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In recent years, mouse nerve growth factor (mNGF) has emerged as an important biological regulator to repair peripheral nerve injury, but its systemic application is restricted by low efficiency and large dosage requirement. These limitations prompted us to search for biomaterials that can be locally loaded. Oxidized sodium alginate hydrogel (OSA) exhibits good biocompatibility and physicochemical properties, and can be loaded with drugs to construct a sustained-release system that can act locally on nerve injury. Here, we constructed a sustained-release system of OSA-mouse nerve growth factor (mNGF), and investigated the loading and release of the drug through Fourier transform infrared spectroscopy and drug release curves. In vitro and in vivo experiments showed that OSA-mNGF significantly promoted the biological activities of RSC-96 cells and facilitated the recovery from sciatic nerve crush injury in rats. This observation may be attributed to the additive effect of OSA on promoting Schwann cell biological activities or its synergistic effect of cross-activating phosphoinositide 3-kinase (PI3K) through extracellular signal regulated kinase (ERK) signaling. Although the specific mechanism of OSA action needs to be explored in the future, the current results provide a valuable preliminary research basis for the clinical application of the OSA-mNGF sustained-release system for nerve repair. [ABSTRACT FROM AUTHOR]

Mechanical ventilation (MV), used in patients with acute lung injury (ALI), induces diaphragmatic myofiber atrophy and contractile inactivity, termed ventilator-induced diaphragm dysfunction. Phosphoinositide 3-kinase-γ (PI3K-γ) is crucial in modulating fibrogenesis during the reparative phase of ALI; however, the mechanisms regulating the interactions among MV, myofiber fibrosis, and PI3K-γ remain unclear. We hypothesized that MV with or without bleomycin treatment would increase diaphragm muscle fibrosis through the PI3K-γ pathway. Five days after receiving a single bolus of 0.075 units of bleomycin intratracheally, C57BL/6 mice were exposed to 6 or 10 mL/kg of MV for 8 h after receiving 5 mg/kg of AS605240 intraperitoneally. In wild-type mice, bleomycin exposure followed by MV 10 mL/kg prompted significant increases in disruptions of diaphragmatic myofibrillar organization, transforming growth factor-β1, oxidative loads, Masson's trichrome staining, extracellular collagen levels, positive staining of α-smooth muscle actin, PI3K-γ expression, and myonuclear apoptosis (p < 0.05). Decreased diaphragm contractility and peroxisome proliferator-activated receptor-γ coactivator-1α levels were also observed (p < 0.05). MV-augmented bleomycin-induced diaphragm fibrosis and myonuclear apoptosis were attenuated in PI3K-γ-deficient mice and through AS605240-induced inhibition of PI3K-γ activity (p < 0.05). MV-augmented diaphragm fibrosis after bleomycin-induced ALI is partially mediated by PI3K-γ. Therapy targeting PI3K-γ may ameliorate MV-associated diaphragm fibrosis. [ABSTRACT FROM AUTHOR]

Phosphoinositides are a small family of phospholipids that act as signaling hubs and key regulators of cellular function. Detecting their subcellular distribution is crucial to gain insights into membrane organization and is commonly done by the overexpression of biosensors. However, this leads to cellular perturbations and is challenging in systems that cannot be transfected. Here, we present a toolkit for the reliable, fast, multiplex, and super-resolution detection of phosphoinositides in fixed cells and tissue, based on recombinant biosensors with self-labeling SNAP tags. These are highly specific and reliably visualize the subcellular distributions of phosphoinositides across scales, from 2D or 3D cell culture to Drosophila tissue. Further, these probes enable super-resolution approaches, and using STED microscopy, we reveal the nanoscale organization of PI(3)P on endosomes and PI(4)P on the Golgi. Finally, multiplex staining reveals an unexpected presence of PI(3,5)P2-positive membranes in swollen lysosomes following PIKfyve inhibition. This approach enables the versatile, high-resolution visualization of multiple phosphoinositide species in an unprecedented manner. [ABSTRACT FROM AUTHOR]

Nutrient absorption is essential for animal survival and development. Our previous study on zebrafish reported that nutrient absorption in lysosome-rich enterocytes (LREs) is promoted by the voltage-sensing phosphatase (VSP), which regulates phosphoinositide (PIP) homeostasis via electrical signaling in biological membranes. However, it remains unknown whether this VSP function is shared by different absorptive tissues in other species. Here, we focused on the function of VSP in a viviparous teleost Xenotoca eiseni, whose intraovarian embryos absorb nutrients from the maternal ovarian fluid through a specialized hindgutderived pseudoplacental structure called trophotaenia. Xenotoca eiseni VSP (Xe-VSP) is expressed in trophotaenia epithelium, an absorptive tissue functionally similar to zebrafish LREs. Notably, the apical distribution of Xe-VSP in trophotaenia epithelial cells closely resembles zebrafish VSP (Dr-VSP) distribution in zebrafish LREs, suggesting a shared role for VSP in absorptive tissues between the two species. Electrophysiological analysis using a heterologous expression system revealed that Xe-VSP preserves functional voltage sensors and phosphatase activity with the leftward shifted voltage sensitivity compared with zebrafish VSP (Dr-VSP). We also identified a single amino acid variation in the S4 helix of Xe-VSP as one of the factors contributing to the leftward shifted voltage sensitivity. This study highlights the biological variation and significance of VSP in various animal species, as well as hinting at the potential role of VSP in nutrient absorption in X. eiseni trophotaenia. NEW & NOTEWORTHY We investigate the voltage-sensing phosphatase (VSP) in Xenotoca eiseni, a viviparous fish whose intraovarian embryos utilize trophotaenia for nutrient absorption. Although X. eiseni VSP (Xe-VSP) shares key features with known VSPs, its distinct voltage sensitivity arises from species-specific amino acid variation. Xe-VSP in trophotaenia epithelium suggests its involvement in nutrient absorption, similar to VSP in zebrafish enterocytes and potentially in species with similar absorptive cells. Our findings highlight the potential role of VSP across species. [ABSTRACT FROM AUTHOR]

This article discusses the role of phosphoinositide-3 kinase (PI3K) activity in severe eosinophilic asthma with recurrent airway infections and neutrophilic exacerbations. The study found that patients with severe eosinophilic asthma and recurrent airway infections have increased PI3K activity, reduced histone deacetylase (HDAC) expression, and reduced macrophage receptor with collagenous structure (MARCO) expression. These findings suggest that PI3K activation may contribute to steroid subsensitivity and increased susceptibility to infections in this subgroup of patients. The study also highlights the potential for therapeutic studies of PI3K antagonism as a novel treatment strategy for these patients. [Extracted from the article]

Phosphatidylinositol (PI) is the precursor lipid for the minor phosphoinositides (PPIns), which are critical for multiple functions in all eukaryotic cells. It is poorly understood how phosphatidylinositol, which is synthesized in the ER, reaches those membranes where PPIns are formed. Here, we used VT01454, a recently identified inhibitor of class I PI transfer proteins (PITPs), to unravel their roles in lipid metabolism, and solved the structure of inhibitor-bound PITPNA to gain insight into the mode of inhibition. We found that class I PITPs not only distribute PI for PPIns production in various organelles such as the plasma membrane (PM) and late endosomes/lysosomes, but that their inhibition also significantly reduced the levels of phosphatidylserine, di- and triacylglycerols, and other lipids, and caused prominent increases in phosphatidic acid. While VT01454 did not inhibit Golgi PI4P formation nor reduce resting PM PI(4,5)P2 levels, the recovery of the PM pool of PI(4,5)P2 after receptor-mediated hydrolysis required both class I and class II PITPs. Overall, these studies show that class I PITPs differentially regulate phosphoinositide pools and affect the overall cellular lipid landscape. Synopsis: Class I phosphatidylinositol (PI) transfer proteins (PITPs) have been proposed to supply PI to the lipid kinases that produce mono-phosphorylated phosphoinositides, PI4P and PI3P. A novel inhibitor of class I PITPs illuminates how these proteins contribute to the maintenance of various phosphoinositide pools in intact cells. PITPs provide PI to the plasma membrane and late endosomes for generation of PI4P in those organelle membranes. Acute PITP inhibition does not affect PI4P levels in the Golgi and only has a small impact on PI3P levels in early endosomes. PI(4,5)P2 maintenance in the plasma membrane upon phospholipase C activation requires both class I and class II PITPs. The crystal structure of inhibitor-bound PITPNA reveals an "open" structure identical to the reported cargo-free structures. PTIP Inhibition affects the levels of several lipid classes. Inhibition of specific lipid transfer proteins reveals how pools of key signalling lipids are maintained in the plasma membrane and late endosome. [ABSTRACT FROM AUTHOR]

