial Cells Due to Electroporation with JMY.

The ionic environment within the nucleoplasm might diverge from the conditions found in the cytoplasm, potentially playing a role in the cellular stress response. As a result, it is conceivable that interactions of nuclear actin and actin-binding proteins (ABPs) with apoptosis factors may differ in the nucleoplasm and cytoplasm. The primary intracellular stress response is Ca2+ influx. The junctional mediating and regulating Y protein (JMY) is an actin-binding protein and has the capability to interact with the apoptosis factor p53 in a Ca2+-dependent manner, forming complexes that play a regulatory role in cytoskeletal remodelling and motility. JMY's presence is observed in both the cytoplasm and nucleoplasm. Here, we show that ex vivo ectocervical squamous cells subjected to electroporation with JMY protein exhibited varying morphological alterations. Specifically, the highly differentiated superficial and intermediate cells displayed reduced nuclear size. In inflamed samples, nuclear enlargement and simultaneous cytoplasmic reduction were observable and showed signs of apoptotic processes. In contrast, the less differentiated parabasal and metaplastic cells showed increased cytoplasmic activity and the formation of membrane protrusions. Surprisingly, in severe inflammation, vaginosis or ASC-US (Atypical Squamous Cells of Undetermined Significance), JMY appears to influence only the nuclear and perinuclear irregularities of differentiated cells, and cytoplasmic abnormalities still existed after the electroporation. Our observations can provide an appropriate basis for the exploration of the relationship between cytopathologically relevant morphological changes of epithelial cells and the function of ABPs. This is particularly important since ABPs are considered potential diagnostic and therapeutic biomarkers for both cancers and chronic inflammation.

Role of Actin-Binding Proteins in Skeletal Myogenesis.

Maintenance of skeletal muscle quantity and quality is essential to ensure various vital functions of the body. Muscle homeostasis is regulated by multiple cytoskeletal proteins and myogenic transcriptional programs responding to endogenous and exogenous signals influencing cell structure and function. Since actin is an essential component in cytoskeleton dynamics, actin-binding proteins (ABPs) have been recognized as crucial players in skeletal muscle health and diseases. Hence, dysregulation of ABPs leads to muscle atrophy characterized by loss of mass, strength, quality, and capacity for regeneration. This comprehensive review summarizes the recent studies that have unveiled the role of ABPs in actin cytoskeletal dynamics, with a particular focus on skeletal myogenesis and diseases. This provides insight into the molecular mechanisms that regulate skeletal myogenesis via ABPs as well as research avenues to identify potential therapeutic targets. Moreover, this review explores the implications of non-coding RNAs (ncRNAs) targeting ABPs in skeletal myogenesis and disorders based on recent achievements in ncRNA research. The studies presented here will enhance our understanding of the functional significance of ABPs and mechanotransduction-derived myogenic regulatory mechanisms. Furthermore, revealing how ncRNAs regulate ABPs will allow diverse therapeutic approaches for skeletal muscle disorders to be developed.

Coronin-1 promotes directional cell rearrangement in Drosophila wing epithelium.

Directional cell rearrangement is a critical process underlying correct tissue deformation during morphogenesis. Although the involvement of F-actin regulation in cell rearrangement has been established, the role and regulation of actin binding proteins (ABPs) in this process are not well understood. In this study, we investigated the function of Coronin-1, a WD-repeat actin-binding protein, in controlling directional cell rearrangement in the Drosophila pupal wing. Transgenic flies expressing Coronin-1-EGFP were generated using CRISPR-Cas9. We observed that Coronin-1 localizes at the reconnecting junction during cell rearrangement, which is dependent on actin interacting protein 1 (AIP1) and cofilin, actin disassemblers and known regulators of wing cell rearrangement. Loss of Coronin-1 function reduces cell rearrangement directionality and hexagonal cell fraction. These results suggest that Coronin-1 promotes directional cell rearrangement via its interaction with AIP1 and cofilin, highlighting the role of ABPs in the complex process of morphogenesis.Key words: morphogenesis, cell rearrangement, actin binding proteins (ABPs).

Alternative splicing of a single exon causes a major impact on the affinity of Caenorhabditis elegans tropomyosin isoforms for actin filaments.

