Regulation Of Cell Signaling Research Articles

The transcriptional landscape of plant infection by the rice blast fungus Magnaporthe oryzae reveals distinct families of temporally co-regulated and structurally conserved effectors.

The rice blast fungus Magnaporthe oryzae causes a devastating disease that threatens global rice (Oryza sativa) production. Despite intense study, the biology of plant tissue invasion during blast disease remains poorly understood. Here we report a high-resolution transcriptional profiling study of the entire plant-associated development of the blast fungus. Our analysis revealed major temporal changes in fungal gene expression during plant infection. Pathogen gene expression could be classified into 10 modules of temporally co-expressed genes, providing evidence for the induction of pronounced shifts in primary and secondary metabolism, cell signaling, and transcriptional regulation. A set of 863 genes encoding secreted proteins are differentially expressed at specific stages of infection, and 546 genes named MEP (Magnaportheeffector protein) genes were predicted to encode effectors. Computational prediction of structurally related MEPs, including the MAX effector family, revealed their temporal co-regulation in the same co-expression modules. We characterized 32 MEP genes and demonstrate that Mep effectors are predominantly targeted to the cytoplasm of rice cells via the biotrophic interfacial complex and use a common unconventional secretory pathway. Taken together, our study reveals major changes in gene expression associated with blast disease and identifies a diverse repertoire of effectors critical for successful infection.

Activity-based Tools for Interrogating Host Biology During Infection.

Host cells sense and respond to pathogens by dynamically regulating cell signaling. The rapid modulation of signaling pathways is achieved by post-translational modifications (PTMs) that can alter protein structure, function, and/or binding interactions. By using chemical probes to broadly profile changes in enzyme function or side-chain reactivity, activity-based protein profiling (ABPP) can reveal PTMs that regulate host-microbe interactions. While ABPP has been widely utilized to uncover microbial mechanisms of pathogenesis, in this review, we focus on more recent applications of this technique to the discovery of host PTMs and enzymes that modulate signaling within infected cells. Collectively, these advances underscore the importance of ABPP as a tool for interrogating the host response to infection and identifying potential targets for host-directed therapies.

Heterogeneity of B cell lymphopoiesis in patients with premalignant and active myeloma

To better characterize the heterogeneity of multiple myeloma (MM), we profiled plasma cells (PCs) and their B cell lymphopoiesis in the BM samples from patients with monoclonal gammopathy of undetermined significance, smoldering MM, and active MM by mass cytometry (CyTOF) analysis. Characterization of intra- and interneoplastic heterogeneity of malignant plasmablasts and PCs revealed overexpression of the MM SET domain (MMSET), Notch-1, and CD47. Variations in upregulation of B cell signaling regulators (IFN regulatory factor 4 [IRF-4], CXCR4, B cell lymphoma 6 [Bcl-6], c-Myc, myeloid differentiation primary response protein 88 [MYD88], and spliced X box-binding protein 1 [sXBP-1]) and aberrant markers (CD319, CD269, CD200, CD117, CD56, and CD28) were associated with different clinical outcomes in clonal PC subsets. In addition, prognosis was related to heterogeneity in subclonal expression of stemness markers, including neuroepithelial stem cell protein (Nestin), SRY-box transcription factor 2 (Sox2), Krüppel-like factor 4 (KLF-4), and Nanog. Furthermore, we have defined significantly elevated levels of MMSET, MYD88, c-Myc, CD243, Notch-1, and CD47 from hematopoietic stem cells to PCs in myeloma B cell lymphopoiesis, noted even in premalignant conditions, with variably modulated expression of B cell development regulators, including IRF-4, Bcl-2, Bcl-6, and sXBP-1; aberrant PC markers (such as CD52, CD44, CD200, CD81, CD269, CD117, and CXCR4); and stemness-controlling regulators, including Nanog, KLF-4, octamer-binding transcription factor 3/4 (Oct3/4), Sox2, and retinoic acid receptor α2 (RARα2). This study provides the rationale for precise molecular profiling of patients with MM by CyTOF technology to define disease heterogeneity and prognosis.

Cumulus Cell Transcriptome after Cumulus-Oocyte Complex Exposure to Nanomolar Cadmium in an In Vitro Animal Model of Prepubertal and Adult Age.