C‐peptide, a byproduct of insulin synthesis believed to be biologically inert, is emerging as a multifunctional molecule. C‐peptide serves an anti‐inflammatory and anti‐atherogenic role in type 1 diabetes mellitus (T1DM) and early T2DM. C‐peptide protects endothelial cells by activating AMP‐activated protein kinase α, thus suppressing the activity of NAD(P)H oxidase activity and reducing reactive oxygen species (ROS) generation. It also prevents apoptosis by regulating hyperglycemia‐induced p53 upregulation and mitochondrial adaptor p66shc overactivation, as well as reducing caspase‐3 activity and promoting expression of B‐cell lymphoma‐2. Additionally, C‐peptide suppresses platelet‐derived growth factor (PDGF)‐beta receptor and p44/p42 mitogen‐activated protein (MAP) kinase phosphorylation to inhibit vascular smooth muscle cells (VSMC) proliferation. It also diminishes leukocyte adhesion by virtue of its capacity to abolish nuclear factor kappa B (NF‐kB) signaling, a major pro‐inflammatory cascade. Consequently, it is envisaged that supplementation of C‐peptide in T1DM might ameliorate or even prevent end‐organ damage. In marked contrast, C‐peptide increases monocyte recruitment and migration through phosphoinositide 3‐kinase (PI‐3 kinase)‐mediated pathways, induces lipid accumulation via peroxisome proliferator‐activated receptor γ upregulation, and stimulates VSMC proliferation and CD4+ lymphocyte migration through Src‐kinase and PI‐3K dependent pathways. Thus, it promotes atherosclerosis and microvascular damage in late T2DM. Indeed, C‐peptide is now contemplated as a potential biomarker for insulin resistance in T2DM and linked to increased coronary artery disease risk. This shift in the understanding of the pathophysiology of diabetes from being a single hormone deficiency to a dual hormone disorder warrants a careful consideration of the role of C‐peptide as a unique molecule with promising diagnostic, prognostic, and therapeutic applications. [ABSTRACT FROM AUTHOR]

The Hippo pathway effectors Yes-associated protein 1 (YAP) and its homolog TAZ are transcriptional coactivators that control gene expression by binding to TEA domain (TEAD) family transcription factors. The YAP/TAZ–TEAD complex is a key regulator of cancer-specific transcriptional programs, which promote tumor progression in diverse types of cancer, including breast cancer. Despite intensive efforts, the YAP/TAZ–TEAD complex in cancer has remained largely undruggable due to an incomplete mechanistic understanding. Here, we report that nuclear phosphoinositides function as cofactors that mediate the binding of YAP/TAZ to TEADs. The enzymatic products of phosphoinositide kinases PIPKIα and IPMK, including phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) and phosphatidylinositol 3,4,5-trisphosphate (P(I3,4,5)P3), bridge the binding of YAP/TAZ to TEAD. Inhibiting these kinases or the association of YAP/TAZ with PI(4,5)P2 and PI(3,4,5)P3 attenuates YAP/TAZ interaction with the TEADs, the expression of YAP/TAZ target genes, and breast cancer cell motility. Although we could not conclusively exclude the possibility that other enzymatic products of IPMK such as inositol phosphates play a role in the mechanism, our results point to a previously unrecognized role of nuclear phosphoinositide signaling in control of YAP/TAZ activity and implicate this pathway as a potential therapeutic target in YAP/TAZ-driven breast cancer. Synopsis: The YAP-TEAD transcriptional regulators play a pivotal role in tumorigenesis, but activation of this pathway remains incompletely understood at the molecular level. This study implicates nuclear inositol phospholipids as molecular glue mediating YAP-TEAD interaction, required for full activation of the YAP-TEAD complex in breast cancer. YAP binds to phosphoinositide species via a polybasic motif located in its TEAD-binding domain. Phosphoinositides generated by lipid kinases, i.e., PIPKIa and IPMK, facilitate the binding of YAP to TEAD. Inhibition of PIPKIa and IPMK, or blockade of phosphoinositide binding to YAP, diminishes the expression of YAP target genes. The phosphoinositide-driven enhancement of YAP-TEAD complex formation promotes breast cancer cell motility. Phosphoinositide products of lipid kinases PIPKIa and IPMK bind to the transcriptional co-activator YAP and enhance its interaction with the transcription factor TEAD. [ABSTRACT FROM AUTHOR]

According to the World Health Organization, Chagas disease (CD) is the most prevalent poverty-promoting neglected tropical disease. Alarmingly, climate change is accelerating the geographical spreading of CD causative parasite, Trypanosoma cruzi, which additionally increases infection rates. Still, CD treatment remains challenging due to a lack of safe and efficient drugs. In this work, we analyze the viability of T. cruzi Akt-like kinase (TcAkt) as drug target against CD including primary structural and functional information about a parasitic Akt protein. Nuclear Magnetic Resonance derived information in combination with Molecular Dynamics simulations offer detailed insights into structural properties of the pleckstrin homology (PH) domain of TcAkt and its binding to phosphatidylinositol phosphate ligands (PIP). Experimental data combined with Alpha Fold proposes a model for the mechanism of action of TcAkt involving a PIP-induced disruption of the intramolecular interface between the kinase and the PH domain resulting in an open conformation enabling TcAkt kinase activity. Further docking experiments reveal that TcAkt is recognized by human inhibitors PIT-1 and capivasertib, and TcAkt inhibition by UBMC-4 and UBMC-6 is achieved via binding to TcAkt kinase domain. Our in-depth structural analysis of TcAkt reveals potential sites for drug development against CD, located at activity essential regions. [ABSTRACT FROM AUTHOR]

The TTG2 transcription factor of Arabidopsis regulates a set of epidermal traits, including the differentiation of leaf trichomes, flavonoid pigment production in cells of the inner testa (or seed coat) layer and mucilage production in specialized cells of the outer testa layer. Despite the fact that TTG2 has been known for over twenty years as an important regulator of multiple developmental pathways, little has been discovered about the downstream mechanisms by which TTG2 co-regulates these epidermal features. In this study, we present evidence of phosphoinositide lipid signaling as a mechanism for the regulation of TTG2-dependent epidermal pathways. Overexpression of the AtPLC1 gene rescues the trichome and seed coat phenotypes of the ttg2-1 mutant plant. Moreover, in the case of seed coat color rescue, AtPLC1 overexpression restored expression of the TTG2 flavonoid pathway target genes, TT12 and TT13/AHA10. Consistent with these observations, a dominant AtPLC1 T-DNA insertion allele (plc1-1D) promotes trichome development in both wild-type and ttg2-3 plants. Also, AtPLC1 promoter:GUS analysis shows expression in trichomes and this expression appears dependent on TTG2. Taken together, the discovery of a genetic interaction between TTG2 and AtPLC1 suggests a role for phosphoinositide signaling in the regulation of trichome development, flavonoid pigment biosynthesis and the differentiation of mucilage-producing cells of the seed coat. This finding provides new avenues for future research at the intersection of the TTG2-dependent developmental pathways and the numerous molecular and cellular phenomena influenced by phospholipid signaling. [ABSTRACT FROM AUTHOR]

Synaptojanin-1 (SJ1) is a major neuronal-enriched PI(4, 5)P2 4-and 5-phosphatase implicated in the shedding of endocytic factors during endocytosis. A mutation (R258Q) that impairs selectively its 4-phosphatase activity causes Parkinsonism in humans and neurological defects in mice (SJ1RQKI mice). Studies of these mice showed, besides an abnormal assembly state of endocytic factors at synapses, the presence of dystrophic nerve terminals selectively in a subset of nigro-striatal dopamine (DA)-ergic axons, suggesting a special lability of DA neurons to the impairment of SJ1 function. Here we have further investigated the impact of SJ1 on DA neurons using iPSC-derived SJ1 KO and SJ1RQKI DA neurons and their isogenic controls. In addition to the expected enhanced clustering of endocytic factors in nerve terminals, we observed in both SJ1 mutant neuronal lines increased cilia length. Further analysis of cilia of SJ1RQDA neurons revealed abnormal accumulation of the Ca2+ channel Cav1.3 and of ubiquitin chains, suggesting a defect in the clearing of ubiquitinated proteins at the ciliary base, where a focal concentration of SJ1 was observed. We suggest that SJ1 may contribute to the control of ciliary protein dynamics in DA neurons, with implications on cilia-mediated signaling. [ABSTRACT FROM AUTHOR]

In the last three decades, the presence of phospholipids in the nucleus has been shown and thoroughly investigated. A considerable amount of interest has been raised about nuclear inositol lipids, mainly because of their role in signaling acting. Here, we review the main issues of nuclear phospholipid localization and the role of nuclear inositol lipids and their related enzymes in cellular signaling, both in physiological and pathological conditions. [ABSTRACT FROM AUTHOR]

