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MedComm Oct 2023Tissue-type plasminogen activator (tPA) encoded by is a major mediator that promotes fibrinolysis and prevents thrombosis. Pathogenetic mutations in associated with...
Tissue-type plasminogen activator (tPA) encoded by is a major mediator that promotes fibrinolysis and prevents thrombosis. Pathogenetic mutations in associated with venous thromboembolism have rarely been reported. Here, we report the first case of a homozygous point mutation c.1411T>C (p.Y471H) in leading to thromboembolic events and conduct related functional studies. The corresponding tPA mutant protein (tPA-Y471H) and wild-type tPA (tPA-WT) were synthesized in vitro, and mutant mice ( mice) were constructed. The molecular docking and surface plasmon resonance results indicated that the mutation impeded the hydrogen-bonding interactions between the protease domain of tPA and the kringle 4 domain of plasminogen, and the binding affinity of tPA and plasminogen was significantly reduced with a difference of one order of magnitude. mRNA half-life assay showed that the half-life of tPA-Y471H was shortened. The inferior vena cava thrombosis model showed that the rate of venous thrombosis in mice was 80% compared with 53% in wild-type mice. Our data suggested a novel role for the protease domain of tPA in efficient plasminogen activation, and demonstrated that this tPA mutation could reduce the fibrinolysis function of the body and lead to an increased propensity for thrombosis.
PubMed: 37808270
DOI: 10.1002/mco2.392 -
Redox Biology Sep 2023The production of reactive oxygen species (ROS) by NADPH oxidase (NOX) 2 has been linked to both insulin resistance and exercise training adaptations in skeletal muscle....
The production of reactive oxygen species (ROS) by NADPH oxidase (NOX) 2 has been linked to both insulin resistance and exercise training adaptations in skeletal muscle. This study explores the previously unexamined role of NOX2 in the interplay between diet-induced insulin resistance and exercise training (ET). Using a mouse model that harbors a point mutation in the essential NOX2 regulatory subunit, p47phox (Ncf1*), we investigated the impact of this mutation on various metabolic adaptations. Wild-type (WT) and Ncf1* mice were assigned to three groups: chow diet, 60% energy fat diet (HFD), and HFD with access to running wheels (HFD + E). After a 16-week intervention, a comprehensive phenotypic assessment was performed, including body composition, glucose tolerance, energy intake, muscle insulin signaling, redox-related proteins, and mitochondrial adaptations. The results revealed that NOX2 deficiency exacerbated the impact of HFD on body weight, body composition, and glucose intolerance. Moreover, in Ncf1* mice, ET did not improve glucose tolerance or increase muscle cross-sectional area. ET normalized body fat independently of genotype. The lack of NOX2 activity during ET reduced several metabolic adaptations in skeletal muscle, including insulin signaling and expression of Hexokinase II and oxidative phosphorylation complexes. In conclusion, these findings suggest that NOX2 mediates key beneficial effects of exercise training in the context of diet-induced obesity.
Topics: Animals; Mice; Insulin Resistance; Diet, High-Fat; Obesity; Muscle, Skeletal; Insulin; Glucose; Mice, Inbred C57BL
PubMed: 37572454
DOI: 10.1016/j.redox.2023.102842 -
Cell Reports Nov 2023TmaR, the only known pole-localizer protein in Escherichia coli, was shown to cluster at the cell poles and control localization and activity of the major sugar...
TmaR, the only known pole-localizer protein in Escherichia coli, was shown to cluster at the cell poles and control localization and activity of the major sugar regulator in a tyrosine phosphorylation-dependent manner. Here, we show that TmaR assembles by phase separation (PS) via heterotypic interactions with RNA in vivo and in vitro. An unbiased automated mutant screen combined with directed mutagenesis and genetic manipulations uncovered the importance of a predicted nucleic-acid-binding domain, a disordered region, and charged patches, one containing the phosphorylated tyrosine, for TmaR condensation. We demonstrate that, by protecting flagella-related transcripts, TmaR controls flagella production and, thus, cell motility and biofilm formation. These results connect PS in bacteria to survival and provide an explanation for the linkage between metabolism and motility. Intriguingly, a point mutation or increase in its cellular concentration induces irreversible liquid-to-solid transition of TmaR, similar to human disease-causing proteins, which affects cell morphology and division.
