Biology subjects

Liu, G.

Publications and source records attributed to Liu, G..

At least 181 records · Page 10Linked to original sources

bioRxiv2022

Large clones of pre-existing T cells drive early immunity against SARS-COV-2 and LCMV infection.

We analyzed the dynamics of the earliest T cell response to SARS-COV-2. A wave of TCRs strongly but transiently expand during infection, frequently peaking the same week as the first positive PCR test. These expanding TCR CDR3s were enriched for sequences functionally annotated as SARS-COV-2 specific. Most epitopes recognized by the expanding TCRs were highly conserved between SARS-COV-2 strains, but not with circulating human coronaviruses. Many expanding CDR3s were also present at high precursor frequency in pre-pandemic TCR repertoires. A similar set of early response TCRs specific for lymphocytic choriomeningitis virus epitopes were also found at high frequency in the pre-infection naive repertoire. High frequency naive precursors may allow the T cell response to respond rapidly during the crucial early phases of acute viral infection. One-Sentence SummaryHigh frequency naive precursors underly the rapid T cell response during the crucial early phases of acute viral infection.

immunology↗

bioRxiv2022

Membrane-remodeling protein ESCRT-III homologs incarnate the evolution and morphogenesis of multicellular magnetotactic bacteria

Endosomal sorting complex required transport (ESCRT) III proteins are essential for membrane remodeling and repair across all domains of life. Eukaryotic ESCRT-III and the cyanobacterial homologs PspA and Vipp1/Imm30 remodel membrane into vesicles, rings, filaments and tubular rods structures. Here our microscopy analysis showed that multicellular bacteria, referred to as magnetoglobules, possess multiple compartments including magnetosome organelles, polyphosphate granules, vesicles, rings, tubular rods, filaments and MVB-like structures. Therefore, membrane remodeling protein PspA might be required for the formation of these compartments, and contribute to the morphogenesis and evolution of multicellularity. To assess these hypotheses, we sequenced nine genomes of magnetoglobules and found a significant genome expansion compared to unicellular magnetotactic bacteria. Moreover, PspA was ubiquitous in magnetoglobules and formed a distinct clade on the tree of eubacterial and archaeal ESCRT-III. The phylogenetic feature suggested the evolution of magnetoglobules from a unicellular ancestor of deltaproteobacterium. Hetero-expression of ellipsoidal magnetoglobule pspA2 gene alone in Escherichia coli resulted in intracellular membrane aggregation. GFP fusion labeling revealed polar location of PspA2 in rod-shaped unicells and regular interval location in filamentous cells. Cryo-electron tomography analysis showed filament bundle, membrane sacculus, vesicles and MVB-like structure in the cells expressing PspA2. Moreover, electron-dense area with a similar distribution as GFP-PspA2 foci in filamentous cells changed the inward orientation of the septum, which might interfere with the cell division. Collectively, these results show the membrane remodeling function of magnetoglobule PspA proteins, which may contribute to morphogenesis and the evolution of multicellularity of magnetotactic bacteria.

microbiology↗

bioRxiv2022

Proliferative Exhausted CD8 T Cells Exacerbate Long-lasting Anti-tumor Effects in Human Papillomavirus Positive Head and Neck Squamous Cell Carcinoma

The survival prognosis of human papillomavirus (HPV)-positive and HPV-negative head and neck squamous cell carcinoma (HNSCC) is largely different, and little is known about the anti-tumor mechanism of tumor-infiltrated CD8+ exhausted T cells (Tex) in HNSCC. We performed cell level multi-omics sequencing on human HNSCC samples to decipher the multi-dimensional characteristics of Tex cells. A proliferative exhausted CD8+ T cell cluster (P-Tex) which was beneficial to survival outcomes of patients with HPV-positive HNSCC was identified. Interestingly, P-Tex cells expressed CDK4 genes as high as cancer cells, which could be simultaneously inhibited by CDK4 inhibitors and might be a potential reason for the ineffectiveness of CDK4 inhibitors in treating HPV-positive HNSCC. P-Tex cells could aggregate in the antigen presenting cell niches and activate certain signaling pathways. Together, our findings suggest a promising role for P-Tex cells in the prognosis of patients with HPV-positive HNSCC by providing modest but persistent anti-tumor effects. Graphic abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=105 SRC="FIGDIR/small/513993v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@1f8810forg.highwire.dtl.DTLVardef@1e11d84org.highwire.dtl.DTLVardef@58fb6aorg.highwire.dtl.DTLVardef@1e20a13_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2022