Background: Colorectal cancer (CRC) lacks established biomarkers or molecular targets for predicting or enhancing radiation response. Phosphatidylinositol-3,4,5-triphosphate-dependent Rac exchange factor 2 (PREX2) exhibits intricate implications in tumorigenesis and progression. Nevertheless, the precise role and underlying mechanisms of PREX2 in CRC radioresistance remain unclear. Methods: RNA-seq was employed to identify differentially expressed genes between radioresistant CRC cell lines and their parental counterparts. PREX2 expression was scrutinized using Western blotting, real-time PCR, and immunohistochemistry. The radioresistant role of PREX2 was assessed through in vitro colony formation assay, apoptosis assay, comet assay, and in vivo xenograft tumor models. The mechanism of PREX2 was elucidated using RNA-seq and Western blotting. Finally, a PREX2 small-molecule inhibitor, designated PREX-in1, was utilized to enhance the efficacy of ionizing radiation (IR) therapy in CRC mouse models. Results: PREX2 emerged as the most significantly upregulated gene in radioresistant CRC cells. It augmented the radioresistant capacity of CRC cells and demonstrated potential as a marker for predicting radioresistance efficacy. Mechanistically, PREX2 facilitated DNA repair by upregulating DNA-PKcs, suppressing radiation-induced immunogenic cell death, and impeding CD8+ T cell infiltration through the cGAS/STING/IFNs pathway. In vivo, the blockade of PREX2 heightened the efficacy of IR therapy. Conclusions: PREX2 assumes a pivotal role in CRC radiation resistance by inhibiting the cGAS/STING/IFNs pathway, presenting itself as a potential radioresistant biomarker and therapeutic target for effectively overcoming radioresistance in CRC. [ABSTRACT FROM AUTHOR]

Neuroinflammation is the major cause of neurodegenerative disorders such as Alzheimer's and Parkinson's disease. Currently available drugs present relatively low efficacy and are not capable of modifying the course of the disease or delaying its progression. Identifying well-tolerated and brain-penetrant agents of plant origin could fulfil the pressing need for novel treatment techniques for neuroinflammation. Attention has been drawn to a large family of flavonoids in citrus fruits, which may function as strong nutraceuticals in slowing down the development and progression of neuroinflammation. This review is aimed at elucidating and summarizing the effects of the flavonoid tangeretin (TAN) in the management of neuroinflammation-mediated neurodegenerative disorders. A literature survey was performed using various resources, including ScienceDirect, PubMed, Google Scholar, Springer, and Web of Science. The data revealed that TAN exhibited immense neuroprotective effects in addition to its anti-oxidant, anti-diabetic, and peroxisome proliferator-activated receptor-γ agonistic effects. The effects of TAN are mainly mediated through the inhibition of oxidative and inflammatory pathways via regulating multiple signaling pathways, including c-Jun N-terminal kinase, phosphoinositide 3-kinase, mitogen-activated protein kinase, nuclear factor erythroid-2-related factor 2, extracellular-signal-regulated kinase, and CRE-dependent transcription. In conclusion, the citrus flavonoid TAN has the potential to prevent neuronal death mediated by neuroinflammatory pathways and can be developed as an auxiliary therapeutic agent in the management of neurodegenerative disorders. [ABSTRACT FROM AUTHOR]

The motility of microglia involves intracellular signaling pathways that are predominantly controlled by changes in cytosolic Ca 2+ and activation of PI3K/Akt (phosphoinositide-3-kinase/protein kinase B). In this letter, we develop a novel biophysical model for cytosolic Ca 2+ activation of the PI3K/Akt pathway in microglia where Ca 2+ influx is mediated by both P2Y purinergic receptors (P2YR) and P2X purinergic receptors (P2XR). The model parameters are estimated by employing optimization techniques to fit the model to phosphorylated Akt (pAkt) experimental modeling/in vitro data. The integrated model supports the hypothesis that Ca 2+ influx via P2YR and P2XR can explain the experimentally reported biphasic transient responses in measuring pAkt levels. Our predictions reveal new quantitative insights into P2Rs on how they regulate Ca 2+ and Akt in terms of physiological interactions and transient responses. It is shown that the upregulation of P2X receptors through a repetitive application of agonist results in a continual increase in the baseline [Ca 2+ ], which causes the biphasic response to become a monophasic response which prolongs elevated levels of pAkt. [ABSTRACT FROM AUTHOR]

EH domain‐containing protein 2 (EHD2) is a member of the EHD protein family and is mainly located in the plasma membrane, but can also be found in the cytoplasm and endosomes. EHD2 is also a nuclear‐cytoplasmic shuttle protein. After entering the cell nuclear, EHD2 acts as a corepressor of transcription to inhibit gene transcription. EHD2 regulates a series of biological processes. As a key regulator of endocytic transport, EHD2 is involved in the formation and maintenance of endosomal tubules and vesicles, which are critical for the intracellular transport of proteins and other substances. The N‐terminal of EHD2 is attached to the cell membrane, while its C‐terminal binds to the actin‐binding protein. After binding, EHD2 connects with the actin cytoskeleton, forming the curvature of the membrane and promoting cell endocytosis. EHD2 is also associated with membrane protein trafficking and receptor signaling, as well as in glucose metabolism and lipid metabolism. In this review, we highlight the recent advances in the function of EHD2 in various cellular processes and its potential implications in human diseases such as cancer and metabolic disease. We also discussed the prospects for the future of EHD2. EHD2 has a broad prospect as a therapeutic target for a variety of diseases. Further research is needed to explore its mechanism, which could pave the way for the development of targeted treatments. Significance statement: The overview and biological function of EH domain‐containing protein 2 (EHD2) are discussed in this article.The article focuses on the role of EHD2 in metabolic diseases and cancer.The article also discussed the prospects for the future of EHD2 (Figure 1). [ABSTRACT FROM AUTHOR]

Phosphatidylinositols and their phosphorylated derivatives, known as phosphoinositides, are crucial in cellular processes, with their abnormalities linked to various diseases. Thus, identifying and measuring phosphoinositide levels in tissues are crucial for understanding their contributions to cellular processes and disease development. One powerful technique for mapping the spatial distribution of molecules in biological samples is matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI). This technique allows for the simultaneous detection and analysis of multiple lipid classes in situ, making it invaluable for unbiased lipidomic studies. However, detecting phosphoinositides with MALDI-MSI is challenging due to their relatively low abundance in tissues and complex matrix effects. Addressing this, our study focused on optimizing matrix selection and thickness for better detection of phosphatidylinositols and their phosphorylated forms in mouse kidney tissues. Various matrices were assessed, including 9AA, DAN, CMBT, and DHA, adjusting their coating to improve ionization efficiency. Our results demonstrate that DAN, DHA, and CMBT matrices produced high-intensity chemical images of phosphatidylinositol distributions within kidney sections. These matrices, particularly DAN, DHA, and CMBT, allowed the identification of even low-abundance phosphoinositides, through tentative identifications. Notably, DAN and DHA served as optimal candidates due to their prominent detection and ability to map a majority of phosphatidylinositol species, while CMBT showed potential detection capability for phosphatidylinositol triphosphate compounds. These findings not only provide valuable insights for future research on the involvement of phosphoinositides in kidney pathophysiology, but also propose the use of the identified optimal matrices, particularly DAN and DHA, as the preferred choices for enhanced detection and mapping of these lipid species in future studies. [ABSTRACT FROM AUTHOR]

Septins play a key regulatory role in cell division, cytokinesis, and cell polar growth of the rice blast fungus (Magnaporthe oryzae). We found that the organization of the septin ring, which is essential for appressorium-mediated infection in M. oryzae, requires long-chain fatty acids (LCFAs), which act as mediators of septin organization at membrane interfaces. However, it is unclear how septin ring formation and LCFAs regulate the pathogenicity of the rice blast fungus. In this study, a novel protein was named MoLfa1 because of its role in LCFAs utilization. MoLfa1 affects the utilization of LCFAs, lipid metabolism, and the formation of the septin ring by binding with phosphatidylinositol phosphates (PIPs), thereby participating in the construction of penetration pegs of M. oryzae. In addition, MoLfa1 is localized in the endoplasmic reticulum (ER) and interacts with the ER-related protein MoMip11 to affect the phosphorylation level of Mps1. (Mps1 is the core protein in the MPS1-MAPK pathway.) In conclusion, MoLfa1 affects conidia morphology, appressorium formation, lipid metabolism, LCFAs utilization, septin ring formation, and the Mps1-MAPK pathway of M. oryzae, influencing pathogenicity. [ABSTRACT FROM AUTHOR]

