Search Results (407)

Small (monomeric) GTP-binding proteins (smgs; Cdc42 and Rac1) play requisite roles in islet beta cell function, including glucose-stimulated insulin secretion. In addition, emerging evidence suggests that sustained (constitutive) activation of smgs (e.g., Rac1) culminates in the genesis of islet beta cell dysfunction under the duress of chronic hyperglycemia. It is noteworthy that functions (i.e., activation–deactivation) of smgs in many cells, including the islet beta cell, have been shown to be under the regulatory control of at least three factors, namely the guanine nucleotide exchange factors (GEFs), the GTPase-activating proteins (GAPs), and the GDP-dissociation inhibitors (GDIs). The overall objective of this review is to highlight our current understanding of the regulatory roles of the RhoGDIβ-Rac1-CARD9 signalome in the pathology of beta cell dysfunction under chronic hyperglycemic stress. For brevity, this review is structured by an overview of smgs and their regulatory proteins/factors in the beta cell, followed by a discussion of potential roles of the RhoGDIβ-Rac1-CARD9 axis in the onset of cellular dysfunction under the duress of metabolic stress. Overall conclusions, potential knowledge gaps, and opportunities for future research in this field of islet biology are highlighted in the last section. Full article

Respiratory syncytial virus (RSV) is a major human respiratory pathogen, particularly affecting infants, the elderly, and immunocompromised individuals. RSV exists in both spherical and filamentous forms, with the filamentous morphology associated with enhanced infectivity and cell-to-cell spread. Here, we demonstrate that RhoA, a small GTPase involved in cytoskeletal regulation, is essential for filamentous RSV morphogenesis through its role in organizing lipid raft microdomains. Rhosin, a selective RhoA inhibitor developed through structure-guided screening, disrupts GEF–RhoA interactions to block RhoA activation. The pharmacological inhibition of RhoA with Rhosin significantly reduced filamentous virion formation, disrupted RSV fusion (F) protein colocalization with lipid rafts, and diminished cell-to-cell fusion, without affecting overall viral replication. Scanning electron microscopy revealed that Rhosin-treated infected HEp-2 cells exhibited fewer and shorter filamentous projections compared to the extensive filament formation seen in untreated cells. β-galactosidase-based fusion assays confirmed that reduced filamentation corresponded with decreased cell-to-cell fusion. The biophysical separation of RSV spherical and filamentous particles by sucrose gradient velocity sedimentation, coupled with fluorescence and transmission electron microscopy, showed that Rhosin treatment shifted virion morphology toward spherical forms. This suggests that RhoA activity is critical for filamentous virion assembly, which may enhance viral spread. Immunofluorescence microscopy using lipid raft-selective dyes (DiIC₁₆) and fusion protein-specific antibodies revealed the strong co-localization of RSV proteins with lipid rafts. Importantly, the pharmacological inhibition of RhoA with Rhosin disrupted F protein partitioning into raft domains, underscoring the requirement for intact lipid rafts in assembly. These findings highlight a novel role for host RhoA signaling in regulating viral assembly through raft microdomain organization, offering a potential target for host-directed antiviral intervention aimed at altering RSV structural phenotypes and limiting pathogenesis. Full article

Axon guidance, a fundamental process in neural circuit formation, is intricately regulated by Fibroblast Growth Factors (FGFs) and their receptors (FGFRs) through dynamic cytoskeletal remodeling. FGF signaling, mediated by heparan sulfate proteoglycans or Klotho co-factors, activates key downstream pathways: PI3K-Akt, JAK-STAT, PLCγ, and RAS-MAPK. These pathways orchestrate actin filament dynamics, microtubule stability, and the organization of intermediate filaments. These pathways converge on Rho GTPases, cofilin, profilin, and tau to balance the cytoskeletal assembly−disassembly cycles, enabling growth cone navigation. Unresolved questions, such as the mechanisms underlying FGF-mediated growth cone steering, highlight critical future research directions. This review integrates structural, molecular, and functional insights into how FGF-FGFR interactions regulate axon pathfinding, emphasizing the crosstalk between signaling cascades and cytoskeletal plasticity. Elucidating these mechanisms not only advances our understanding of neural development but also opens therapeutic avenues for neuro-developmental disorders, nerve injury, and neurodegenerative diseases by targeting FGF-driven cytoskeletal dynamics. Full article