Topics: Humans; Biomolecular Condensates; Bacteria; Proteins; Flagella; Escherichia coli; Tyrosine
PubMed: 37934665
DOI: 10.1016/j.celrep.2023.113393 -
Proceedings of the National Academy of... Oct 2023The nuclear cleavage of a suboptimal primary miRNA hairpin by the Drosha/DGCR8 complex ("Microprocessor") can be enhanced by an optimal miRNA neighbor, a phenomenon...
The nuclear cleavage of a suboptimal primary miRNA hairpin by the Drosha/DGCR8 complex ("Microprocessor") can be enhanced by an optimal miRNA neighbor, a phenomenon termed cluster assistance. Several features and biological impacts of this new layer of miRNA regulation are not fully known. Here, we elucidate the parameters of cluster assistance of a suboptimal miRNA and also reveal competitive interactions amongst optimal miRNAs within a cluster. We exploit cluster assistance as a functional assay for suboptimal processing and use this to invalidate putative suboptimal substrates, as well as identify a "solo" suboptimal miRNA. Finally, we report complexity in how specific mutations might affect the biogenesis of clustered miRNAs in disease contexts. This includes how an operon context can buffer the effect of a deleterious processing variant, but reciprocally how a point mutation can have a nonautonomous effect to impair the biogenesis of a clustered, suboptimal, neighbor. These data expand our knowledge regarding regulated miRNA biogenesis in humans and represent a functional assay for empirical definition of suboptimal Microprocessor substrates.
Topics: Humans; MicroRNAs; RNA Processing, Post-Transcriptional; RNA-Binding Proteins; Ribonuclease III
PubMed: 37788316
DOI: 10.1073/pnas.2306727120 -
Bioinformatics (Oxford, England) Nov 2023The two strands of the DNA double helix locally and spontaneously separate and recombine in living cells due to the inherent thermal DNA motion. This dynamics results in...
MOTIVATION
The two strands of the DNA double helix locally and spontaneously separate and recombine in living cells due to the inherent thermal DNA motion. This dynamics results in transient openings in the double helix and is referred to as "DNA breathing" or "DNA bubbles." The propensity to form local transient openings is important in a wide range of biological processes, such as transcription, replication, and transcription factors binding. However, the modeling and computer simulation of these phenomena, have remained a challenge due to the complex interplay of numerous factors, such as, temperature, salt content, DNA sequence, hydrogen bonding, base stacking, and others.
RESULTS
We present pyDNA-EPBD, a parallel software implementation of the Extended Peyrard-Bishop-Dauxois (EPBD) nonlinear DNA model that allows us to describe some features of DNA dynamics in detail. The pyDNA-EPBD generates genomic scale profiles of average base-pair openings, base flipping probability, DNA bubble probability, and calculations of the characteristically dynamic length indicating the number of base pairs statistically significantly affected by a single point mutation using the Markov Chain Monte Carlo algorithm.
AVAILABILITY AND IMPLEMENTATION
pyDNA-EPBD is supported across most operating systems and is freely available at https://github.com/lanl/pyDNA_EPBD. Extensive documentation can be found at https://lanl.github.io/pyDNA_EPBD/.
Topics: Computer Simulation; Models, Chemical; DNA; Software; Base Pairing; Nucleic Acid Conformation
PubMed: 37991847
DOI: 10.1093/bioinformatics/btad699 -
Cell Reports Dec 2023Heterotrimeric G proteins transduce extracellular chemical messages to generate appropriate intracellular responses. Point mutations in GNAO1, encoding the G protein α...
