Nat Commun. 2023 Jun 8;14(1):3379. doi: 10.1038/s41467-023-39130-z.

ABSTRACT

In plant communities, diversity often increases productivity and functioning, but the specific underlying drivers are difficult to identify. Most ecological theories attribute positive diversity effects to complementary niches occupied by different species or genotypes. However, the specific nature of niche complementarity often remains unclear, including how it is expressed in terms of trait differences between plants. Here, we use a gene-centred approach to study positive diversity effects in mixtures of natural Arabidopsis thaliana genotypes. Using two orthogonal genetic mapping approaches, we find that between-plant allelic differences at the AtSUC8 locus are strongly associated with mixture overyielding. AtSUC8 encodes a proton-sucrose symporter and is expressed in root tissues. Genetic variation in AtSUC8 affects the biochemical activities of protein variants and natural variation at this locus is associated with different sensitivities of root growth to changes in substrate pH. We thus speculate that - in the particular case studied here - evolutionary divergence along an edaphic gradient resulted in the niche complementarity between genotypes that now drives overyielding in mixtures. Identifying genes important for ecosystem functioning may ultimately allow linking ecological processes to evolutionary drivers, help identify traits underlying positive diversity effects, and facilitate the development of high-performance crop variety mixtures.

PMID:37291153 | DOI:10.1038/s41467-023-39130-z

Nat Commun. 2023 Jun 8;14(1):3371. doi: 10.1038/s41467-023-38817-7.

ABSTRACT

In the Neurospora circadian system, the White Collar Complex (WCC) drives expression of the principal circadian negative arm component frequency (frq). FRQ interacts with FRH (FRQ-interacting RNA helicase) and CKI, forming a stable complex that represses its own expression by inhibiting WCC. In this study, a genetic screen identified a gene, designated as brd-8, that encodes a conserved auxiliary subunit of the NuA4 histone acetylation complex. Loss of brd-8 reduces H4 acetylation and RNA polymerase (Pol) II occupancy at frq and other known circadian genes, and leads to a long circadian period, delayed phase, and defective overt circadian output at some temperatures. In addition to strongly associating with the NuA4 histone acetyltransferase complex, BRD-8 is also found complexed with the transcription elongation regulator BYE-1. Expression of brd-8, bye-1, histone h2a.z, and several NuA4 subunits is controlled by the circadian clock, indicating that the molecular clock both regulates the basic chromatin status and is regulated by changes in chromatin. Taken together, our data identify auxiliary elements of the fungal NuA4 complex having homology to mammalian components, which along with conventional NuA4 subunits, are required for timely and dynamic frq expression and thereby a normal and persistent circadian rhythm.

PMID:37291101 | DOI:10.1038/s41467-023-38817-7

Semin Cell Dev Biol. 2023 Jun 6:S1084-9521(23)00114-3. doi: 10.1016/j.semcdb.2023.05.005. Online ahead of print.

NO ABSTRACT

PMID:37291029 | DOI:10.1016/j.semcdb.2023.05.005

Curr Biol. 2023 Jun 5:S0960-9822(23)00678-4. doi: 10.1016/j.cub.2023.05.046. Online ahead of print.

ABSTRACT

White campion (Silene latifolia, Caryophyllaceae) was the first vascular plant where sex chromosomes were discovered. This species is a classic model for studies on plant sex chromosomes due to presence of large, clearly distinguishable X and Y chromosomes that originated de novo about 11 million years ago (mya), but lack of genomic resources for this relatively large genome (∼2.8 Gb) remains a significant hurdle. Here we report S. latifolia female genome assembly integrated with sex-specific genetic maps of this species, focusing on sex chromosomes and their evolution. The analysis reveals a highly heterogeneous recombination landscape with strong reduction in recombination rate in the central parts of all chromosomes. Recombination on the X chromosome in female meiosis primarily occurs at the very ends, and over 85% of the X chromosome length is located in a massive (∼330 Mb) gene-poor, rarely recombining pericentromeric region (Xpr). The results indicate that the non-recombining region on the Y chromosome (NRY) initially evolved in a relatively small (∼15 Mb), actively recombining region at the end of the q-arm, possibly as a result of inversion on the nascent X chromosome. The NRY expanded about 6 mya via linkage between the Xpr and the sex-determining region, which may have been caused by expanding pericentromeric recombination suppression on the X chromosome. These findings shed light on the origin of sex chromosomes in S. latifolia and yield genomic resources to assist ongoing and future investigations into sex chromosome evolution.