The neurotrophic system includes neurotrophins, such as brain-derived neurotrophic factor (BDNF) and its precursor proBDNF, which play conflicting roles in neuronal survival and apoptosis, with their balance having a significant impact on neurodegenerative outcomes. While BDNF is widely acknowledged as a potent neurotrophin that promotes neuronal survival and differentiation, its precursor, proBDNF, has the opposite effect, promoting apoptosis and neuronal death. This review highlights the new and unique aspects of BDNF/proBDNF interaction in the modulation of neuronal apoptotic pathways in neurodegenerative disorders. It systematically discusses the cross-talk in apoptotic signaling at the molecular level, whereby BDNF activates survival pathways such as PI3K/Akt and MAPK/ERK, whereas proBDNF activates p75NTR and sortilin to induce neuronal apoptosis via JNK, RhoA, NFkB, and Rac-GTPase pathways such as caspase activation and mitochondrial injury. Moreover, this review emphasizes the factors that affect the balance between proBDNF and BDNF levels within the context of neurodegeneration, including proteolytic processing, the expression of TrkB and p75NTR receptors, and extrinsic gene transcription regulators. Cellular injury, stress, or signaling pathway alterations can disrupt the balance of BDNF/proBDNF, which may be involved in apoptotic-related neurodegenerative diseases like Alzheimer’s, Parkinson’s, and Huntington’s diseases. This review provides a comprehensive framework for targeting neurotrophin signaling in the development of innovative therapies for neuronal survival and managing apoptotic-related neurodegenerative disorders, addressing the mechanistic complexity and clinical feasibility of BDNF/proBDNF interaction. Full article

The Arhgap29 gene encodes Rho-GTPase-activating protein 29 (Arhgap29), which plays a crucial role in embryonic tissue development. Mutations in the Arhgap29 gene are significantly associated with non-syndromic cleft lip and palate (NSCL/P). Our study demonstrated that the deletion of Arhgap29 leads to syndromic cleft lip and palate (SCL/P) characteristics in mice, where, in addition to cleft palate, the mice exhibit craniofacial and systemic skeletal abnormalities. However, the mechanisms underlying these skeletal abnormalities remain unclear. Through micro-CT imaging, histological analysis, and transcriptomic methods, we discovered that the knockout of Arhgap29 delays the fusion of Meckel’s cartilage, widens cranial sutures, reduces bone quality, and alters the expression of osteoblasts and osteoclasts in the mandible. Digit defects, including ectrodactyly and impaired endochondral ossification, were also observed. Immunohistochemical analysis demonstrated the expression of Arhgap29 in both osteoblasts and osteoclasts, indicating its dual role in maintaining matrix homeostasis and regulating bone resorption equilibrium. Transcriptomic analysis revealed disrupted calcium and MAPK signaling pathways, while in vitro studies demonstrated impaired osteogenesis in Arhgap29-deficient calvarial cells, mirroring the in vivo defects. Furthermore, spatial transcriptomics linked the loss of Arhgap29 to defective bone differentiation and protein synthesis. Our findings underscore the critical role of Arhgap29 in the development of the mandible and digits, suggesting its potential as a pathogenic gene associated with syndromic cleft lip and palate (SCL/P). Full article

The aim of this work is to highlight the beneficial effects of bioactive peptides present in fermented whey (FW) and carotenoids from pumpkin (P) against the pro-oxidant effects of aflatoxin B1 and ochratoxin A at the neuronal level. For this purpose, SH-SY5Y human neuroblastoma differentiated cells were exposed to (A) mycotoxins, (B) the digesta of mycotoxin-contaminated bread formulated with P, or (C) bread enriched with FW + P. A proteomic approach using HPLC-MS/MS-QTOF was then employed to characterize the metabolic pathways affected by the presence of these components, as well as their ability to modulate the toxic effects exacerbated by mycotoxins. Gene ontology functional analysis revealed proteins primarily associated with nucleosome structure, such as the H3-H4 tetramer, H2A-H2B dimer, and HIRA, which were overexpressed in the presence of mycotoxins and, interestingly, downregulated with the addition of the functional ingredients. Additionally, important metabolic pathways associated with the RHO GTPase family, estrogen-dependent gene expression, and androgen receptor transcription stimulated by PKN1 activation were discovered. Network interaction analysis highlighted the modulation of cytoskeletal dynamics, cell migration, and stress responses. These findings provide novel insights into the neuroprotective potential of functional food components, supporting their use in mitigating mycotoxin-induced neuronal damage and opening new avenues for dietary-based neuroprotection strategies. Full article