Background: Phosphoinositide 3-kinases (PI3Ks) are critical regulators of diverse cellular functions and have emerged as promising targets in cancer therapy. Despite significant progress, existing PI3K inhibitors encounter various challenges such as suboptimal bioavailability, potential off-target effects, restricted therapeutic indices, and cancer-acquired resistance. Hence, novel inhibitors that overcome some of these challenges are needed. Here, we describe the characterization of KTC1101, a novel pan-PI3K inhibitor that simultaneously targets tumor cell proliferation and the tumor microenvironment. Our studies demonstrate that KTC1101 significantly increases the anti-PD-1 efficacy in multiple pre-clinical mouse models. Methods: KTC1101 was synthesized and characterized employing chemical synthesis, molecular modeling, Nuclear Magnetic Resonance (NMR), and mass spectrometry. Its target specificity was confirmed through the kinase assay, JFCR39 COMPARE analysis, and RNA-Seq analysis. Metabolic stability was verified via liver microsome and plasma assays, pharmacokinetics determined by LC–MS/MS, and safety profile established through acute toxicity assays to determine the LD50. The antiproliferative effects of KTC1101 were evaluated in a panel of cancer cell lines and further validated in diverse BALB/c nude mouse xenograft, NSG mouse xenograft and syngeneic mouse models. The KTC1101 treatment effect on the immune response was assessed through comprehensive RNA-Seq, flow cytometry, and immunohistochemistry, with molecular pathways investigated via Western blot, ELISA, and qRT-PCR. Results: KTC1101 demonstrated strong inhibition of cancer cell growth in vitro and significantly impeded tumor progression in vivo. It effectively modulated the Tumor Microenvironment (TME), characterized by increased infiltration of CD8+ T cells and innate immune cells. An intermittent dosing regimen of KTC1101 enhanced these effects. Notably, KTC1101 synergized with anti-PD-1 therapy, significantly boosting antitumor immunity and extending survival in preclinical models. Conclusion: KTC1101's dual mechanism of action—directly inhibiting tumor cell growth and dynamically enhancing the immune response— represents a significant advancement in cancer treatment strategies. These findings support incorporating KTC1101 into future oncologic regimens to improve the efficacy of immunotherapy combinations. [ABSTRACT FROM AUTHOR]

Voltage-gated sodium channels (Naᵥ) are membrane proteins which open to facilitate the inward flux of sodium ions into excitable cells. In response to stimuli, Naᵥ channels transition from the resting, closed state to an open, conductive state, before rapidly inactivating. Dysregulation of this functional cycle due to mutations causes diseases including epilepsy, pain conditions, and cardiac disorders, making Naᵥ channels a significant pharmacological target. Phosphoinositides are important lipid cofactors for ion channel function. The phosphoinositide PI(4,5)P2 decreases Naᵥ1.4 activity by increasing the difficulty of channel opening, accelerating fast inactivation and slowing recovery from fast inactivation. Using multiscale molecular dynamics simulations, we show that PI(4,5)P2 binds stably to inactivated Naᵥ at a conserved site within the DIV S4-S5 linker, which couples the voltage-sensing domain (VSD) to the pore. As the Naᵥ C-terminal domain is proposed to also bind here during recovery from inactivation, we hypothesize that PI(4,5)P2 prolongs inactivation by competitively binding to this site. In atomistic simulations, PI(4,5)P2 reduces the mobility of both the DIV S4-S5 linker and the DIII-IV linker, responsible for fast inactivation, slowing the conformational changes required for the channel to recover to the resting state. We further show that in a resting state Naᵥ model, phosphoinositides bind to VSD gating charges, which may anchor them and impede VSD activation. Our results provide a mechanism by which phosphoinositides alter the voltage dependence of activation and the rate of recovery from inactivation, an important step for the development of novel therapies to treat Naᵥ-related diseases. [ABSTRACT FROM AUTHOR]

Cell motility processes highly depend on the membrane distribution of Phosphoinositides, giving rise to cytoskeleton reshaping and membrane trafficking events. Membrane contact sites serve as platforms for direct lipid exchange and calcium fluxes between two organelles. Here, we show that VAPA, an ER transmembrane contact site tether, plays a crucial role during cell motility. CaCo2 adenocarcinoma epithelial cells depleted for VAPA exhibit several collective and individual motility defects, disorganized actin cytoskeleton and altered protrusive activity. During migration, VAPA is required for the maintenance of PI(4)P and PI(4,5)P2 levels at the plasma membrane, but not for PI(4)P homeostasis in the Golgi and endosomal compartments. Importantly, we show that VAPA regulates the dynamics of focal adhesions (FA) through its MSP domain, is essential to stabilize and anchor ventral ER-PM contact sites to FA, and mediates microtubule-dependent FA disassembly. To conclude, our results reveal unknown functions for VAPA-mediated membrane contact sites during cell motility and provide a dynamic picture of ER-PM contact sites connection with FA mediated by VAPA. [ABSTRACT FROM AUTHOR]

Regulator of ribosome synthesis 1 (RRS1), a crucial regulatory factor in ribosome biogenesis, exerts a remarkable impact on the progression of breast cancer (BC). However, the exact mechanisms and pathways have not yet been fully elucidated. To investigate the impact of RRS1 on BC growth and metastasis, along with its underlying mechanisms. We discovered that RRS1 is overexpressed in BC tissues and cell lines. This study aims to regulate the level of RRS1 through lentiviral transfection technology to explore its potential function in BC cells. Knockdown of RRS1 resulted in the inhibition of cell proliferation, invasion, and migration, whereas overexpression had the opposite effects. We firstly identified the interaction between RRS1 and Glucose-Regulated Protein 78 (GRP78) using Co-immunoprecipitation (Co-IP) combined with mass spectrometry analysis, providing evidences of co-localization and positive regulation between RRS1 and GRP78. We observed that RRS1 inhibited the degradation of GRP78 through the ubiquitin–proteasome pathway, resulting in the stabilization of GRP78. In addition, our findings suggested that RRS1 promoted BC progression by activating the GRP78-mediated phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT) signaling pathway. In conclusion, this newly discovered RRS1/GRP78 signaling axis provides a molecular and theoretical basis for further exploring the mechanisms of breast cancer invasion and metastasis. [ABSTRACT FROM AUTHOR]

Phosphoinositides are amphipathic lipid molecules derived from phosphatidylinositol that represent low abundance components of biological membranes. Rather than serving as mere structural elements of lipid bilayers, they represent molecular switches for a broad range of biological processes, including cell signaling, membrane dynamics and remodeling, and many other functions. Here, we focus on the molecular mechanisms that turn phosphoinositides into molecular switches and how the dysregulation of these processes can lead to disease. [ABSTRACT FROM AUTHOR]

Lamins, the nuclear intermediate filaments, are important regulators of nuclear structural integrity as well as nuclear functional processes such as DNA transcription, replication and repair, and epigenetic regulations. A portion of phosphorylated lamin A/C localizes to the nuclear interior in interphase, forming a lamin A/C pool with specific properties and distinct functions. Nucleoplasmic lamin A/C molecular functions are mainly dependent on its binding partners; therefore, revealing new interactions could give us new clues on the lamin A/C mechanism of action. In the present study, we show that lamin A/C interacts with nuclear phosphoinositides (PIPs), and with nuclear myosin I (NM1). Both NM1 and nuclear PIPs have been previously reported as important regulators of gene expression and DNA damage/repair. Furthermore, phosphorylated lamin A/C forms a complex with NM1 in a phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2)-dependent manner in the nuclear interior. Taken together, our study reveals a previously unidentified interaction between phosphorylated lamin A/C, NM1, and PI(4,5)P2 and suggests new possible ways of nucleoplasmic lamin A/C regulation, function, and importance for the formation of functional nuclear microdomains. [ABSTRACT FROM AUTHOR]

Aim: The etiology of bipolar disorder (BD) remains unknown; however, lipid abnormalities in BD have received increasing attention in recent years. In this study, we examined the expression levels of enzyme proteins associated with the metabolic pathway of phosphoinositides (PIs) and their downstream effectors, protein kinase B (Akt1) and glycogen synthase kinase 3β (GSK3β), which have been assumed to be the targets of mood stabilizers such as lithium, in the postmortem brains of patients with BD. Methods: The protein expression levels of phosphatidylinositol 4‐phosphate 5‐kinase type‐1 gamma (PIP5K1C), phosphatidylinositol 4‐kinase alpha (PIK4CA), phosphatase and tensin homolog deleted from chromosome 10 (PTEN), Akt1, and GSK3β were measured using enzyme‐linked immunosorbent assays and multiplex fluorescent bead‐based immunoassays in the prefrontal cortex (PFC). Specifically, PTEN, Akt1, GSK3β, and PIP5K1C were measured in seven BD patients and 48 controls. Additionally, PIK4CA was analyzed in 10 cases and 34 controls. Results: PTEN expression levels were markedly decreased in the PFCs of patients with BD, whereas those of Akt and GSK3β were prominently elevated. Moreover, patients medicated with lithium exhibited higher Akt1 expression levels and lower PTEN expression levels in comparison with the untreated group. Conclusion: Our results suggest that the expression levels of Akt1/GSK3β and its upstream regulator PTEN are considerably altered. [ABSTRACT FROM AUTHOR]

This study aims to examine the impact and underlying mechanism of micro ribonucleic acid-515-5p on the biological behavior of colorectal cancer cells. The study conducted an experiment categorizing into control (blank), colorectal cancer, and micro ribonucleic acid-515-5p overexpression groups. Western blot, quantitative polymerase chain reaction techniques, cell counting kit-8, and Transwell methods were used to detect the growth and metastasis ability of the cells, the expression level of the related protein and messenger ribonucleic acid. The micro ribonucleic acid-515-5p messenger ribonucleic acid was found to be significantly reduced in colorectal cancer group cells compared to control group cells. The proliferation activity and migration number of the cells in micro ribonucleic acid-515-5p overexpression group were reduced compared to the colorectal cancer group cells. The first apoptosis signal, B-cell lymphoma 2-associated X protein, and first apoptosis signal ligand in the micro ribonucleic acid-515-5p overexpression group were higher compared to the colorectal cancer group, while the expression level of B-cell lymphoma 2 was lower compared to the colorectal cancer group. Protein kinase B, phosphoinositide 3-kinase, messenger ribonucleic acid and protein in the micro ribonucleic acid-515-5p overexpression group were reduced than the colorectal cancer group. The overexpression of micro ribonucleic acid-515-5p has been observed to exert inhibitory effects on proliferation, migration, and invasion of colorectal cancer cells, while induce apoptosis of colorectal cancer cells by inhibiting phosphoinositide 3-kinase/protein kinase B signaling pathway. [ABSTRACT FROM AUTHOR]