Tropomyosin is generally known as an actin-binding protein that regulates actomyosin interaction and actin filament stability. In metazoans, multiple tropomyosin isoforms are expressed, and some of them are involved in generating subpopulations of actin cytoskeleton in an isoform-specific manner. However, functions of many tropomyosin isoforms remain unknown. Here, we report identification of a novel alternative exon in the Caenorhabditis elegans tropomyosin gene and characterization of the effects of alternative splicing on the properties of tropomyosin isoforms. Previous studies have reported six tropomyosin isoforms encoded by the C. elegans lev-11 tropomyosin gene. We identified a seventh isoform, LEV-11U, that contained a novel alternative exon, exon 7c (E7c). LEV-11U is a low-molecular-weight tropomyosin isoform that differs from LEV-11T only at the exon 7-encoded region. In silico analyses indicated that the E7c-encoded peptide sequence was unfavorable for coiled-coil formation and distinct from other tropomyosin isoforms in the pattern of electrostatic surface potentials. In vitro, LEV-11U bound poorly to actin filaments, whereas LEV-11T bound to actin filaments in a saturable manner. When these isoforms were transgenically expressed in the C. elegans striated muscle, LEV-11U was present in the diffuse cytoplasm with tendency to form aggregates, whereas LEV-11T co-localized with sarcomeric actin filaments. Worms with a mutation in E7c showed reduced motility and brood size, suggesting that this exon is important for the optimal health. These results indicate that alternative splicing of a single exon can produce biochemically diverged tropomyosin isoforms and suggest that a tropomyosin isoform with poor actin affinity has a novel biological function.

Cytochalasans and Their Impact on Actin Filament Remodeling.

The eukaryotic actin cytoskeleton comprises the protein itself in its monomeric and filamentous forms, G- and F-actin, as well as multiple interaction partners (actin-binding proteins, ABPs). This gives rise to a temporally and spatially controlled, dynamic network, eliciting a plethora of motility-associated processes. To interfere with the complex inter- and intracellular interactions the actin cytoskeleton confers, small molecular inhibitors have been used, foremost of all to study the relevance of actin filaments and their turnover for various cellular processes. The most prominent inhibitors act by, e.g., sequestering monomers or by interfering with the polymerization of new filaments and the elongation of existing filaments. Among these inhibitors used as tool compounds are the cytochalasans, fungal secondary metabolites known for decades and exploited for their F-actin polymerization inhibitory capabilities. In spite of their application as tool compounds for decades, comprehensive data are lacking that explain (i) how the structural deviances of the more than 400 cytochalasans described to date influence their bioactivity mechanistically and (ii) how the intricate network of ABPs reacts (or adapts) to cytochalasan binding. This review thus aims to summarize the information available concerning the structural features of cytochalasans and their influence on the described activities on cell morphology and actin cytoskeleton organization in eukaryotic cells.

A census of actin-associated proteins in humans.

Actin filaments help in maintaining the cell structure and coordinating cellular movements and cargo transport within the cell. Actin participates in the interaction with several proteins and also with itself to form the helical filamentous actin (F-actin). Actin-binding proteins (ABPs) and actin-associated proteins (AAPs) coordinate the actin filament assembly and processing, regulate the flux between globular G-actin and F-actin in the cell, and help maintain the cellular structure and integrity. We have used protein-protein interaction data available through multiple sources (STRING, BioGRID, mentha, and a few others), functional annotation, and classical actin-binding domains to identify actin-binding and actin-associated proteins in the human proteome. Here, we report 2482 AAPs and present an analysis of their structural and sequential domains, functions, evolutionary conservation, cellular localization, abundance, and tissue-specific expression patterns. This analysis provides a base for the characterization of proteins involved in actin dynamics and turnover in the cell.

The Role of non-muscle actin paralogs in cell cycle progression and proliferation.

Uncontrolled cell proliferation leads to several pathologies, including cancer. Thus, this process must be tightly regulated. The cell cycle accounts for cell proliferation, and its progression is coordinated with changes in cell shape, for which cytoskeleton reorganization is responsible. Rearrangement of the cytoskeleton allows for its participation in the precise division of genetic material and cytokinesis. One of the main cytoskeletal components is filamentous actin-based structures. Mammalian cells have at least six actin paralogs, four of which are muscle-specific, while two, named ß- and γ-actin, are abundantly present in all types of cells. This review summarizes the findings that establish the role of non-muscle actin paralogs in regulating cell cycle progression and proliferation. We discuss studies showing that the level of a given non-muscle actin paralog in a cell influences the cell's ability to progress through the cell cycle and, thus, proliferation. Moreover, we elaborate on the non-muscle actins' role in regulating gene transcription, interactions of actin paralogs with proteins involved in controlling cell proliferation, and the contribution of non-muscle actins to different structures in a dividing cell. The data cited in this review show that non-muscle actins regulate the cell cycle and proliferation through varying mechanisms. We point to the need for further studies addressing these mechanisms.