Chronic liver diseases are marked by persistent inflammation and can evolve into liver fibrosis, cirrhosis, and hepatocellular carcinoma. In an affected liver, hepatic stellate cells (HSCs) transition from a quiescent to an activated state and adopt a myofibroblast-like cell phenotype. While these activated cells play a role in supporting liver regeneration, they can also have detrimental effects on liver function as the disease progresses to fibrosis and cirrhosis. These findings highlight the dynamic switching between different signaling pathways involving Ras, Rho GTPases, and Notch signaling. Notably, two specific members of the Ras and Rho GTPases, Eras and Rnd3, are predominantly expressed in quiescent HSCs, while Mras and Rhoc are more abundant in their activated forms. In addition, this study highlights the critical role of cytosolic Notch1 in quiescent HSCs and Rock in activated HSCs. We hypothesize that distinct yet interdependent intracellular signaling networks regulate HSC fate decisions in two key ways: by maintaining HSC quiescence and homeostasis and by facilitating HSC activation, thereby influencing processes such as proliferation, transdifferentiation, and mesenchymal transition. The proposed signaling model, combined with specific methodological tools for maintaining HSCs in a quiescent state, will deepen our understanding of the mechanisms underlying chronic liver disease and may also pave the way for innovative therapies. These therapies could include small molecule drugs targeting Ras- and Rho-dependent pathways. Full article

The p21-activated kinases (PAKs) are a group of evolutionarily conserved serine/threonine protein kinases and serve as a downstream target of the small GTPases Rac and Cdc42, both of which belong to the Rho family. PAKs play pivotal roles in various physiological processes, including cytoskeletal rearrangement and cellular signal transduction. Group II PAKs (PAK4-6) are particularly closely linked to human tumors, such as breast and pancreatic cancers, while Group I PAKs (PAK1-3) are indispensable for normal physiological functions such as cardiovascular development and neurogenesis. In recent years, the association of PAKs with diseases like cancer and the rise of small-molecule inhibitors targeting PAKs have attracted significant attention. This article focuses on the analysis of PAKs’ role in tumor progression and immune infiltration, as well as the current small-molecule inhibitors of PAKs and their mechanisms. Full article

Male infertility is often linked to sperm quality issues; however, the mechanisms behind these alterations remain unclear in certain contexts. This study investigates the impact of anomalous Rho GTPase activation—a process triggered by bacterial toxins—on human sperm structure and function. Human spermatozoa were exposed in vitro to a Rho GTPase activator derived from Escherichia coli under both capacitating and non-capacitating conditions. The results showed increased RhoA GTPase activity in non-capacitating conditions, without affecting viability or mitochondrial membrane potential. However, progressive motility decreased across both conditions, while non-progressive motility and acrosome reaction rates increased. Additionally, intracellular calcium levels rose exclusively in non-capacitating conditions. Structural analysis revealed an increase in abnormal sperm morphology, particularly vacuoles in the sperm head. These findings highlight that anomalous Rho GTPase activation disrupts essential processes like motility and capacitation, which are crucial for successful fertilization. This study provides novel insights into how bacterial infections may induce sperm damage, proposing that Rho GTPase activity could serve as a biomarker for evaluating sperm quality in cases of infertility linked to urogenital infections. Understanding these mechanisms may improve diagnostic and therapeutic approaches for male infertility associated with bacterial pathogens. Human spermatozoa were exposed in vitro to a Rho GTPase activator derived from Escherichia coli under both capacitating and non-capacitating conditions. Full article