Heterotrimeric G proteins transduce extracellular chemical messages to generate appropriate intracellular responses. Point mutations in GNAO1, encoding the G protein α subunit, have been implicated in a pathogenic condition characterized by seizures, movement disorders, intellectual disability, and developmental delay (GNAO1 disorder). However, the effects of these mutations on G protein structure and function are unclear. Here, we report the effects of 55 mutations on Gα conformation, thermostability, nucleotide binding, and hydrolysis, as well as interaction with Gβγ subunits, receptors, and effectors. Our effort reveals four functionally distinct groups of mutants, including one group that sequesters receptors and another that sequesters Gβγ, both acting in a genetically dominant manner. These findings provide a more comprehensive understanding of disease-relevant mutations and reveal that GNAO1 disorder is likely composed of multiple mechanistically distinct disorders that will likely require multiple therapeutic strategies.
Topics: Humans; Mutation; Movement Disorders; Point Mutation; GTP-Binding Proteins; GTP-Binding Protein alpha Subunits, Gi-Go
PubMed: 37980565
DOI: 10.1016/j.celrep.2023.113462 -
Frontiers in Bioscience (Landmark... Oct 2023In the past 10 years, significant progress has been made in understanding the pathogenic chain of events that causes Alzheimer's disease (AD). According to the most...
BACKGROUND
In the past 10 years, significant progress has been made in understanding the pathogenic chain of events that causes Alzheimer's disease (AD). According to the most widely accepted concept, the production and aggregation of β-amyloid (Aβ) peptides play a critical role in AD. As a result, therapeutic intervention with these processes is the focus of intense research. The Aβ peptide is cleaved by the α-secretase, β-secretase, and γ-secretase enzymes in a region near the pathogenic amyloid precursor protein (APP) and mutations occurring site.
METHODS
In the current review, a complete picture of the risk factors behind AD has been investigated. Mutations involved in AD progression have also been screened in various studies.
RESULTS
Most of the mutations in the amyloid precursor protein (APP) can lead to the accumulation of APP oligomers in the brain, leading to AD. Several point mutations in APP can cause familial AD (FAD), including the Swedish mutation (K>M670/671N>L) and the A673>V mutation. The pathogenic A673>V mutation and Swedish mutation (M670>K/N671>L) are present in the same region of amyloid precursor protein (). However, the A673>T mutation has been shown to confer protection against AD.
CONCLUSION
More investigations are needed from geographically distinct regions on mutations associated with AD development and applications of nanomedicines for better management of the disease burden in the future. Nanotechnology-produced metal nanoparticles (NPs) have gotten much attention because of their wide range of uses in the medicinal and agricultural industries. Nanomedicine containing potential phytochemicals, including GX-50 and curcumin conjugated with NPs, maybe a potential candidate for treating AD.
Topics: Humans; Alzheimer Disease; Amyloid beta-Protein Precursor; Amyloid beta-Peptides; Mutation; Amyloid Precursor Protein Secretases
PubMed: 37919079
DOI: 10.31083/j.fbl2810258 -
Nature Communications Feb 2024The concept of precision cell therapy targeting tumor-specific mutations is appealing but requires surface-exposed neoepitopes, which is a rarity in cancer. B cell...
The concept of precision cell therapy targeting tumor-specific mutations is appealing but requires surface-exposed neoepitopes, which is a rarity in cancer. B cell receptors (BCR) of mature lymphoid malignancies are exceptional in that they harbor tumor-specific-stereotyped sequences in the form of point mutations that drive self-engagement of the BCR and autologous signaling. Here, we use a BCR light chain neoepitope defined by a characteristic point mutation (IGLV3-21) for selective targeting of a poor-risk subset of chronic lymphocytic leukemia (CLL) with chimeric antigen receptor (CAR) T cells. We develop murine and humanized CAR constructs expressed in T cells from healthy donors and CLL patients that eradicate IGLV3-21 expressing cell lines and primary CLL cells, but neither cells expressing the non-pathogenic IGLV3-21 light chain nor polyclonal healthy B cells. In vivo experiments confirm epitope-selective cytolysis in xenograft models in female mice using engrafted IGLV3-21 expressing cell lines or primary CLL cells. We further demonstrate in two humanized mouse models lack of cytotoxicity towards human B cells. These data provide the basis for advanced approaches of resistance-preventive and biomarker-guided cellular targeting of functionally relevant lymphoma driver mutations sparing normal B cells.