Development of an affinity-enhanced clinical candidate TCR targeting NY-ESO-1 with optimal potency and high specificity

The clinical success of T-cell receptor (TCR)-based immunotherapy depends on the efficacy and specificity of TCRs. Naturally occurring TCRs have limited anti-tumor potency due to their low affinity for tumor antigens. Affinity enhancement is a promising strategy to generate highly potent TCRs. However, it is concerned that affinity-enhanced TCRs are prone to lose specificity. We isolated low affinity TCRs specific for NY-ESO-1157-165/HLA-A*02:01 from peripheral blood mononuclear cells of healthy donors. An affinity-enhanced TCR candidate with optimal affinity and specificity was generated using phage display and an extensive set of in vitro and in vivo assays. Alanine scanning mutagenesis showed that the TCR candidate retained specificity by making extensive contacts to the side chains of NY-ESO-1157-165 peptide. Adoptive transfer of T cells engineered with this candidate (termed TAEST16001) significantly inhibited tumor growth in subcutaneous, metastatic, and patient-derived xenograft (PDX) mouse tumor models. This study demonstrates that sophisticated engineering and screening techniques can be utilized to generate a clinical candidate TCR with potent anti-tumor activity without losing specificity. TAEST16001 was approved by the Center for Drug Evaluation (CDE) as the first TCR-based immunotherapy clinical trial in China (ClinicalTrials.gov Identifier: NCT03159585).

immunology↗

bioRxiv2022

Role of spike in the pathogenic and antigenic behavior of SARS-CoV-2 BA.1Omicron

The recently identified, globally predominant SARS-CoV-2 Omicron variant (BA.1) is highly transmissible, even in fully vaccinated individuals, and causes attenuated disease compared with other major viral variants recognized to date1-7. The Omicron spike (S) protein, with an unusually large number of mutations, is considered the major driver of these phenotypes3,8. We generated chimeric recombinant SARS-CoV-2 encoding the S gene of Omicron in the backbone of an ancestral SARS-CoV-2 isolate and compared this virus with the naturally circulating Omicron variant. The Omicron S-bearing virus robustly escapes vaccine-induced humoral immunity, mainly due to mutations in the receptor-binding motif (RBM), yet unlike naturally occurring Omicron, efficiently replicates in cell lines and primary-like distal lung cells. In K18-hACE2 mice, while Omicron causes mild, non-fatal infection, the Omicron S-carrying virus inflicts severe disease with a mortality rate of 80%. This indicates that while the vaccine escape of Omicron is defined by mutations in S, major determinants of viral pathogenicity reside outside of S.

microbiology↗

bioRxiv2022

A pan-variant mRNA-LNP T cell vaccine protects HLA transgenic mice from mortality after infection with SARS-CoV-2 Beta

Clinically licensed COVID-19 vaccines ameliorate viral infection by inducing vaccinee production of neutralizing antibodies that bind to the SARS-CoV-2 Spike protein to inhibit viral cellular entry (Walsh et al., 2020; Baden et al., 2021), however the clinical effectiveness of these vaccines is transitory as viral variants arise that escape antibody neutralization (Tregoning et al., 2021; Willett et al., 2022). Vaccines that solely rely upon a T cell response to combat viral infection could be transformational because they can be based on highly conserved short peptide epitopes that hold the potential for pan-variant immunity, but a mRNA-LNP T cell vaccine has not been shown to be sufficient for effective antiviral prophylaxis. Here we show that a mRNA-LNP vaccine based on highly conserved short peptide epitopes activates a CD8+ and CD4+ T cell response that prevents mortality in HLA-A*02:01 transgenic mice infected with the SARS-CoV-2 Beta variant of concern (B.1.351). In mice vaccinated with the T cell vaccine, 24% of the nucleated cells in lung were CD8+ T cells on day 7 post infection. This was 5.5 times more CD8+ T cell infiltration of the lungs in response to infection compared to the Pfizer-BioNTech Comirnaty(R) vaccine. Between days 2 and 7 post infection, the number of CD8+ T cells in the lung increased in mice vaccinated with the T cell vaccine and decreased in mice vaccinated with Comirnaty(R). The T cell vaccine did not produce neutralizing antibodies, and thus our results demonstrate that SARS-CoV-2 viral infection can be controlled by a T cell response alone. Our results suggest that further study is merited for pan-variant T cell vaccines, and that T cell vaccines may be relevant for individuals that cannot produce neutralizing antibodies or to help mitigate Long COVID.