Cadmium (Cd), a highly toxic pollutant, impairs oocyte fertilization, through oxidative damage on cumulus cells (CCs). This study analysed the transcriptomic profile of CCs of cumulus-oocyte complexes (COCs) from adult and prepubertal sheep, exposed to Cd nanomolar concentration during in vitro maturation. In both age-groups, CCs of matured oocytes underwent RNA-seq, data analysis and validation. Differentially expressed genes (DEGs) were identified in adult (n = 99 DEGs) and prepubertal (n = 18 DEGs) CCs upon Cd exposure. Transcriptomes of adult CCs clustered separately between Cd-exposed and control samples, whereas prepubertal ones did not as observed by Principal Component Analysis. The transcriptomic signature of Cd-induced CC toxicity was identified by gene annotation and literature search. Genes associated with previous studies on ovarian functions and/or Cd effects were confirmed and new genes were identified, thus implementing the knowledge on their involvement in such processes. Enrichment and validation analysis showed that, in adult CCs, Cd acted as endocrine disruptor on DEGs involved in hormone biosynthesis, cumulus expansion, regulation of cell signalling, growth and differentiation and oocyte maturation, whereas in prepubertal CCs, Cd affected DEGs involved in CC development and viability and CC-oocyte communications. In conclusion, these DEGs could be used as valuable non-invasive biomarkers for oocyte competence.

Structural basis for the recruitment of PARP4 and NAD+to the vault complex.

Vault is a massive ribonucleoprotein complex that contains 78 copies of the major vault protein (MVP), which oligomerize into a barrel-like cage. This cage envelops several constitutively-bound minor vault components as well as transiently interacting “cargo” molecules. Vault mediates key biological functions including cell metabolism, innate immunity, and the cell's responses to disparate stressors. Previous studies suggest that vault may contribute to these processes by regulating cell signaling or guiding the intracellular localization of potential cargo, but its molecular-level mechanism remains unclear. In this study, we shed light on vault's functional mechanism by using cryo-EM to resolve the structure of human MVP in complex with the understudied minor vault component poly (ADP)-ribose polymerase 4 (PARP4) and PARP4's small-molecule substrate, nicotinamide adenine dinucleotide (NAD+). PARP4 is an ADP-ribosyltransferase that relies on available NAD+ to deposit ADP-ribose moieties onto macromolecular targets. It therefore has the potential to direct vault-mediated signaling activity, modify its cargo, or alter the vault cage architecture. Our sub-3.5 Å structures are the first to characterize MVP and PARP4 binding at the atomic level, and they also reveal a surprising interaction between NAD+ and a highly conserved region of MVP. Intriguingly, the presence of NAD+ causes the N-terminal region of PARP4 to undergo a substantial conformational change within the vault cage, drawing it into proximity with MVP's NAD+ binding pocket. Experiments are underway to determine the strength of the interaction between MVP and NAD+, MVP's specificity for the metabolite versus other nucleotides, and the extent to which the interaction between MVP and NAD+ impacts PARP4 activity.

Effects of a key negatively charged residue mutation on the binding pathway of SH3 domain complex and ArkA peptide.

SH3 domains are common protein interaction domains found across all forms of life and serve important functions in cell signaling and cytoskeletal regulation. SH3 domains often bind to intrinsically disordered peptides (IDPs) which are peptides that lack a well-defined secondary structure. However, little is known about the binding mechanism in these complexes. Prior to this experiment, our lab has simulated the disordered ArkA peptide binding to the yeast SH3 domain AbpSH3, and it was observed that the binding proceeds through a flexible disordered encounter complex before reaching a fully bound state. While the encounter complex forms quickly, the slow step of binding is the transition from the disordered encounter ensemble to fully engaged state. We mutated glutamic acid to glutamine due to its interaction with the ArkA lysine (−3) in the bound state, a residue that is located at the center of 12-residue ArkA peptide and is required for binding. In this experiment, we used molecular dynamics simulations to simulate the binding between AbpSH3, which has a net negative charge, and ArkA, which has a net positive charge. We analyzed the effect a single residue mutation has on the transitioning of the ArkA peptide and AbpSH3 domain from the unbound stage to the disordered encounter complex and finally to the fully bound complex. We found that the mutated domain forms similar contacts with ArkA in both the encounter complex and bound state compared to the wild type, despite the disruption of the favorable interaction between glutamic acid and lysine. For future research, we will examine the effects of mutating the ArkA peptide on both the encounter complex and the fully bound state.