Topics: Humans; Female; Mice; Animals; Leukemia, Lymphocytic, Chronic, B-Cell; B-Lymphocytes; Mutation; Receptors, Antigen, B-Cell; T-Lymphocytes
PubMed: 38307904
DOI: 10.1038/s41467-024-45378-w -
The Journal of Cell Biology Nov 2023Astrocytes control the formation of specific synaptic circuits via cell adhesion and secreted molecules. Astrocyte synaptogenic functions are dependent on the...
Astrocytes control the formation of specific synaptic circuits via cell adhesion and secreted molecules. Astrocyte synaptogenic functions are dependent on the establishment of their complex morphology. However, it is unknown if distinct neuronal cues differentially regulate astrocyte morphogenesis. δ-Catenin was previously thought to be a neuron-specific protein that regulates dendrite morphology. We found δ-catenin is also highly expressed by astrocytes and required both in astrocytes and neurons for astrocyte morphogenesis. δ-Catenin is hypothesized to mediate transcellular interactions through the cadherin family of cell adhesion proteins. We used structural modeling and biochemical analyses to reveal that δ-catenin interacts with the N-cadherin juxtamembrane domain to promote N-cadherin surface expression. An autism-linked δ-catenin point mutation impaired N-cadherin cell surface expression and reduced astrocyte complexity. In the developing mouse cortex, only lower-layer cortical neurons express N-cadherin. Remarkably, when we silenced astrocytic N-cadherin throughout the cortex, only lower-layer astrocyte morphology was disrupted. These findings show that δ-catenin controls astrocyte-neuron cadherin interactions that regulate layer-specific astrocyte morphogenesis.
Topics: Animals; Mice; Astrocytes; Cadherins; Delta Catenin; Morphogenesis; Neurons
PubMed: 37707499
DOI: 10.1083/jcb.202303138 -
JCI Insight Sep 2023A T50I substitution in the K-Ras interswitch domain causes Noonan syndrome and emerged as a third-site mutation that restored the in vivo transforming activity and...
A T50I substitution in the K-Ras interswitch domain causes Noonan syndrome and emerged as a third-site mutation that restored the in vivo transforming activity and constitutive MAPK pathway activation by an attenuated KrasG12D,E37G oncogene in a mouse leukemia model. Biochemical and crystallographic data suggested that K-RasT50I increases MAPK signal output through a non-GTPase mechanism, potentially by promoting asymmetric Ras:Ras interactions between T50 and E162. We generated a "switchable" system in which K-Ras mutant proteins expressed at physiologic levels supplant the fms like tyrosine kinase 3 (FLT3) dependency of MOLM-13 leukemia cells lacking endogenous KRAS and used this system to interrogate single or compound G12D, T50I, D154Q, and E162L mutations. These studies support a key role for the asymmetric lateral assembly of K-Ras in a plasma membrane-distal orientation that promotes the formation of active Ras:Raf complexes in a membrane-proximal conformation. Disease-causing mutations such as T50I are a valuable starting point for illuminating normal Ras function, elucidating mechanisms of disease, and identifying potential therapeutic opportunities for Rasopathy disorders and cancer.
Topics: Animals; Mice; Disease Models, Animal; Germ Cells; Germ-Line Mutation; Leukemia; Mutation; Proto-Oncogene Proteins p21(ras); ras Proteins
PubMed: 37681415
DOI: 10.1172/jci.insight.168445