PMID:37290443 | DOI:10.1016/j.cub.2023.05.046

Cell Chem Biol. 2023 Jun 6:S2451-9456(23)00150-2. doi: 10.1016/j.chembiol.2023.05.007. Online ahead of print.

ABSTRACT

Recurrent JAK2 alterations are observed in myeloproliferative neoplasms, B-cell acute lymphoblastic leukemia, and other hematologic malignancies. Currently available type I JAK2 inhibitors have limited activity in these diseases. Preclinical data support the improved efficacy of type II JAK2 inhibitors, which lock the kinase in the inactive conformation. By screening small molecule libraries, we identified a lead compound with JAK2 selectivity. We highlight analogs with on-target biochemical and cellular activity and demonstrate in vivo activity using a mouse model of polycythemia vera. We present a co-crystal structure that confirms the type II binding mode of our compounds with the "DFG-out" conformation of the JAK2 activation loop. Finally, we identify a JAK2 G993A mutation that confers resistance to the type II JAK2 inhibitor CHZ868 but not to our analogs. These data provide a template for identifying novel type II kinase inhibitors and inform further development of agents targeting JAK2 that overcome resistance.

PMID:37290440 | DOI:10.1016/j.chembiol.2023.05.007

Comput Biol Med. 2023 Jun 3;162:107068. doi: 10.1016/j.compbiomed.2023.107068. Online ahead of print.

ABSTRACT

Ubiquitin-specific protease 7 (USP7) is one of the most abundant deubiquitinases and plays an important role in various malignant tumors. However, the molecular mechanisms underlying USP7's structures, dynamics, and biological significance are yet to be investigated. In this study, we constructed the full-length models of USP7 in both the extended and compact state, and applied elastic network models (ENM), molecular dynamics (MD) simulations, perturbation response scanning (PRS) analysis, residue interaction networks as well as allosteric pocket prediction to investigate allosteric dynamics in USP7. Our analysis of intrinsic and conformational dynamics revealed that the structural transition between the two states is characterized by global clamp motions, during which the catalytic domain (CD) and UBL4-5 domain exhibit strong negative correlations. The PRS analysis, combined with the analysis of disease mutations and post-translational modifications (PTMs) further highlighted the allosteric potential of the two domains. The residue interaction network based on MD simulations captured an allosteric communication path which starts at CD domain and ends at UBL4-5 domain. Moreover, we identified a pocket at the TRAF-CD interface as a high-potential allosteric site for USP7. Overall, our studies not only provide molecular insights into the conformational changes of USP7, but also aid in the design of allosteric modulators that target USP7.

PMID:37290391 | DOI:10.1016/j.compbiomed.2023.107068

Biochem Biophys Res Commun. 2023 May 18;671:10-17. doi: 10.1016/j.bbrc.2023.05.071. Online ahead of print.

ABSTRACT

α-amylase plays a crucial role in regulating metabolism and health by hydrolyzing of starch and glycogen. Despite comprehensive studies of this classic enzyme spanning over a century, the function of its carboxyl terminal domain (CTD) with a conserved eight β-strands is still not fully understood. Amy63, identified from a marine bacterium, was reported as a novel multifunctional enzyme with amylase, agarase and carrageenase activities. In this study, the crystal structure of Amy63 was determined at 1.8 Å resolution, revealing high conservation with some other amylases. Interestingly, the independent amylase activity of the carboxyl terminal domain of Amy63 (Amy63_CTD) was newly discovered by the plate-based assay and mass spectrometry. To date, the Amy63_CTD alone could be regarded as the smallest amylase subunit. Moreover, the significant amylase activity of Amy63_CTD was measured over a wide range of temperature and pH, with optimal activity at 60 °C and pH 7.5. The Small-angle X-ray scattering (SAXS) data showed that the high-order oligomeric assembly gradually formed with increasing concentration of Amy63_CTD, implying the novel catalytic mechanism as revealed by the assembly structure. Therefore, the discovery of the novel independent amylase activity of Amy63_CTD suggests a possible missing step or a new perspective in the complex catalytic process of Amy63 and other related α-amylases. This work may shed light on the design of nanozymes to process marine polysaccharides efficiently.