The Curcumin and Gingerol Combination as an Immune Regulator and Anti-Inflammatory Agent of SARS-CoV Infection According to a Nutrigenomic Approach: A Mini-Review

Introduction: The COVID-19 pathophysiology caused by SARS-Cov-2 is closely related to immunoregulation and the process of inflammation. There are therapeutic targets in both, which are ideal for the healing process of infected patients. Phytonutrients are closely related to nutrigenomics. Curcumin and gingerol are two types of phytonutrients that have been studied, researched, and developed as therapeutic agents for diseases. Objective: This study aimed to examine the potential of curcumin and gingerol as immune regulators and anti-inflammatory agents in SARS-CoV-2 infections using a nutrigenomic approach. Methods: The literature review method was used in this study. Relevant information was gathered from scientific engines and databases (Google Scholar, Elsevier, Science Direct, Scopus, Wiley Online Library, PubMed) published during 2010-2021, and the data were analyzed by deductive qualitative descriptive technique. Result and Discussion: Curcumin in turmeric and gingerol in ginger have the potential to be used as a therapy for COVID-19 as they could be immune regulators and anti-inflammatory agents for SARS-CoV-2 infection. Curcumin and gingerol can act as primary and secondary antioxidants that can activate endogenous antioxidant enzymes, regulate cell signaling related to immunity such as interferons, nuclear factor-kappa beta, nitric oxide, and tumor necrosis factor-alpha, as well as stimulate anti-inflammatory and pro-inflammatory cytokine homeostasis, especially interleukins (IL-1β, IL-6, IL-17, IL-8). In silico, these two compounds were also proven to have potential as SARS-CoV-2 antivirals by acting as viral protease inhibitors. Conlclusion: The combination of curcumin and gingerol showed synergistic activity with increasing antioxidant and anti-inflammatory capacities. Thus, it has great potential for use in COVID-19 therapy.

De novo design of BMP mimics.

In traditional structure-guided drug designs, scientists often design drugs to target the active site of proteins of interest with detailed knowledge about the active site. However, despite the success rate, targeting the active site often leads to off-target side effects. With the development of protein binder design in the Baker Lab, one may imagine rewiring this drug development process by using de novo designed binders to regulate cell signaling pathways. Here I will present my efforts toward designing agonists/antagonists targeting different cell signaling pathways. I will focus on our efforts in developing bone morphogenetic protein 2/9/10 (BMP2/9/10) mimics, which are a group of growth factors to constitute a group of morphogenetic and angiogenetic signals. Recombinant human BMPs are used in orthopedics including oral surgery and spinal surgery. This project aims to specifically activate/inhibit BMP signaling pathways of interest by designing protein binders targeting specific BMP receptors, therefore, providing new therapeutics.

FcεRI clusters coupling with signaling condensates and membrane raft domains in mast cell activation.

T cell receptor (TCR) binding to foreign peptides presented by MHC (major histocompatibility complex) triggers T cell activation. Our recent work shows that this T cell activation is promoted by coupling membrane domains with protein condensates containing T cell signal transduction proteins LAT, Grb2, and Sos1. To test whether these observations are generalizable to other cellular systems, we evaluated the coupling of protein condensates to membrane domains in mast cells. Mast cells share many components in common with T cell activation, specifically, LAT, Grb2, and Sos1 are required for mast cell activation downstream of FcεRI clustering induced by multivalent antigens binding IgE. First, using giant plasma membrane vesicles (GPMVs) derived from a model mast cell line (RBL-2H3), we found that FcεRI partitions into the liquid disorder phase prior to antigen stimulation. The addition of a model antigen crosslinks the IgE-FcεRI and changes its partitioning, leading to enrichment in the relatively ordered (lipid raft) phase. Thus, antigen-induced clustering induces FcεRI raft affinity. Concurrently, TIRF imaging showed the formation of LAT+Grb2 condensates after RBL activation, which colocalized with activated FcεRI. These condensates recruited distinct membrane environments, demonstrating the potential coupling of raft domains with FcεRI and protein condensates. Our results indicate that raft coupling with signaling condensates may be a general mechanism for immune cell signaling regulation.