Clostridioides difficile Infection (CDI) continues to be a major cause of antibiotic-associated diarrhea and pseudomembranous colitis, fueled in large measure by virulence factors TcdA and TcdB. These giant glucosyltransferase toxins interfere with host cytoskeletal integrity and inflammatory signaling by inhibiting Rho GTPase; however, the detailed structural dynamics, receptor selectivity, and subcellular trafficking mechanisms remain in part unspecified. This review integrates recent insights from cryo-electron microscopy (cryo-EM) and X-ray crystallography to describe the quaternary architecture of TcdA/B, emphasizing conformational changes key to pore formation and endosomal escape. We also examine the genomic heterogeneity of hypervirulent C. difficile strains (e.g., ribotype 027), correlating toxin gene polymorphisms (e.g., tcdC mutations) with increased toxin production and virulence. Mechanistic explanations of toxin-driven inflammasome activation and epithelial barrier dysfunction are situated within host immune evasion mechanisms, including microbiota-derived bile acid regulation of toxin stability. Subsequent innovative therapeutic strategies, encompassing the utilization of engineered neutralizing antibodies that specifically target the autoprocessing domain alongside structure-guided small-molecule inhibitors, are subjected to a rigorous evaluation. By integrating structural biology, systems-level omics, and clinical epidemiology, this review establishes a comprehensive framework for understanding C. difficile toxin pathogenesis and guiding next-generation precision antimicrobials. Full article

Aberrant Rho-associated kinase function could be associated with increased bone fragility. Since cigarette smoke (CS) exposure promotes the increase in bone fragility due to changes in bone tissue components, this study aimed to investigate how CS exposure could modulate the Rho kinase-associated bone structural changes. Mice were assigned to four groups: control; smoke; control with Rho kinase inhibitor administration; and smoke with a Rho kinase inhibitor. Bone samples were obtained to assess bone histomorphometry analysis, type I collagen composition, and MEPE expression in trabeculae. We observed that CS exposure induced decreased trabecular and osteoid thickness. A concomitant increase in the osteoclastic and erosion surfaces and a decrease in the mineralization surface were observed. Additionally, CS exposure decreased the type I collagen and MEPE expression. Rho kinase inhibitor administration recovered the bone mineralization and the collagen type I deposition. Conclusions: CS exposure increases Rho kinase activity in bone cells, leading to structural changes. The administration of a Rho GTPases inhibitor partially reverses these effects, likely due to the recovery in osteoblast activity. Full article

Mitochondria-ER contact sites (MERCS) are vital for mitochondrial dynamics, lipid exchange, Ca²⁺ homeostasis, and energy metabolism. We examined whether mitochondrial metabolism changes during the cell cycle depend on MERCS dynamics and are regulated by the outer mitochondrial protein mitochondrial rho GTPase 1 (MIRO1). Wound healing was assessed in mice with fibroblast-specific deletion of MIRO1. Wild-type and MIRO1^-/- fibroblasts and vascular smooth muscle cells were evaluated for proliferation, cell cycle progression, number of MERCS, distance, and protein composition throughout the cell cycle. Restoration of MIRO1 mutants was used to test the role of MIRO1 domains; Ca²⁺ transients and mitochondrial metabolism were evaluated using biochemical, immunodetection, and fluorescence techniques. MERCS increased in number during G1/S compared with during G0, which was accompanied by a notable rise in protein–protein interactions involving VDAC1 and IP3R as well as GRP75 and MIRO1 by proximity-ligation assays. Split-GFP ER/mitochondrial contacts of 40 nm also increased. Mitochondrial Ca²⁺ concentration ([Ca²⁺]), membrane potential, and ATP levels correlated with the formation of MERCS during the cell cycle. MIRO1 deficiency blocked G1/S progression and the cell-cycle-dependent formation of MERCS and altered ER Ca²⁺ release and mitochondrial Ca²⁺ uptake. MIRO1 mutants lacking the Ca²⁺-sensitive EF hands or the transmembrane domain did not rescue cell proliferation or the formation of MERCS. MIRO1 controls an increase in the number of MERCS during cell cycle progression and increases mitochondrial [Ca²⁺], driving metabolic activity and proliferation through its EF hands. Full article