PMID:37290279 | DOI:10.1016/j.bbrc.2023.05.071

PLoS Genet. 2023 Jun 8;19(6):e1010759. doi: 10.1371/journal.pgen.1010759. Online ahead of print.

ABSTRACT

Gene regulation is highly cell type-specific and understanding the function of non-coding genetic variants associated with complex traits requires molecular phenotyping at cell type resolution. In this study we performed single nucleus ATAC-seq (snATAC-seq) and genotyping in peripheral blood mononuclear cells from 13 individuals. Clustering chromatin accessibility profiles of 96,002 total nuclei identified 17 immune cell types and sub-types. We mapped chromatin accessibility QTLs (caQTLs) in each immune cell type and sub-type using individuals of European ancestry which identified 6,901 caQTLs at FDR < .10 and 4,220 caQTLs at FDR < .05, including those obscured from assays of bulk tissue such as with divergent effects on different cell types. For 3,941 caQTLs we further annotated putative target genes of variant activity using single cell co-accessibility, and caQTL variants were significantly correlated with the accessibility level of linked gene promoters. We fine-mapped loci associated with 16 complex immune traits and identified immune cell caQTLs at 622 candidate causal variants, including those with cell type-specific effects. At the 6q15 locus associated with type 1 diabetes, in line with previous reports, variant rs72928038 was a naïve CD4+ T cell caQTL linked to BACH2 and we validated the allelic effects of this variant on regulatory activity in Jurkat T cells. These results highlight the utility of snATAC-seq for mapping genetic effects on accessible chromatin in specific cell types.

PMID:37289818 | DOI:10.1371/journal.pgen.1010759

AMB Express. 2023 Jun 8;13(1):55. doi: 10.1186/s13568-023-01558-3.

ABSTRACT

LukS-PV is a component of Panton-Valentine leucocidin (PVL) and is secreted by Staphylococcus aureus. Silver nanoparticles exhibit considerable potential as anticancer agents and drug delivery systems. Drug delivery is a way to deliver medicinal combinations to achieve a beneficial therapeutic effect. In the current study, recombinant LukS-PV protein-loaded silver nanoparticles were prepared and their cytotoxicity effect was analyzed on human breast cancer cells and human normal embryonic kidneys cells by MTT assay. Apoptosis was investigated by staining with Annexin V/propidium iodide. The recombinant LukS-PV protein-loaded silver nanoparticles showed dose-dependent cytotoxicity and induced apoptosis in the MCF7 cells and had a lesser effect on HEK293 cells. After 24 h exposure to the recombinant LukS-PV protein-loaded silver nanoparticles (IC50), Annexin V-FITC/PI FCM revealed that 33.2% of MCF7 cells were apoptotic. In conclusion, recombinant LukS-PV protein-loaded silver nanoparticles probably cannot be a better alternative for the targeted healing approaches to cancer therapies. Hence, it is suggested that silver nanoparticles could be utilized as a delivery system for releasing toxins into cancer cells.

PMID:37289339 | DOI:10.1186/s13568-023-01558-3

J Virol. 2023 Jun 8:e0040023. doi: 10.1128/jvi.00400-23. Online ahead of print.