Targeted Quantification of Protein Phosphorylation and Its Contributions towards Mathematical Modeling of Signaling Pathways.

Post-translational modifications (PTMs) are key regulatory mechanisms that can control protein function. Of these, phosphorylation is the most common and widely studied. Because of its importance in regulating cell signaling, precise and accurate measurements of protein phosphorylation across wide dynamic ranges are crucial to understanding how signaling pathways function. Although immunological assays are commonly used to detect phosphoproteins, their lack of sensitivity, specificity, and selectivity often make them unreliable for quantitative measurements of complex biological samples. Recent advances in Mass Spectrometry (MS)-based targeted proteomics have made it a more useful approach than immunoassays for studying the dynamics of protein phosphorylation. Selected reaction monitoring (SRM)-also known as multiple reaction monitoring (MRM)-and parallel reaction monitoring (PRM) can quantify relative and absolute abundances of protein phosphorylation in multiplexed fashions targeting specific pathways. In addition, the refinement of these tools by enrichment and fractionation strategies has improved measurement of phosphorylation of low-abundance proteins. The quantitative data generated are particularly useful for building and parameterizing mathematical models of complex phospho-signaling pathways. Potentially, these models can provide a framework for linking analytical measurements of clinical samples to better diagnosis and treatment of disease.

The Role of Membrane-Type 1 Matrix Metalloproteinase-Substrate Interactions in Pathogenesis.

A protease is an enzyme with a proteolytic activity that facilitates the digestion of its substrates. Membrane-type I matrix metalloproteinase (MT1-MMP), a member of the broader matrix metalloproteinases (MMP) family, is involved in the regulation of diverse cellular activities. MT1-MMP is a very well-known enzyme as an activator of pro-MMP-2 and two collagenases, MMP-8 and MMP-13, all of which are essential for cell migration. As an anchored membrane enzyme, MT1-MMP has the ability to interact with a diverse group of molecules, including proteins that are not part of the extracellular matrix (ECM). Therefore, MT1-MMP can regulate various cellular activities not only by changing the extra-cellular environment but also by regulating cell signaling. The presence of both intracellular and extra-cellular portions of MT1-MMP can allow it to interact with proteins on both sides of the cell membrane. Here, we reviewed the MT1-MMP substrates involved in disease pathogenesis.

Matrix metalloproteinases as therapeutic targets in breast cancer.

Matrix metalloproteinases (MMPs) are the most prominent proteinases involved in tumorigenesis. They were initially recognized to promote tumor progression by remodeling the extracellular matrix through their proteolytic activity. However, accumulating evidence has revealed that some MMPs have protective roles in cancer progression, and the same MMP can exert opposing roles depending on the cell type in which it is expressed or the stage of cancer. Moreover, studies have shown that MMPs are involved in cancer progression through their roles in other biological processes such as cell signaling and immune regulation, independent of their catalytic activity. Despite the prognostic significance of tumoral or stromal expression of MMPs in breast cancer, their roles and molecular mechanisms in breast cancer progression remain unclear. As the failures of early clinical trials with broad-spectrum MMP inhibitors were mainly due to a lack of drug specificity, substantial efforts have been made to develop highly selective MMP inhibitors. Some recently developed MMP inhibitory monoclonal antibodies demonstrated promising anti-tumor effects in preclinical models of breast cancer. Importantly, anti-tumor effects of these antibodies were associated with the modulation of tumor immune microenvironment, suggesting that the use of MMP inhibitors in combination with immunotherapy can improve the efficacy of immunotherapy in HER2-positive or triple-negative breast cancer. In this review, the current understanding of the roles of tumoral or stromal MMPs in breast cancer is summarized, and recent advances in the development of highly selective MMP inhibitors are discussed.