immunology↗

bioRxiv2022

Whole-genome sequencing and single acute toxicity of heavy metal to Photobacterium kishitanii FJ21

In this study, the growth morphology of FJ21 strain was observed, and its 16S rRNA and whole genome were sequenced. Then, related software was used to make genome assembly, gene structure and function annotation, genome phylogenetic tree analysis, genome collinearity analysis and prediction of secondary metabolic gene cluster analysis. Finally, the single acute toxicity of five heavy metals to FJ21 strain was detected. There were luxC, luxD, luxA, luxB, luxF, luxE and luxG genes in FJ21, and the protein encoded by lux operon had certain hydrophilicity. The genome of this strain FJ21 contains a chromosome with a total length of 4853277bp and a GC content of 39.23%. The genome of FJ21 was compared with that of Photobacterium kishitanii ATCCBAA-1194, Photobacterium phosphoreum JCM21184, Photobacterium aquimaris LC2-065, Photobacterium malacitanum CECT9190, and Photobacterium carnosum TMW 2.2021. The average nucleotide identity(ANI), tetra nucleotide signatures (Tetra), comparative genome, and phylogenetic analysis proposed that FJ21 is a strain of Photobacterium kishitanii. In the acute toxicity test, the toxicity of heavy metals to the strain FJ21 is Pb(NO3)2 > ZnSO4{middle dot}7H2O > CdCl2{middle dot}2.5H2O > CuSO4{middle dot}5H2O > K2Cr2O7.

genomics↗

bioRxiv2022

Optimization and redevelopment of single-cell data analysis workflow based on deep generative models

The present single-cell RNA sequencing(scRNA-seq) analysis pipelines require a combination of appropriate normalization, dimension reduction, clustering, and specific-gene analysis algorithms, but the rationale for the choice of these algorithms is relatively subjective because of the lack of ground truth assessment conclusions. As the number of captured single-cells increases, the number of different types of noise cells also increases, which can strongly affect the analysis efficiency. For scRNA-seq, a technology that generates data through multi-process operations, the deep generative model should be a good choice for this type of data analysis, allowing simultaneous estimation of multiple unobservable parameters assumed in the data generation process. Hence, in our study, we sequenced a pool of pre-labeled single cells to obtain a batch of scRNA-seq data with main and fine labels, which was then used to evaluate the clustering and specific-gene analysis methods. Afterward, we applied two deep generative models to infer the probabilities of pseudo and impurity cells. And by stepwise removing the inferred noise cells, the clustering performance and the consistency of different specific-gene analysis methods are both greatly improved. After that, we applied Deep-LDA (a latent Dirichlet allocation-based deep generative model) to scRNA-seq data analysis. And this model takes the count matrix as input, and makes the classification and specific gene optimization process mutually dependent, which has more practical sense and simplifies the analysis workflow. At last, we successfully implemented the model with transferred knowledge to make single-cell annotation and verified its superior performance.

bioinformatics↗

bioRxiv2022

Capillary electrophoresis mass spectrometry identifies new isomers of inositol pyrophosphates in mammalian tissues

Technical challenges have to date prevented a complete profiling of the levels of myo-inositol phosphates (InsPs) and pyrophosphates (PP-InsPs) in mammalian tissues. Here, we have deployed capillary electrophoresis mass spectrometry to identify and record the levels of InsPs and PP-InsPs in several tissues obtained from wild type mice and a newly-created PPIP5K2 knockout strain. We observe that the mouse colon harbours unusually high levels of InsPs and PP-InsPs. Additionally, the PP-InsP profile is considerably more complex than previously reported for animal cells: using chemically synthesized internal stable isotope references, and high-resolution mass spectra, we characterize two new PP-InsP isomers as 4/6-PP-InsP5 and 2-PP-InsP5. The latter has not previously been described in Nature. Analysis of feces and the commercial mouse diet suggest the latter is one potential source of noncanonical isomers in the colon. However, we also identify both molecules in the heart, indicating unknown synthesis pathways in mammals. We also demonstrate that the CE-MS method is sensitive enough to measure PP-InsPs from patient samples such as colon biopsies and peripheral blood mononuclear cells (PBMCs). Strikingly, PBMCs also contain 4/6-PP-InsP5 and 2-PP-InsP5. In summary, our study substantially expands PP-InsP biology in mammals.

biochemistry↗

bioRxiv2022

Efficient repair of human homozygous genetic mutation by CRISPR/Cas9 mediated interlocus gene conversion