Actin: Static and Dynamic Studies.

The actin cytoskeleton is a highly dynamic network in plant cells, which is precisely regulated by numerous actin-binding proteins. Hence, characterizing the biochemical activities of actin-binding proteins is of great importance. Here we describe methods for determining the binding and bundling of microfilaments as well as methods for visualizing microfilaments using fluorescent phalloidin and single-molecule TIRF imaging.

The Role of Liquid-Liquid Phase Separation in Actin Polymerization.

To date, it has been shown that the phenomenon of liquid-liquid phase separation (LLPS) underlies many seemingly completely different cellular processes. This provided a new idea of the spatiotemporal organization of the cell. The new paradigm makes it possible to provide answers to many long-standing, but still unresolved questions facing the researcher. In particular, spatiotemporal regulation of the assembly/disassembly of the cytoskeleton, including the formation of actin filaments, becomes clearer. To date, it has been shown that coacervates of actin-binding proteins that arise during the phase separation of the liquid-liquid type can integrate G-actin and thereby increase its concentration to initiate polymerization. It has also been shown that the activity intensification of actin-binding proteins that control actin polymerization, such as N-WASP and Arp2/3, can be caused by their integration into liquid droplet coacervates formed by signaling proteins on the inner side of the cell membrane.

Chlamydia trachomatis Subverts Alpha-Actinins To Stabilize Its Inclusion.

Chlamydia trachomatis is the leading cause of sexually transmitted bacterial disease and a global health burden. As an obligate intracellular pathogen, Chlamydia has evolved many strategies to manipulate its host and establish its intracellular niche called the inclusion. C. trachomatis reorganizes the host actin cytoskeleton to form scaffolds around the inclusion and reinforce the growing inclusion membrane. To control the kinetics and formation of actin scaffolds, Chlamydia expresses the effector InaC/CT813, which activates the host GTPase RhoA. Here, we have discovered that InaC stabilizes actin scaffolds through the host actin cross-linking proteins α-actinins 1 and 4. We demonstrate that α-actinins are recruited to the inclusion membrane in an InaC-dependent manner and associate with actin scaffolds that envelop the inclusion. Small interfering RNA (siRNA)-mediated knockdown of α-actinins differentially regulate the frequency of actin scaffolds and impair inclusion stability, leaving them susceptible to rupture and to nonionic detergent extraction. Overall, our data identify new host effectors that are subverted by InaC to stabilize actin scaffolds, highlighting the versatility of InaC as a key regulator of the host cytoskeletal network during Chlamydia infection. IMPORTANCE Despite antibiotics, recurrent C. trachomatis infections cause significant damage to the genital tract in men and women. Without a preventative vaccine, it is paramount to understand the virulence mechanisms that Chlamydia employs to cause disease. In this context, manipulation of the host cytoskeleton is a critical component of Chlamydia development. Actin scaffolds reinforce the integrity of Chlamydia's infectious vacuole, which is a critical barrier between Chlamydia and the host environment. Having previously established that InaC co-opts RhoA to promote the formation of actin scaffolds around the inclusion, we now show that Chlamydia hijacks a new class of host effectors, α-actinins, to cross-link these scaffolds and further stabilize the inclusion. We also establish that a core function of the chlamydial effector InaC is the regulation of cytoskeletal stability during Chlamydia infection. Ultimately, this work expands our understanding of how bacterial pathogens subvert the actin cytoskeleton by targeting fundamental host effector proteins.

Cellular force-sensing through actin filaments.

The actin cytoskeleton orchestrates cell mechanics and facilitates the physical integration of cells into tissues, while tissue-scale forces and extracellular rigidity in turn govern cell behaviour. Here, we discuss recent evidence that actin filaments (F-actin), the core building blocks of the actin cytoskeleton, also serve as molecular force sensors. We delineate two classes of proteins, which interpret forces applied to F-actin through enhanced binding interactions: 'mechanically tuned' canonical actin-binding proteins, whose constitutive F-actin affinity is increased by force, and 'mechanically switched' proteins, which bind F-actin only in the presence of force. We speculate mechanically tuned and mechanically switched actin-binding proteins are biophysically suitable for coordinating cytoskeletal force-feedback and mechanical signalling processes, respectively. Finally, we discuss potential mechanisms mediating force-activated actin binding, which likely occurs both through the structural remodelling of F-actin itself and geometric rearrangements of higher-order actin networks. Understanding the interplay of these mechanisms will enable the dissection of force-activated actin binding's specific biological functions.