SF3B4 promotes Twist1 expression and clear cell renal cell carcinoma progression by facilitating the export of KLF 16 mRNA from the nucleus to the cytoplasm

Splicing factor 3B subunit 4 (SF3B4) plays important functional roles not only in pre-mRNA splicing, but also in the regulation of transcription, translation, and cell signaling, and its dysregulation contributes to various diseases including Nager syndrome and tumorigenesis. However, the role of SF3B4 and underlying mechanisms in clear cell renal cell carcinoma (ccRCC) remain obscure. In the present study, we found that the expression of SF3B4 was significantly elevated in ccRCC tissues and negatively correlated with the overall survival of ccRCC patients. Upregulation of SF3B4 promotes migration and invasion of ccRCC cells in vitro and in vivo. The promoting effect of SF3B4 on cell migration and invasion is mediated by Twist1, a key transcription factor to mediate EMT. Interestingly, SF3B4, a component of the pre-mRNA spliceosome, is able to promote KLF16 expression by facilitating the transport of KLF16 mRNA into the cytoplasm. Mechanistically, SF3B4 promotes the export of KLF16 mRNA from the nucleus to the cytoplasm and thus enhances KLF16 expression, and in turn elevated KLF16 directly binds to the Twist1 promoter to activate its transcription, leading to EMT and ccRCC progression. Our findings provide evidence that the SF3B4-KLF16-Twist1 axis plays important functional roles in the development and progression of ccRCC, and manipulating this pathway may be a novel therapeutic target for the treatment of ccRCC.

Free Radicals Mediated Redox Signaling in Plant Stress Tolerance.

Abiotic and biotic stresses negatively affect plant cellular and biological processes, limiting their growth and productivity. Plants respond to these environmental cues and biotrophic attackers by activating intricate metabolic-molecular signaling networks precisely and coordinately. One of the initial signaling networks activated is involved in the generation of reactive oxygen species (ROS), reactive nitrogen species (RNS), and reactive sulfur species (RSS). Recent research has exemplified that ROS below the threshold level can stimulate plant survival by modulating redox homeostasis and regulating various genes of the stress defense pathway. In contrast, RNS regulates the stress tolerance potential of crop plants by modulating post-translation modification processes, such as S-nitrosation and tyrosine nitration, improving the stability of protein and DNA and activating the expression of downstream stress-responsive genes. RSS has recently emerged as a new warrior in combating plant stress-induced oxidative damage by modulating various physiological and stress-related processes. Several recent findings have corroborated the existence of intertwined signaling of ROS/RNS/RSS, playing a substantial role in crop stress management. However, the molecular mechanisms underlying their remarkable effect are still unknown. This review comprehensively describes recent ROS/RNS/RSS biology advancements and how they can modulate cell signaling and gene regulation for abiotic stress management in crop plants. Further, the review summarizes the latest information on how these ROS/RNS/RSS signaling interacts with other plant growth regulators and modulates essential plant functions, particularly photosynthesis, cell growth, and apoptosis.

Flavonoids as Modulators of Potassium Channels.

Potassium channels are widely distributed integral proteins responsible for the effective and selective transport of K+ ions through the biological membranes. According to the existing structural and mechanistic differences, they are divided into several groups. All of them are considered important molecular drug targets due to their physiological roles, including the regulation of membrane potential or cell signaling. One of the recent trends in molecular pharmacology is the evaluation of the therapeutic potential of natural compounds and their derivatives, which can exhibit high specificity and effectiveness. Among the pharmaceuticals of plant origin, which are potassium channel modulators, flavonoids appear as a powerful group of biologically active substances. It is caused by their well-documented anti-oxidative, anti-inflammatory, anti-mutagenic, anti-carcinogenic, and antidiabetic effects on human health. Here, we focus on presenting the current state of knowledge about the possibilities of modulation of particular types of potassium channels by different flavonoids. Additionally, the biological meaning of the flavonoid-mediated changes in the activity of K+ channels will be outlined. Finally, novel promising directions for further research in this area will be proposed.

Study of Ubiquitin Pathway Genes in a French Population with Amyotrophic Lateral Sclerosis: Focus on HECW1 Encoding the E3 Ligase NEDL1.

The ubiquitin pathway, one of the main actors regulating cell signaling processes and cellular protein homeostasis, is directly involved in the pathophysiology of amyotrophic lateral sclerosis (ALS). We first analyzed, by a next-generation sequencing (NGS) strategy, a series of genes of the ubiquitin pathway in two cohorts of familial and sporadic ALS patients comprising 176 ALS patients. We identified several pathogenic variants in different genes of this ubiquitin pathway already described in ALS, such as FUS, CCNF and UBQLN2. Other variants of interest were discovered in new genes studied in this disease, in particular in the HECW1 gene. We have shown that the HECT E3 ligase called NEDL1, encoded by the HECW1 gene, is expressed in neurons, mainly in their somas. Its overexpression is associated with increased cell death in vitro and, very interestingly, with the cytoplasmic mislocalization of TDP-43, a major protein involved in ALS. These results give new support for the role of the ubiquitin pathway in ALS, and suggest further studies of the HECW1 gene and its protein NEDL1 in the pathophysiology of ALS.