Metastatic prostate cancer occurs when the tumor spreads from the prostate gland to other parts of the body. Previous studies have shown that Gα_i2, a subunit of the heterotrimeric G protein complex, plays a critical role in inducing cell migration and invasion in prostate cancer cells in response to diverse stimuli. Rac1 is a small rho-GTPase, which is activated by the phosphoinositide 3-kinase (PI3K)/AKT pathway and plays an essential role during cell migration. Previous studies have shown that the knockdown of Gα_i2 attenuates cell migration without causing any reduction in basal Rac1 activity in both PC3 and DU145 cells, and has only marginal effects on the epidermal growth facotor (EGF)-induced increase in Rac1 activity. Therefore, Gα_i2 may be involved in the regulation of cell motility and invasion independently or downstream of Rac1 activation. In this study, we investigated the possible mechanism of Gα_i2 at the level of the Rac1-dependent activation of Wiskott-Aldrich Syndrome Protein)-family verprolin homologous protein2 (Wave2) and actin related protein 2/3 (Arp 2/3) proteins, downstream effectors of activated Rac1. PC3 cells with a stable overexpression of constitutively active Rac1 were transfected with control siRNA or Gα_i2 siRNA to knockdown endogenous Gα_i2 expression. Western blot analysis showed that the Rac1-dependent activation of Wave2 was impaired in the absence of Gα_i2. The overexpression of constitutively active Gα_i2 (Gα_i2-Q205L) in PC3 cells significantly increased cell migration compared to cells transfected with control plasmids. In the parallel experiments, a specific Gα_i2 inhibitor blocked G_iα2-Q205L-induced cell migration in PC3 cells. Furthermore, the Rac1 inhibitor did not block increased cell migration in PC3 cells overexpressing constitutively active Gα_i2. We conclude that activated Gα_i2 plays a crucial role in cell migration in prostate cancer cells independent of Rac1 activation. Full article

Several molecular pathways are likely involved in the regulation of cancer stem cells (CSCs) via Ras-associated C3 botulinum toxin substrate 2, RAC2, and pituitary tumor-transforming gene 1 product, PTTG1, given their roles in cellular signaling, survival, proliferation, and metastasis. RAC2 is a member of the Rho GTPase family and plays a crucial role in actin cytoskeleton dynamics, reactive oxygen species production, and cell migration, contributing to epithelial–mesenchymal transition (EMT), immune evasion, and therapy resistance. PTTG1, also known as human securin, regulates key processes such as cell cycle progression, apoptosis suppression, and EMT, promoting metastasis and enhancing cancer cell survival. This article aims to describe the molecular pathways involved in the proliferation, invasiveness, and drug response of cancer cells through RAC2 and PTTG1, aiming to clarify their respective roles in neoplastic process dependencies. Both proteins are involved in critical signaling pathways, including PI3K/AKT, TGF-β, and NF-κB, which facilitate tumor progression by modulating CSC properties, angiogenesis, and immune response. This review highlights the molecular mechanisms by which RAC2 and PTTG1 influence tumorigenesis and describes their potential and efficacy as prognostic biomarkers and therapeutic targets in managing various neoplasms. Full article

Acute myeloid leukemia (AML) is an aggressive bone marrow malignancy that can be cured only by intensive chemotherapy possibly combined with allogeneic stem cell transplantation. We compared the pretreatment proteomic profiles of AML cells derived from 50 patients at the time of first diagnosis with normal CD34⁺ bone marrow cells. A comparison based on all AML and CD34⁺ normal cell populations identified 121 differentially abundant proteins that showed at least 2-fold differences, and these proteins included several markers of neutrophil differentiation (e.g., TLR2, the integrins ITGM and ITGX, and downstream mediators including RHO GTPase, S100A8, S100A9, S100A22). However, the expression of these 121 proteins varied between patients, and a subset of 28 patients was characterized by increased long-term AML-free survival, signs of myeloid AML cell differentiation, and favorable genetic abnormalities. These two main patient subsets (28 with differentiation versus 22 with fewer signs of differentiation) also differed with regard to the phosphorylation of 16 differentially abundant proteins. Furthermore, we also classified our patients based on their expression of 16 proteins involved in the regulation of iron metabolism/ferroptosis and showing differential expression when comparing AML cells and normal CD34+ cells. Among the 22 patients with less favorable prognosis, we could then identify a genetically heterogeneous subset characterized by adverse prognosis (i.e., death from primary resistance/relapse) and an iron metabolism/ferroptosis protein profile showing similarities with normal CD34⁺ cells. We conclude that proteomic profiles differ between AML and normal CD34⁺ cells; especially, proteomic differences reflecting differentiation and regulation of iron metabolism/ferroptosis are associated with risk of relapse after intensive conventional therapy. Full article