To investigate the effect of liraglutide on myocardial injury in acute myocardial infarction rats by regulating phosphoinositide 3 kinase/protein kinase B signaling. Sprague-Dawley rats were randomly divided into sham group, model group, liraglutide group (0.6 mg/kg liraglutide), LY294002 (phosphoinositide 3 kinase inhibitor) (1.5 mg/kg LY294002) and liraglutide+LY294002 (LY294002 after liraglutide), with 12 rats in each group. Except for the sham group, the acute myocardial infarction rat model was constructed by coronary artery ligation. After successful mold making, the drug group was treated according to the administration method of each group, and the group was injected with equal amount of normal saline intraperitoneally for 2 w. Cardiac function was measured by echocardiography, hemodynamics were recorded by polygraph, myocardial histopathological changes were detected by hematoxylin and eosin staining, the degree of fibrosis was observed by Masson staining, and the expression of autophagy-related factors and proteins related to phosphoinositide 3 kinase/protein kinase B signaling pathway was detected in rats by Western blot. As compared to the sham group, a significant decrease in cardiac function, the degree of myocardial tissue damage and the degree of fibrosis, the expression of autophagy-related proteins and phosphoinositide 3 kinase/protein kinase B pathway-related proteins was significantly decreased; compared to the model group, the cardiac function, degree of fibrosis in liraglutide rats, the expression of autophagy-related proteins and phosphoinositide 3 kinase/protein kinase B pathway-related proteins was significantly increased; in contrast to the liraglutide group, cardiac function was significantly reduced in the liraglutide+LY2940002 group, the degree of myocardial tissue damage and the degree of fibrosis, the expression of autophagy-related proteins and phosphoinositide 3 kinase/protein kinase B pathway-related proteins was significantly decreased; compared with the LY2940002 group, the extent of cardiac function, myocardial tissue damage and fibrosis were alleviated in liraglutide+LY2940002 group rats, the expression of autophagy-related proteins and phosphoinositide 3 kinase/protein kinase B pathway-related proteins was significantly increased. Liraglutide can improve the myocardial injury and protect the cardiac function by activating the phosphoinositide 3 kinase/protein kinase B signal pathway in acute myocardial infarction rats. [ABSTRACT FROM AUTHOR]

The lipid phosphatidylinositol 4-phosphate (PI4P) plays a master regulatory role at Golgi membranes, orchestrating membrane budding, non-vesicular lipid transport and membrane organization. It follows that harmonious Golgi function requires strictly maintained PI4P homeostasis. One of the most abundant PI4P effector proteins is the oxysterol binding protein (OSBP), a lipid transfer protein that exchanges trans-Golgi PI4P for ER cholesterol. Although this protein consumes PI4P as part of its lipid anti-porter function, whether it actively contributes to Golgi PI4P homeostasis has been questioned. Here, we employed a series of acute and chronic genetic manipulations, together with orthogonal targeting of OSBP, to interrogate its control over Golgi PI4P abundance. Modulating OSBP levels at ER:Golgi membrane contact sites produces reciprocal changes in PI4P levels. Additionally, we observe that OSBP has a high capacity for PI4P turnover, even at orthogonal organelle membranes. However, despite also visiting the plasma membrane, endogenous OSBP makes no impact on PI4P levels in this compartment. We conclude that OSBP is a major determinant of Golgi PI4P homeostasis. [ABSTRACT FROM AUTHOR]

Vacuolar ATPases (V-ATPases), proton pumps composed of 16 subunits, are necessary for a variety of cellular functions. Subunit "a" has four isoforms, a1–a4, each with a distinct cellular location. We identified a phosphoinositide (PIP) interaction motif, KXnK(R)IK(R), conserved in all four isoforms, and hypothesize that a/PIP interactions regulate V-ATPase recruitment/retention to different organelles. Among the four isoforms, a2 is enriched on Golgi with a2 mutations in the PIP motif resulting in cutis laxa. We hypothesize that the hydrophilic N-terminal (NT) domain of a2 contains a lipid-binding domain, and mutations in this domain prevent interaction with Golgi-enriched PIPs, resulting in cutis laxa. We recreated the cutis laxa-causing mutation K237_V238del, and a double mutation in the PIP-binding motif, K237A/V238A. Circular dichroism confirmed that there were no protein structure alterations. Pull-down assays with PIP-enriched liposomes revealed that wildtype a2NT preferentially binds phosphatidylinositol 4-phosphate (PI(4)P), while mutants decreased binding to PI(4)P. In HEK293 cells, wildtype a2NT was localized to Golgi and co-purified with microsomal membranes. Mutants reduced Golgi localization and membrane association. Rapamycin depletion of PI(4)P diminished a2NT-Golgi localization. We conclude that a2NT is sufficient for Golgi retention, suggesting the lipid-binding motif is involved in V-ATPase targeting and/or retention. Mutational analyses suggest a molecular mechanism underlying how a2 mutations result in cutis laxa. [ABSTRACT FROM AUTHOR]

Transient receptor potential (TRP) ion channels have diverse activation mechanisms including physical stimuli, such as high or low temperatures, and a variety of intracellular signaling molecules. Regulation by phosphoinositides and their derivatives is their only known common regulatory feature. For most TRP channels, phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] serves as a cofactor required for activity. Such dependence on PI(4,5)P2 has been demonstrated for members of the TRPM subfamily and for the epithelial TRPV5 and TRPV6 channels. Intracellular TRPML channels show specific activation by PI(3,5)P2. Structural studies uncovered the PI(4,5)P2 and PI(3,5)P2 binding sites for these channels and shed light on the mechanism of channel opening. PI(4,5)P2 regulation of TRPV1–4 as well as some TRPC channels is more complex, involving both positive and negative effects. This review discusses the functional roles of phosphoinositides in TRP channel regulation and molecular insights gained from recent cryo-electron microscopy structures. [ABSTRACT FROM AUTHOR]

Oxysterol-binding protein (OSBP)-related proteins (ORPs) 5 and 8 have been shown to deplete the lipid phosphatidylinositol 4-phosphate (PI4P) at sites of membrane contact between the endoplasmic reticulum (ER) and plasma membrane (PM). This is believed to be caused by transport of PI4P from the PM to the ER, where PI4P is degraded by an ER-localized SAC1 phosphatase. This is proposed to power the anti-port of phosphatidylserine (PS) lipids from ER to PM, up their concentration gradient. Alternatively, ORPs have been proposed to sequester PI4P, dependent on the concentration of their alternative lipid ligand. Here, we aimed to distinguish these possibilities in living cells by orthogonal targeting of PI4P transfer and degradation to PM-mitochondria contact sites. Surprisingly, we found that orthogonal targeting of SAC1 to mitochondria enhanced PM PI4P turnover independent of targeting to contact sites with the PM. This turnover could be slowed by knock-down of soluble ORP2, which also has a major impact on PM PI4P levels even without SAC1 over-expression. The data reveal a role for contact site-independent modulation of PM PI4P levels and lipid antiport. [ABSTRACT FROM AUTHOR]

Solute carrier family 27 member 5, a key enzyme in fatty acid transport and bile acid metabolism in the liver, is frequently expressed in low quantities in patients with hepatocellular carcinoma, resulting in poor prognosis. However, it is unclear whether SLC27A5 plays non‐canonical functions and regulates HCC progression. Here, an unexpected non‐canonical role of SLC27A5 is reported: regulating the alternative splicing of mRNA to inhibit the metastasis of HCC independently of its metabolic enzyme activity. Mechanistically, SLC27A5 interacts with IGF2BP3 to prevent its translocation into the nucleus, thereby inhibiting its binding to target mRNA and modulating PIP4K2A pre‐mRNA splicing. Loss of SLC27A5 results in elevated levels of the PIP4K2A‐S isoform, thus positively regulating phosphoinositide 3‐kinase signaling via enhanced p85 stability in HCC. SLC27A5 restoration by AAV‐Slc27a5 or IGF2BP3 RNA decoy oligonucleotides exerts an inhibitory effect on HCC metastasis with reduced expression of the PIP4K2A‐S isoform. Therefore, PIP4K2A‐S may be a novel target for treating HCC with SLC27A5 deficiency. [ABSTRACT FROM AUTHOR]