The Role of Inflammation in the Pathogenesis of Psoriasis

"Psoriasis is a chronic inflammatory skin condition with genetic determinism characterized by the presence of welldefined, erythematous plaques, covered by white, pearly, stratified scales, located on the extension areas, the skin of the scalp, intertriginous regions. The origin of psoriasis is multifactorial, involving hereditary and environmental pathogenic mechanisms. It is triggered by various risk factors involving a variety of processes, such as inflammation, antigen presentation, cell signaling, and transcriptional regulation. "

Sickle red blood cell-derived extracellular vesicles activate endothelial cells and enhance sickle red cell adhesion mediated by von Willebrand factor.

Endothelial activation and sickle red blood cell (RBC) adhesion are central to the pathogenesis of sickle cell disease (SCD). Quantitatively, RBC-derived extracellular vesicles (REVs) are more abundant from SS RBCs compared with healthy RBCs (AA RBCs). Sickle RBC-derived REVs (SS REVs) are known to promote endothelial cell (EC) activation through cell signalling and transcriptional regulation at longer terms. However, the SS REV-mediated short-term non-transcriptional response of EC is unclear. Here, we examined the impact of SS REVs on acute microvascular EC activation and RBC adhesion at 2 h. Compared with AA REVs, SS REVs promoted human pulmonary microvascular ECs (HPMEC) activation indicated by increased von Willebrand factor (VWF) expression. Under microfluidic conditions, we found abnormal SS RBC adhesion to HPMECs exposed to SS REVs. This enhanced SS RBC adhesion was reduced by haeme binding protein haemopexin or VWF cleaving protease ADAMTS13 to a level similar to HPMECs treated with AA REVs. Consistent with these observations, haemin- or SS REV-induced microvascular stasis in SS mice with implanted dorsal skin-fold chambers that was inhibited by ADAMTS13. The adhesion induced by SS REVs was variable and was higher with SS RBCs from patients with increased markers of haemolysis (lactate dehydrogenase and reticulocyte count) or a concomitant clinical diagnosis of deep vein thrombosis. Our results emphasise the critical contribution made by REVs to the pathophysiology of SCD by triggering acute microvascular EC activation and abnormal RBC adhesion. These findings may help to better understand acute pathophysiological mechanism of SCD and thereby the development of new treatment strategies using VWF as a potential target.

Spatial Chemoproteomics for Mapping the Active Proteome.

Functional regulation of cell signaling through dynamic changes in protein activity state as well as spatial organization represent two dynamic, complex, and conserved phenomena in biology. Seemingly separate areas of -omics method development have focused on building tools that can detect and quantify protein activity states, as well as map sub-cellular and intercellular protein organization. Integration of these efforts, through the development of chemical tools and platforms that enable detection and quantification of protein functional states with spatial resolution provide opportunities to better understand heterogeneity in the proteome within cell organelles, multi-cellular tissues, and whole organisms. This review provides an overview of and considerations for major classes of chemical proteomic probes and technologies that enable protein activity mapping from sub-cellular compartments to live animals.

The rational design of ARUK2007145, a dual inhibitor of the α and γ isoforms of the lipid kinase phosphatidylinositol 5-phosphate 4-kinase (PI5P4K).

The phosphatidylinositol 5-phosphate 4-kinases (PI5P4Ks) are therapeutic targets for diseases such as cancer, neurodegeneration and immunological disorders as they are key components in regulating cell signalling pathways. In an effort to make probe molecules available for further exploring these targets, we have previously reported PI5P4Kα-selective and PI5P4Kγ-selective ligands. Herein we report the rational design of PI5P4Kα/γ dual inhibitors, using knowledge gained during the development of selective inhibitors for these proteins. ARUK2007145 (39) is disclosed as a potent, cell-active probe molecule with ADMET properties amenable to conducting experiments in cells.