ABSTRACT

Induction of type I interferon (IFN) gene expression is among the first lines of cellular defense a virus encounters during primary infection. We previously identified the tegument protein M35 of murine cytomegalovirus (MCMV) as an essential antagonist of this antiviral system, showing that M35 interferes with type I IFN induction downstream of pattern-recognition receptor (PRR) activation. Here, we report structural and mechanistic details of M35's function. Determination of M35's crystal structure combined with reverse genetics revealed that homodimerization is a key feature for M35's immunomodulatory activity. In electrophoretic mobility shift assays (EMSAs), purified M35 protein specifically bound to the regulatory DNA element that governs transcription of the first type I IFN gene induced in nonimmune cells, Ifnb1. DNA-binding sites of M35 overlapped with the recognition elements of interferon regulatory factor 3 (IRF3), a key transcription factor activated by PRR signaling. Chromatin immunoprecipitation (ChIP) showed reduced binding of IRF3 to the host Ifnb1 promoter in the presence of M35. We furthermore defined the IRF3-dependent and the type I IFN signaling-responsive genes in murine fibroblasts by RNA sequencing of metabolically labeled transcripts (SLAM-seq) and assessed M35's global effect on gene expression. Stable expression of M35 broadly influenced the transcriptome in untreated cells and specifically downregulated basal expression of IRF3-dependent genes. During MCMV infection, M35 impaired expression of IRF3-responsive genes aside of Ifnb1. Our results suggest that M35-DNA binding directly antagonizes gene induction mediated by IRF3 and impairs the antiviral response more broadly than formerly recognized. IMPORTANCE Replication of the ubiquitous human cytomegalovirus (HCMV) in healthy individuals mostly goes unnoticed but can impair fetal development or cause life-threatening symptoms in immunosuppressed or -deficient patients. Like other herpesviruses, CMV extensively manipulates its hosts and establishes lifelong latent infections. Murine CMV (MCMV) presents an important model system as it allows the study of CMV infection in the host organism. We previously showed that during entry into host cells, MCMV virions release the evolutionary conserved protein M35 protein to immediately dampen the antiviral type I interferon (IFN) response induced by pathogen detection. Here, we show that M35 dimers bind to regulatory DNA elements and interfere with recruitment of interferon regulatory factor 3 (IRF3), a key cellular factor for antiviral gene expression. Thereby, M35 interferes with expression of type I IFNs and other IRF3-dependent genes, reflecting the importance for herpesviruses to avoid IRF3-mediated gene induction.

PMID:37289084 | DOI:10.1128/jvi.00400-23

Elife. 2023 Jun 8;12:e83393. doi: 10.7554/eLife.83393. Online ahead of print.

ABSTRACT

Globular bushy cells (GBCs) of the cochlear nucleus play central roles in the temporal processing of sound. Despite investigation over many decades, fundamental questions remain about their dendrite structure, afferent innervation, and integration of synaptic inputs. Here, we use volume electron microscopy (EM) of the mouse cochlear nucleus to construct synaptic maps that precisely specify convergence ratios and synaptic weights for auditory- nerve innervation and accurate surface areas of all postsynaptic compartments. Detailed biophysically-based compartmental models can help develop hypotheses regarding how GBCs integrate inputs to yield their recorded responses to sound. We established a pipeline to export a precise reconstruction of auditory nerve axons and their endbulb terminals together with high-resolution dendrite, soma, and axon reconstructions into biophysically-detailed compartmental models that could be activated by a standard cochlear transduction model. With these constraints, the models predict auditory nerve input profiles whereby all endbulbs onto a GBC are subthreshold (coincidence detection mode), or one or two inputs are suprathreshold (mixed mode). The models also predict the relative importance of dendrite geometry, soma size, and axon initial segment length in setting action potential threshold and generating heterogeneity in sound-evoked responses, and thereby propose mechanisms by which GBCs may homeostatically adjust their excitability. Volume EM also reveals new dendritic structures and dendrites that lack innervation. This framework defines a pathway from subcellular morphology to synaptic connectivity, and facilitates investigation into the roles of specific cellular features in sound encoding. We also clarify the need for new experimental measurements to provide missing cellular parameters, and predict responses to sound for further in vivo studies, thereby serving as a template for investigation of other neuron classes.

PMID:37288824 | DOI:10.7554/eLife.83393

J Biomol Struct Dyn. 2023 Jun 8:1-14. doi: 10.1080/07391102.2023.2220812. Online ahead of print.