SUMMARY: Anionic phospholipids (PS, PA, PI, PIPs) are low‐abundant phospholipids with impactful functions in cell signaling, membrane trafficking and cell differentiation processes. They can be quickly metabolized and can transiently accumulate at defined spots within the cell or an organ to respond to physiological or environmental stimuli. As even a small change in their composition profile will produce a significant effect on biological processes, it is crucial to develop a sensitive and optimized analytical method to accurately detect and quantify them. While thin‐layer chromatography (TLC) separation coupled with gas chromatography (GC) detection methods already exist, they do not allow for precise, sensitive, and accurate quantification of all anionic phospholipid species. Here we developed a method based on high‐performance liquid chromatography (HPLC) combined with two‐dimensional mass spectrometry (MS2) by MRM mode to detect and quantify all molecular species and classes of anionic phospholipids in one shot. This method is based on a derivatization step by methylation that greatly enhances the ionization, the separation of each peak, the peak resolution as well as the limit of detection and quantification for each individual molecular species, and more particularly for PA and PS. Our method universally works in various plant samples. Remarkably, we identified that PS is enriched with very long chain fatty acids in the roots but not in aerial organs of Arabidopsis thaliana. Our work thus paves the way for new studies on how the composition of anionic lipids is finely tuned during plant development and environmental responses. Significance Statement: While anionic phospholipids have key functions in plant cellular processes, their low concentration in biological samples and their low stability during the analysis complicate their quantification. Here, we present the first one‐shot analytical method for the profiling and quantification of all anionic phospholipid classes and species from plant tissues with unprecedented sensitivity. This method opens the way to future studies requiring a fine quantification of anionic phospholipids to understand their role in plant cell processes. [ABSTRACT FROM AUTHOR]

Background: Treatment failure following androgen deprivation therapy (ADT) presents a significant challenge in the management of advanced prostate cancer. Thus, understanding the genetic factors influencing this process could facilitate the development of personalized treatments and innovative therapeutic strategies. The phosphoinositide 3-kinase (PI3K)/AKT signaling pathway plays a pivotal role in controlling cell growth and tumorigenesis. We hypothesized that genetic variants within this pathway may affect the clinical outcomes of patients undergoing ADT for prostate cancer. Methods: We genotyped 399 single-nucleotide polymorphisms (SNPs) across 28 core PI3K/AKT pathway genes in a cohort of 630 patients with prostate cancer undergoing ADT. We assessed the potential association of the SNPs with patient survival. Functional analyses of the implicated genes were also performed to evaluate their effects on prostate cancer. Results: After multivariate Cox regression analysis and multiple testing correction, GABRB3 rs12591845 exhibited the most significant association with both overall and cancer-specific survivals (P < 0.003). A comprehensive pooled analysis of 16 independent gene expression datasets revealed elevated expression of GABRB3 in prostate cancer tissues compared to that in normal tissues (P < 0.001). Furthermore, gene set enrichment analysis unveiled differential enrichment of pathways such as myogenesis, interferon γ and α responses, and the MYC proto-oncogene pathway in tumors with elevated GABRB3 expression, implying a role for GABRB3 in prostate cancer. Conclusion: Our results suggest that rs12591845 could potentially serve as a valuable prognostic indicator for patients undergoing ADT. The potential role of GABRB3 in promoting prostate tumorigenesis is also highlighted. [ABSTRACT FROM AUTHOR]

Membrane polyphosphoinositides (PPIs) are lipid-signaling molecules that undergo metabolic turnover and influence a diverse range of cellular functions. PPIs regulate the activity and/or spatial localization of a number of actin-binding proteins (ABPs) through direct interactions; however, it is much less clear whether ABPs could also be an integral part in regulating PPI signaling. In this study, we show that ABP profilin1 (Pfn1) is an important molecular determinant of the cellular content of PI(4,5)P2 (the most abundant PPI in cells). In growth factor (EGF) stimulation setting, Pfn1 depletion does not impact PI(4,5)P2 hydrolysis but enhances plasma membrane (PM) enrichment of PPIs that are produced downstream of activated PI3-kinase, including PI(3,4,5)P3 and PI(3,4)P2, the latter consistent with increased PM recruitment of SH2-containing inositol 50 phosphatase (SHIP2) (a key enzyme for PI(3,4)P2 biosynthesis). Although Pfn1 binds to PPIs in vitro, our data suggest that Pfn1's affinity to PPIs and PM presence in actual cells, if at all, is negligible, suggesting that Pfn1 is unlikely to directly compete with SHIP2 for binding to PM PPIs. Additionally, we provide evidence for Pfn1's interaction with SHIP2 in cells and modulation of this interaction upon EGF stimulation, raising an alternative possibility of Pfn1 binding as a potential restrictive mechanism for PM recruitment of SHIP2. In conclusion, our findings challenge the dogma of Pfn1's binding to PM by PPI interaction, uncover a previously unrecognized role of Pfn1 in PI(4,5)P2 homeostasis and provide a new mechanistic avenue of how an ABP could potentially impact PI3K signaling byproducts in cells through lipid phosphatase control. [ABSTRACT FROM AUTHOR]

Introduction: Oncolytic virotherapy is a novel strategy for cancer treatment in humans and companion animals. Canine distemper virus (CDV) is known to induce apoptosis in tumor cells, thus serving as a potential candidate for oncolytic therapy. However, the mechanism of viral oncolytic activity is less studied and varies depending on the type of cancer and cell lines. Methods: In the present study, the susceptibility of the MCF-7 cell line to CDV infection was assessed using the CDV strain, which was confirmed previously through sequence analysis in the Vero cell line. The impact of CDV infection on cell proliferation and apoptosis was studied by evaluating the expression of four target genes including the myeloid cell leukemia 1 (MCL-1), phosphoinositide-3-kinase regulatory subunit 1 (PIK3R1), transcription factor (SP1), and DNA (cytosine-5)-methyltransferase 3A (DNMT3A). Results: CDV replication in the cells induced cytopathic effect and decreased in the cell proliferation rates compared to the uninfected control. MCL-1, SP1, and PIK3R1 gene expression was down-regulated, while the expression of DNMT3A was up-regulated 3 days post-infection. The expression levels of the target genes suggest that CDV may be inducing the intrinsic apoptotic pathway in the cancer cell line. Conclusion: Overall, the results strongly propose CDV strain as a potential candidate for cancer therapy after detailed studies. [ABSTRACT FROM AUTHOR]

In response to different immune challenges, immune cells migrate to specific sites in the body, where they perform their functions such as defense against infection, inflammation regulation, antigen recognition, and immune surveillance. Therefore, the migration ability is a fundamental aspect of immune cell function. Phosphoinositide signaling plays critical roles in modulating immune cell migration by controlling cell polarization, cytoskeletal rearrangement, protrusion formation, and uropod contraction. Upon chemoattractant stimulation, specific phosphoinositide kinases and phosphatases control the local phosphoinositide levels to establish polarized phosphoinositide distribution, which recruits phosphoinositide effectors to distinct subcellular locations to facilitate cell migration. In this Special Issue of "Molecular Mechanisms Underlying Cell Adhesion and Migration", we discuss the significance of phosphoinositide production and conversion by phosphoinositide kinases and phosphatases in the migration of different types of immune cells. [ABSTRACT FROM AUTHOR]

CD14 is a co-receptor of Toll-like receptor (TLR)− 4, with a critical role in innate immune responses. CD14 recognizes bacterial lipopolysaccharides, pathogen-, and damage-associated molecular patterns, thereby facilitating inflammatory immune responses. In addition to its well-established association with TLR4, CD14 is also implicated in TLR4-independent signaling, which leads to the apoptotic death of differentiated dendritic cells and activation of the noncanonical inflammasome pathway. CD14 also has a role beyond that of the immune responses. It contributes to tissue homeostasis by promoting the clearance of various apoptotic cells via recognizing externalized phosphatidylinositol phosphates. CD14 also has context-dependent roles, particularly in barrier tissues that include the skin and gastrointestinal tract. For example, CD14+ dendritic cells in the skin can induce immunostimulatory or immunosuppressive responses. In the gastrointestinal system, CD14 is involved in producing inflammatory cytokines in inflammatory bowel disease and maintaining of intestinal integrity. This review focuses on the multifaceted roles of CD14 in innate immunity and its potential regulatory functions in barrier tissues characterized by rapid cell renewal. By providing insights into the diverse functions of CD14, this review offers potential therapeutic implications for this versatile molecule in immune modulation and tissue homeostasis. • CD14, a co-receptor for TLR4, recognizes LPS, PAMPs, and DAMPs in innate immunity. • CD14 has a role in TLR4-dependent and TLR4-independent signaling pathways. • CD14 helps clear apoptotic cells by recognizing phospholipids externalized on the cell surface. • In barrier tissues, CD14 can mediate inflammatory and tolerogenic responses. • Knowledge of CD14 ligand interactions will inform specifically directed therapies. [ABSTRACT FROM AUTHOR]

Nitric oxide (NO) is a key signaling molecule affecting the response of plants to salt stress; however, the underlying molecular mechanism is poorly understood. In this study, we conducted a phenotype analysis and found that the small GTPase RABG3E (RAB7) promotes salt tolerance in Arabidopsis thaliana. NO promotes the S‐nitrosylation of RAB7 at Cys‐171, which in turn helps maintain the ion balance in salt‐stressed plants. Furthermore, the S‐nitrosylation of RAB7 at Cys‐171 enhances the enzyme's GTPase activity, thereby promoting vesicle trafficking and increasing its interaction with phosphatidylinositol phosphates—especially phosphatidylinositol‐4‐phosphate (PI4P). Exogenously applied PI4P increases vesicle trafficking and promotes salt tolerance depending on the S‐nitrosylation of RAB7 at Cys‐171. These findings illustrate a unique mechanism in salt tolerance, by which NO regulates vesicle trafficking and ion homeostasis through the S‐nitrosylation of RAB7 and its interaction with PI4P. Summary statement: Nitric oxide‐mediated S‐nitrosylation of a RAB GTPase RABG3E modulates vesicle transport relying on its interaction with phosphatidylinositol‐4‐phosphate to enhance salt tolerance in Arabidopsis thaliana. [ABSTRACT FROM AUTHOR]

Vacuolar H+-ATPases (V-ATPases) are highly conserved multisubunit enzymes that maintain the distinct pH of eukaryotic organelles. The integral membrane a-subunit is encoded by tissue- and organelle-specific isoforms, and its cytosolic N-terminal domain (aNT) modulates organellespecific regulation and targeting of V-ATPases. Organelle membranes have specific phosphatidylinositol phosphate (PIP) lipid enrichment linked to maintenance of organelle pH. In yeast, the aNT domains of the two a-subunit isoforms bind PIP lipids enriched in the organelle membranes where they reside; these interactions affect activity and regulatory properties of the V-ATPases containing each isoform. Humans have four asubunit isoforms, and we hypothesize that the aNT domains of these isoforms will also bind to specific PIP lipids. The a1 and a2 isoforms of human V-ATPase a-subunits are localized to endolysosomes and Golgi, respectively. We determined that bacterially expressed Hua1NT and Hua2NT bind specifically to endolysosomal PIP lipids PI(3)P and PI(3,5)P2 and Golgi enriched PI(4)P, respectively. Despite the lack of canonical PIP-binding sites, we identified potential binding sites in the HuaNT domains by sequence comparisons and existing subunit structures and models. We found that mutations at a similar location in the distal loops of both HuaNT isoforms compromise binding to their cognate PIP lipids, suggesting that these loops encode PIP specificity of the a-subunit isoforms. These data suggest a mechanism through which PIP lipid binding could stabilize and activate V-ATPases in distinct organelles. [ABSTRACT FROM AUTHOR]

Phosphoinositide 3-kinases (PI3K) and phosphoinositide 3-kinase-related protein kinases (PIKK) are two structurally related families of kinases that play vital roles in cell growth and DNA damage repair. Dysfunction of PIKK members and aberrant stimulation of the PI3K/AKT/mTOR signalling pathway are linked to a plethora of diseases including cancer. In recent decades, numerous inhibitors related to the PI3K/AKT/mTOR signalling have made great strides in cancer treatment, like copanlisib and sirolimus. Notably, most of the PIKK inhibitors (such as VX-970 and M3814) related to DNA damage response have also shown good efficacy in clinical trials. However, these drugs still require a suitable combination therapy to overcome drug resistance or improve antitumor activity. Based on the aforementioned facts, we summarised the efficacy of PIKK, PI3K, and AKT inhibitors in the therapy of human malignancies and the resistance mechanisms of targeted therapy, in order to provide deeper insights into cancer treatment. [ABSTRACT FROM AUTHOR]

Background & Aims: The class I‐ phosphatidylinositol‐3 kinases (PI3Ks) signalling is dysregulated in almost all human cancers whereas the isoform‐specific roles remain poorly investigated. We reported that the isoform δ (PI3Kδ) regulated epithelial cell polarity and plasticity and recent developments have heightened its role in hepatocellular carcinoma (HCC) and solid tumour progression. However, its role in cholangiocarcinoma (CCA) still lacks investigation. Approach & Results: Immunohistochemical analyses of CCA samples reveal a high expression of PI3Kδ in the less differentiated CCA. The RT‐qPCR and immunoblot analyses performed on CCA cells stably overexpressing PI3Kδ using lentiviral construction reveal an increase of mesenchymal and stem cell markers and the pluripotency transcription factors. CCA cells stably overexpressing PI3Kδ cultured in 3D culture display a thick layer of ECM at the basement membrane and a wide single lumen compared to control cells. Similar data are observed in vivo, in xenografted tumours established with PI3Kδ‐overexpressing CCA cells in immunodeficient mice. The expression of mesenchymal and stemness genes also increases and tumour tissue displays necrosis and fibrosis, along with a prominent angiogenesis and lymphangiogenesis, as in mice liver of AAV8‐based‐PI3Kδ overexpression. These PI3Kδ‐mediated cell morphogenesis and stroma remodelling were dependent on TGFβ/Src/Notch signalling. Whole transcriptome analysis of PI3Kδ using the cancer cell line encyclopedia allows the classification of CCA cells according to cancer progression. Conclusions: Overall, our results support the critical role of PI3Kδ in the progression and aggressiveness of CCA via TGFβ/src/Notch‐dependent mechanisms and open new directions for the classification and treatment of CCA patients. [ABSTRACT FROM AUTHOR]

Phosphoinositide 3‐kinase (PI3K) p110δ signalling negatively regulates the production of mouse IgE. However, there are disparities between the mouse and human IgE biology, and the role of PI3K p110δ in the production of human IgE is yet to be determined. To investigate the effect of PI3K p110δ inhibition in the production of human IgE we isolated human B cells from tonsil tissue and stimulated them with IL‐4 and anti‐CD40 antibody to induce class switching to IgE and IgG1 in the presence or absence of IC87114, a small molecule inhibitor of PI3K p110δ. Using FACS, RT‐PCR and ELISA we examined the effect of PI3K p110δ inhibition on IgE production and determined the mechanisms involved. Unlike in mice, we observed that PI3K p110δ inhibition significantly reduces the number of IgE+ switched cells and the amounts of secreted IgE in IL4 and anti‐CD40 cultures. However, the number of IgG1+ cells and secreted IgG1 were largely unaffected by PI3K p110δ inhibition. The expression levels of AID, ε and γ1 germinal transcripts or other factors involved in the regulation of CSR to IgE and IgG1 were also unaffected by IC87114. However, we found that IC87114 significantly decreases the proliferation of tonsil B cells stimulated with IL‐4 and anti‐CD40, specifically reducing the frequency of cells that had undergone 4 divisions or more. In addition, PI3K p110δ inhibition reduced the levels of IRF4 expression in IgE+ germinal centre‐like B cells leading to a block in plasma cell differentiation. In conclusion, PI3K p110δ signalling is required for the production of human IgE, which makes it a pharmacological target for the treatment of allergic disease. [ABSTRACT FROM AUTHOR]

African trypanosomes evade host immune clearance by antigenic variation, causing persistent infections in humans and animals. These parasites express a homogeneous surface coat of variant surface glycoproteins (VSGs). They transcribe one out of hundreds of VSG genes at a time from telomeric expression sites (ESs) and periodically change the VSG expressed by transcriptional switching or recombination. The mechanisms underlying the control of VSG switching and its developmental silencing remain elusive. We report that telomeric ES activation and silencing entail an on/ off genetic switch controlled by a nuclear phosphoinositide signaling system. This system includes a nuclear phosphatidylinositol 5-phosphatase (PIP5Pase), its substrate PI(3,4,5)P3, and the repressor-activator protein 1 (RAP1). RAP1 binds to ES sequences flanking VSG genes via its DNA binding domains and represses VSG transcription. In contrast, PI(3,4,5)P3 binds to the N-terminus of RAP1 and controls its DNA binding activity. Transient inactivation of PIP5Pase results in the accumulation of nuclear PI(3,4,5)P3, which binds RAP1 and displaces it from ESs, activating transcription of silent ESs and VSG switching. The system is also required for the developmental silencing of VSG genes. The data provides a mechanism controlling reversible telomere silencing essential for the periodic switching in VSG expression and its developmental regulation. [ABSTRACT FROM AUTHOR]

The phosphoinositide 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) signaling pathway regulates proliferation, survival and metabolism, and its dysregulation is one of the most frequent oncogenic events across human malignancies. Over the last two decades, there has been significant focus on the clinical development of PI3K pathway inhibitors. More than 40 different inhibitors of this axis have reached various stages of clinical trials, but only a few of them have been approved by the Food and Drug Administration (FDA) for cancer treatment. These clinical results, however, could be improved given the importance of PI3K signaling in cancer and its role in linking cancer growth with metabolism. In this systematic review, after a glance at PI3K/AKT/mTOR pathway and its different inhibitors, we retrieved registered clinical trials evaluating the efficacy and safety of PI3K/AKT/mTOR inhibitors on Clinicaltrials.gov. Following the extraction of the data, finally we analyzed 2250 included studies in multiple steps, beginning with an overview and moving on to the details about type of malignancies, inhibitors, and treatment strategies. We also took a closer look at more than 100 phase III-IV clinical trials to pinpoint promising therapies, hoping that presenting a comprehensive picture of current clinical trials casts a flash of light on what remains to be done in future clinical trials of PI3K/AKT/mTOR inhibitors in human malignancies. [ABSTRACT FROM AUTHOR]