ABSTRACT

Chlamydia trachomatis (C. trachomatis) is an obligate intracellular bacterium which causes eye and sexually transmitted infections. During pregnancy, the bacterium is associated with preterm complications, low weight of neonates, fetal demise and endometritis leading to infertility. The aim of our study was design of a multi-epitope vaccine (MEV) candidate against C. trachomatis. After protein sequence adoption from the NCBI, potential epitopes toxicity, antigenicity, allergenicity, MHC-I and MHC-II binding, cytotoxic T lymphocytes (CTLs), Helper T lymphocytes (HTLs) and interferon-γ (IFN-γ)- induction were predicted. The adopted epitopes were fused together using appropriate linkers. In the next step, the MEV structural mapping and characterization, three-dimensional (3D) structure homology modeling and refinement were also performed. The MEV candidate interaction with the toll-like receptor 4 (TLR4) was also docked. The immune responses simulation was assessed using the C-IMMSIM server. Molecular dynamic (MD) simulation verified the structural stability of the TLR4-MEV complex. The Molecular Mechanics Poisson-Boltzmann Surface Area (MMPBSA) approach demonstrated the MEV high affinity of binding to the TLR4, MHC-I and MHC-II. The MEV construct was also stable and water soluble and had enough antigenicity and lacked allergenicity with stimulation of T cells and B cells and INF-γ release. The immune simulation confirmed acceptable responses of both the humoral and cellular arms. It is proposed that in vitro and in vivo studies are needed to evaluate the findings of this study.Communicated by Ramaswamy H. Sarma.

PMID:37288800 | DOI:10.1080/07391102.2023.2220812

iScience. 2023 Apr 8;26(6):106584. doi: 10.1016/j.isci.2023.106584. eCollection 2023 Jun 16.

ABSTRACT

EGFR-TKIs were used in NSCLC patients with actionable EGFR mutations and prolong prognosis. However, most patients treated with EGFR-TKIs developed resistance within around one year. This suggests that residual EGFR-TKIs resistant cells may eventually lead to relapse. Predicting resistance risk in patients will facilitate individualized management. Herein, we built an EGFR-TKIs resistance prediction (R-index) model and validate in cell line, mice, and cohort. We found significantly higher R-index value in resistant cell lines, mice models and relapsed patients. Patients with an elevated R-index had significantly shorter relapse time. We also found that the glycolysis pathway and the KRAS upregulation pathway were related to EGFR-TKIs resistance. MDSC is a significant immunosuppression factor in the resistant microenvironment. Our model provides an executable method for assessing patient resistance status based on transcriptional reprogramming and may contribute to the clinical translation of patient individual management and the study of unclear resistance mechanisms.

PMID:37288343 | PMC:PMC10241979 | DOI:10.1016/j.isci.2023.106584

Plant Environ Interact. 2023 Feb 9;4(2):55-69. doi: 10.1002/pei3.10098. eCollection 2023 Apr.

ABSTRACT

As agricultural production is reaching its limits regarding outputs and land use, the need to further improve crop yield is greater than ever. The limited translatability from in vitro lab results into more natural growth conditions in soil remains problematic. Although considerable progress has been made in developing soil-growth assays to tackle this bottleneck, the majority of these assays use pots or whole trays, making them not only space- and resource-intensive, but also hampering the individual treatment of plants. Therefore, we developed a flexible and compact screening system named PhenoWell® in which individual seedlings are grown in wells filled with soil allowing single-plant treatments. The system makes use of an automated image-analysis pipeline that extracts multiple growth parameters from individual seedlings over time, including projected rosette area, relative growth rate, compactness, and stockiness. Macronutrient, hormone, salt, osmotic, and drought stress treatments were tested in the PhenoWell® system. The system is also optimized for maize with results that are consistent with Arabidopsis while different in amplitude. We conclude that the PhenoWell® system enables a high-throughput, precise, and uniform application of a small amount of solution to individually soil-grown plants, which increases the replicability and reduces variability and compound usage.

PMID:37288161 | PMC:PMC10243540 | DOI:10.1002/pei3.10098

Adv Biomed Res. 2023 Apr 28;12:113. doi: 10.4103/abr.abr_267_21. eCollection 2023.

ABSTRACT

BACKGROUND: Bacteriocins are a type of antimicrobial peptide that are produced by probiotics. They have been studied as possible therapeutic drugs and have been used to suppress bacterial development in foods. Nisin is a potent bacteriocin having the anti-microbial and anti-cancer characteristics produced by Lactococcus lactis. The aim of the present paper is to evaluate the influence of Nisin on cell adhesion and its two related genes, mmp-2 and mmp-9, in the colorectal cancer cell line.

MATERIALS AND METHODS: For this purpose, HT-29 cells were treated with various concentrations of Nisin and the cell cytotoxicity, cell adhesion, and gene expression were evaluated using the MTT assay, cell adhesion assay, and real-time PCR.

RESULTS: Our findings showed that 32 to 1024 μg/ml of Nisin resulted in a significant reduction in cell viability (P < 0.05). Furthermore, 128 and 256 μg/ml of Nisin significantly reduced the cell adhesion, and mmp-2 and mmp-9 gene expressions (P < 0.05).

CONCLUSION: Our findings suggested that Nisin could prevent metastasis and cancer progression.

PMID:37288013 | PMC:PMC10241620 | DOI:10.4103/abr.abr_267_21

Curr Opin Syst Biol. 2023 Jun;34:None. doi: 10.1016/j.coisb.2023.100455.

ABSTRACT

Engineering new functions in the microbiome requires understanding how host genetic control and microbe-microbe interactions shape the microbiome. One key genetic mechanism underlying host control is the immune system. The immune system can promote stability in the composition of the microbiome by reshaping the ecological dynamics of its members, but the degree of stability will depend on the interplay between ecological context, immune system development, and higher-order microbe-microbe interactions. The eco-evolutionary interplay affecting composition and stability should inform the strategies used to engineer new functions in the microbiome. We conclude with recent methodological developments that provide an important path forward for both engineering new functionality in the microbiome and broadly understanding how ecological interactions shape evolutionary processes in complex biological systems.

PMID:37287906 | PMC:PMC10242548 | DOI:10.1016/j.coisb.2023.100455

Biophys Rep. 2021 Aug 31;7(4):326-340. doi: 10.52601/bpr.2021.200045.

ABSTRACT

Microbiota-host interaction has attracted more and more attentions in recent years. The association between microbiota and host health is largely attributed to its influence on host immune system. Microbial-derived antigens and metabolites play a critical role in shaping the host immune system, including regulating its development, activation, and function. However, during various diseases the microbiota-host communication is frequently found to be disordered. In particular, gut microbiota dysbiosis associated with or led to the occurrence and progression of infectious diseases, autoimmune diseases, metabolic diseases, and neurological diseases. Pathogenic microbes and their metabolites disturb the protective function of immune system, and lead to disordered immune responses that usually correlate with disease exacerbation. In the other hand, the immune system also regulates microbiota composition to keep host homeostasis. Here, we will discuss the current advances of our knowledge about the interactions between microbiota and host immune system during health and diseases.

PMID:37287759 | PMC:PMC10233470 | DOI:10.52601/bpr.2021.200045

Front Plant Sci. 2023 May 23;14:1182867. doi: 10.3389/fpls.2023.1182867. eCollection 2023.

ABSTRACT

Groundnut productivity and quality have been impeded by rising temperatures in semi-arid environments. Hence, understanding the effects and molecular mechanisms of heat stress tolerance will aid in tackling yield losses. In this context, a recombinant inbred line (RIL) population was developed and phenotyped for eight seasons at three locations for agronomic, phenological, and physiological traits under heat stress. A genetic map was constructed using genotyping-by-sequencing with 478 single-nucleotide polymorphism (SNP) loci spanning a map distance of 1,961.39 cM. Quantitative trait locus (QTL) analysis using phenotypic and genotypic data identified 45 major main-effect QTLs for 21 traits. Intriguingly, three QTL clusters (Cluster-1-Ah03, Cluster-2-Ah12, and Cluster-3-Ah20) harbor more than half of the major QTLs (30/45, 66.6%) for various heat tolerant traits, explaining 10.4%-38.6%, 10.6%-44.6%, and 10.1%-49.5% of phenotypic variance, respectively. Furthermore, important candidate genes encoding DHHC-type zinc finger family protein (arahy.J0Y6Y5), peptide transporter 1 (arahy.8ZMT0C), pentatricopeptide repeat-containing protein (arahy.4A4JE9), Ulp1 protease family (arahy.X568GS), Kelch repeat F-box protein (arahy.I7X4PC), FRIGIDA-like protein (arahy.0C3V8Z), and post-illumination chlorophyll fluorescence increase (arahy.92ZGJC) were the underlying three QTL clusters. The putative functions of these genes suggested their involvement in seed development, regulating plant architecture, yield, genesis and growth of plants, flowering time regulation, and photosynthesis. Our results could provide a platform for further fine mapping, gene discovery, and developing markers for genomics-assisted breeding to develop heat-tolerant groundnut varieties.