The phosphoinositide 3-kinase/protein kinase B/mammalian target of rapamycin (P13K/AKT/mTOR) pathway plays a key role in tuberculosis (TB) pathogenesis and infection. While the activity levels of this pathway during active infection are still debated, manipulating this pathway shows potential benefit for host-directed therapies. Some studies indicate that pathway inhibitors may have potential for TB treatment through upregulation of autophagy, while other studies do not encourage the use of these inhibitors due to possible host tissue destruction by Mycobacterium tuberculosis (M. tb) and increased infection risk. Investigating further clinical trials and their use of pathway inhibitors is necessary in order to ascertain their potential for TB treatment. This paper is particularly focused on the drug everolimus, an mTOR inhibitor. One of the first clinical trials sponsored by the Aurum Institute showed potential benefit in using everolimus as an adjunctive therapy for tuberculosis. Infection with tuberculosis is associated with a metabolic shift from oxidative phosphorylation towards glycolysis. The everolimus arm in the clinical trial showed further reduction than the control for both maximal and peak glycolytic activity. Compared with control, those receiving everolimus demonstrated increased lung function through forced expiratory volume in 1 s (FEV1) measurements, suggesting that everolimus may mitigate inflammation contributing to lung damage. [ABSTRACT FROM AUTHOR]

Background: The endoplasmic reticulum (ER)-mitochondria membrane contact sites (MCS) are extensively studied in aerobic eukaryotes; however, little is known about MCS in anaerobes with reduced forms of mitochondria named hydrogenosomes. In several eukaryotic lineages, the direct physical tether between ER and the outer mitochondrial membrane is formed by ER-mitochondria encounter structure (ERMES). The complex consists of four core proteins (Mmm1, Mmm2, Mdm12, and Mdm10) which are involved in phospholipid trafficking. Here we investigated ERMES distribution in organisms bearing hydrogenosomes and employed Trichomonas vaginalis as a model to estimate ERMES cellular localization, structure, and function. Results: Homology searches revealed that Parabasalia-Anaeramoebae, anaerobic jakobids, and anaerobic fungi are lineages with hydrogenosomes that retain ERMES, while ERMES components were gradually lost in Fornicata, and are absent in Preaxostyla and Archamoebae. In T. vaginalis and other parabasalids, three ERMES components were found with the expansion of Mmm1. Immunofluorescence microscopy confirmed that Mmm1 localized in ER, while Mdm12 and Mmm2 were partially localized in hydrogenosomes. Pull-down assays and mass spectrometry of the ERMES components identified a parabasalid-specific Porin2 as a substitute for the Mdm10. ERMES modeling predicted a formation of a continuous hydrophobic tunnel of TvMmm1-TvMdm12-TvMmm2 that is anchored via Porin2 to the hydrogenosomal outer membrane. Phospholipid-ERMES docking and Mdm12-phospholipid dot-blot indicated that ERMES is involved in the transport of phosphatidylinositol phosphates. The absence of enzymes involved in hydrogenosomal phospholipid metabolism implies that ERMES is not involved in the exchange of substrates between ER and hydrogenosomes but in the unidirectional import of phospholipids into hydrogenosomal membranes. Conclusions: Our investigation demonstrated that ERMES mediates ER-hydrogenosome interactions in parabasalid T. vaginalis, while the complex was lost in several other lineages with hydrogenosomes. [ABSTRACT FROM AUTHOR]

Background: Tyrosine kinase and phosphoinositide kinase pathways play important roles in asthma formation. As a dual tyrosine and phosphoinositide kinase inhibitor, PP121 has shown anticancer efficacy in multiple tumors. However, the study of PP121 in pulmonary diseases is still limited. Herein, we investigated the therapeutic activities of PP121 in asthma treatment. Methods: Tension measurements and patch clamp recordings were made to investigate the anticontractile characteristics of PP121 in vitro. Then, an asthma mouse model was established to further explore the therapeutic characteristics of PP121 via measurement of respiratory system resistance, histological analysis and western blotting. Results: We discovered that PP121 could relax precontracted mouse tracheal rings (mTRs) by blocking certain ion channels, including L-type voltage-dependent Ca2+ channels (L-VDCCs), nonselective cation channels (NSCCs), transient receptor potential channels (TRPCs), Na+/Ca2+ exchangers (NCXs) and K+ channels, and accelerating calcium mobilization. Furthermore, PP121 relieved asthmatic pathological features, including airway hyperresponsiveness, systematic inflammation and mucus secretion, via downregulation of inflammatory factors, mucins and the mitogen-activated protein kinase (MAPK)/Akt signaling pathway in asthmatic mice. Conclusion: In summary, PP121 exerts dual anti-contractile and anti-inflammatory effects in asthma treatment, which suggests that PP121 might be a promising therapeutic compound and shed new light on asthma therapy. Keypoints: PP121 has anticontractile properties in precontracted mTRs. PP121 inhibits calcium mobilization by switching certain ion channels. PP121 relieves typical pathological signs in an asthma mouse model. PP121 downregulates inflammatory factors, mucins and MAPK/Akt signaling. [ABSTRACT FROM AUTHOR]

The purpose of this study was to identify proteins associated with differences in the freezing tolerance of sheep sperm and to analyze their functions. Qualified fresh semen from four breeds of rams, the Australian White, white-head Dorper, Black-head Dorper, and Hu sheep breeds, were selected for cryopreservation. The sperm freezing tolerance was investigated by evaluation of the overall vitality, progressive vitality, and rapidly advance vitality of the sperm. A differential model of sperm freezing tolerance was constructed for sheep breeds showing significant differences. Differentially expressed proteins associated with sperm freezing tolerance were identified using iTRAQ and the protein functions were analyzed. It was found that sperm freezing tolerance was best in Hu sheep and worst in white-head Dorper sheep. These two breeds were used for the construction of a model based on differences in freezing tolerance and the identification of sperm proteins expressed differentially before freezing and after thawing. A total of 128 differentially expressed proteins (88 up-regulated and 40 down-regulated) were identified before freezing and after thawing in Hu sheep sperm (fresh/frozen Hu sheep sperm referred to as HL vs. HF), while 219 differentially expressed proteins (106 up-regulated and 113 down-regulated) were identified in white-head Dorper sheep (fresh/frozen white-head Dorper sheep sperm referred to as WL vs. WF). A comparison of these differentially expressed proteins showed that 57 proteins overlapped between the two breeds while 71 were only expressed in Hu sheep and 162 were only expressed in white-head Dorper sheep. Functional annotation and enrichment analyses of differentially expressed proteins down-regulated in Hu sheep involved in phosphorylation of phosphatidylinositol phosphate kinases, regulation of GTPase activity and glycolysis/gluconeogenesis signaling pathway. Up-regulated proteins of Hu sheep participated in oxidoreductase activity and oxidative phosphorylation process of sperm freezing. Furthermore, down-regulated in white-head Dorper sheep involved in the metabolic regulation of carbohydrate and nuclear sugar metabolism. Up-regulated proteins of white-head Dorper sheep involved in the ferroptosis and oxidative phosphorylation pathways. Collectively, These proteins were found to participate mainly in oxidative phosphorylation as well as phosphorylation and metabolic processes in the mitochondria to affect the freezing tolerance of sheep sperm. • The purpose of this study was to identify proteins associated with differences in freezing tolerance of sheep sperm and analyze their functions. • A differential model of sperm freezing tolerance was constructed for sheep breeds showing significant differences. • It was found that sperm freezing tolerance was best in Hu sheep and worst in white-head Dorper sheep. • Differentially expressed proteins associated with sperm freezing tolerance were identified using iTRAQ and proteins functions were analyzed. [ABSTRACT FROM AUTHOR]

Background: Many gaps remain in our understanding of the immune and molecular characteristics that underlie activated phosphoinositide 3‐kinase delta syndrome (APDS). Methods: We performed RNA sequencing of peripheral blood leukocytes obtained from a child with APDS and his healthy parents and deconvoluted bulk transcriptional data to assess immune cell status. Results: Pathway enrichment analysis suggested signaling pathways enriched in virus infection as well as the PI3K, mitogen‐activated protein kinase (MAPK), natural killer cell‐mediated cytotoxicity, and nucleotide‐binding oligomerization domain (NOD)‐like receptor signaling pathways. The proportion of B cells memory, T cells CD4 memory resting and dendritic cells activated were reduced, whereas B cells naïve, T cells CD8, NK cells resting, monocytes and macrophages M2 were increased in the child. Top 10 hub genes were screened and showed moderate to strong relatedness with immune cell proportions. Conclusion: Deconvolution of bulk RNA sequencing to assess immune cells status can provide further insight into the alterations in immunological features underlying APDS and other rare diseases. [ABSTRACT FROM AUTHOR]