PMID:37287715 | PMC:PMC10243373 | DOI:10.3389/fpls.2023.1182867

Front Cell Infect Microbiol. 2023 May 23;13:1130965. doi: 10.3389/fcimb.2023.1130965. eCollection 2023.

ABSTRACT

INTRODUCTION: Toxoplasma gondii induces a strong CD8 T cell response characterized by the secretion of IFNγ that promotes host survival during infection. The initiation of CD8 T cell IFNγ responses in vitro differs widely between clonal lineage strains of T. gondii, in which type I strains are low inducers, while types II and III strains are high inducers. We hypothesized this phenotype is due to a polymorphic "Regulator Of CD8 T cell Response" (ROCTR).

METHODS: Therefore, we screened F1 progeny from genetic crosses between the clonal lineage strains to identify ROCTR. Naïve antigen-specific CD8 T cells (T57) isolated from transnuclear mice, which are specific for the endogenous and vacuolar TGD057 antigen, were measured for their ability to become activated, transcribe Ifng and produce IFNγ in response to T. gondii infected macrophages.

RESULTS: Genetic mapping returned four non-interacting quantitative trait loci (QTL) with small effect on T. gondii chromosomes (chr) VIIb-VIII, X and XII. These loci encompass multiple gene candidates highlighted by ROP16 (chrVIIb-VIII), GRA35 (chrX), TgNSM (chrX), and a pair of uncharacterized NTPases (chrXII), whose locus we report to be significantly truncated in the type I RH background. Although none of the chromosome X and XII candidates bore evidence for regulating CD8 T cell IFNγ responses, type I variants of ROP16 lowered Ifng transcription early after T cell activation. During our search for ROCTR, we also noted the parasitophorous vacuole membrane (PVM) targeting factor for dense granules (GRAs), GRA43, repressed the response suggesting PVM-associated GRAs are important for CD8 T cell activation. Furthermore, RIPK3 expression in macrophages was an absolute requirement for CD8 T cell IFNγ differentiation implicating the necroptosis pathway in T cell immunity to T. gondii.

DISCUSSION: Collectively, our data suggest that while CD8 T cell IFNγ production to T. gondii strains vary dramatically, it is not controlled by a single polymorphism with strong effect. However, early in the differentiation process, polymorphisms in ROP16 can regulate commitment of responding CD8 T cells to IFNγ production which may have bearing on immunity to T. gondii.

PMID:37287466 | PMC:PMC10242045 | DOI:10.3389/fcimb.2023.1130965

Proteomics. 2023 Jun 7:e2300100. doi: 10.1002/pmic.202300100. Online ahead of print.

ABSTRACT

Increased throughput in proteomic experiments can improve accessibility of proteomic platforms, reduce costs, and facilitate new approaches in systems biology and biomedical research. Here we propose combination of analytical flow rate chromatography with ion mobility separation of peptide ions, data-independent acquisition, and data analysis with the DIA-NN software suite, to achieve high-quality proteomic experiments from limited sample amounts, at a throughput of up to 400 samples per day. For instance, when benchmarking our workflow using a 500-μL/min flow rate and 3-min chromatographic gradients, we report the quantification of 5211 proteins from 2 μg of a mammalian cell-line standard at high quantitative accuracy and precision. We further used this platform to analyze blood plasma samples from a cohort of COVID-19 inpatients, using a 3-min chromatographic gradient and alternating column regeneration on a dual pump system. The method delivered a comprehensive view of the COVID-19 plasma proteome, allowing classification of the patients according to disease severity and revealing plasma biomarker candidates.

PMID:37287406 | DOI:10.1002/pmic.202300100