DNA double-strand breaks (DSBs) induced by gene editing tools are primarily resolved either by non-homologous end joining (NHEJ) or homology-directed repair (HDR) using exogenous synthetic DNA templates. Repaired by error-prone NHEJ may lead to unexpected indels at the targeted site. In the case of most genetic disorders, HDR-mediated precise correction using an exogenous homologous sequence is ideal. However, the therapeutic application of HDR might be especially challenging given the requirement for the codelivery of exogenous DNA templates with toxicity into cells, and the low efficiency of HDR could also limit its clinical application. Here, we used hematopoietic stem cells (HSCs) with genetic mutations to cause {beta}-thalassemia in HBB coding regions and discovered that many cells are actually repaired by CRISPR/Cas9-mediated gene conversion (GC) independent of exogenous synthetic DNA templates. We show that pathogenic mutations in the HBB coding regions of HSCs can be repaired efficiently through CRISPR/GC using the paralog gene HBD as the internal template. Electroporations of Cas9 for ribonucleoprotein with sgRNA into haematopoietic stem and progenitor cells (HSPCs) with a variety of pathogenic gene mutations also resulted in effective conversion of mutations to normal wild-type sequences without exogenous DNA template. Moreover, the edited HSCs can repopulate the haematopoietic system and generate erythroid cells with a greatly reduced propensity for thalassemia after transplantations. Thus, CRISPR/GC, independent of exogenous DNA templates, holds great promise for gene therapy of genetic diseases.

genetics↗

bioRxiv2022

Stable isotopomers of myo-inositol to uncover the complex MINPP1-dependent inositol phosphate network

The water-soluble inositol phosphates (InsPs) represent a functionally diverse group of small-molecule messengers central to a myriad of cellular processes. However, we have an incomplete understanding of InsP metabolism because the available analytical toolset for inositol phosphates is rather limited. Here, we have synthesized and utilized fully and unsymmetrically 13C-labeled myo-inositol and inositol phosphates. These probes were applied in combination with nuclear magnetic resonance spectroscopy (NMR) and capillary electrophoresis mass spectrometry (CE-MS) to further annotate central aspects of InsP metabolism in human cells. The labeling strategy provided detailed structural information via NMR - down to individual enantiomers - which overcomes a crucial blind spot in the analysis of InsPs. We uncovered a novel branch of InsP dephosphorylation in human cells which is dependent on MINPP1, a phytase-like enzyme, that contributes to cellular homeostasis. Full characterization of MINPP1 activity in vitro and in cells, provided a clear picture of this multifunctional phosphatase. Metabolic labeling with stable isotopomers thus constitutes a powerful tool for investigating InsP networks in a variety of different biological contexts.

biochemistry↗

bioRxiv2022

Endothelial TIE1 restricts angiogenic sprouting to coordinate vein assembly in synergy with its homologue TIE2

ObjectiveVascular growth followed by vessel specification is crucial for the establishment of a hierarchical blood vascular network. We have here investigated mechanisms underlying venogenesis, particularly the molecular control over venous fate acquisition during vascular development. Approach and ResultsWe analyzed the function of TIE1 as well as its synergy with TIE2 in the regulation of vein formation by employing genetic mouse models targeting Tie1 and Tek. Cardinal vein growth appeared normal in TIE1 deficient mice, whereas TIE2 deficiency altered the identity of cardinal vein endothelial cells with the aberrant expression of DLL4. Interestingly, the parallel growth of murine cutaneous veins along with arteries, which was initiated at approximately embryonic day 13.5, was retarded in mice lack of TIE1. Tie1 deletion disrupted also venous integrity, displaying increased sprouting angiogenesis and vascular bleeding. Abnormal venous sprouts with defective arteriovenous alignment were also observed in the mesenteries of Tie1 deleted mice. Mechanistically, TIE1 deficiency resulted in the decreased expression of venous regulators including TIE2 while angiogenic regulators were upregulated. The alteration of TIE2 level by TIE1 insufficiency was further confirmed by the siRNA-mediated knockdown of Tie1 in cultured endothelial cells. Additionally, combining the endothelial deletion of Tie1 with one null allele of Tek resulted in a progressive increase of vein-associated angiogenesis leading to the formation of vascular tufts in retinas, whereas the loss of Tie1 alone produced only a relatively mild venous defect. ConclusionsFindings from this study imply that TIE1 and TIE2 act in a synergistic manner to restrict sprouting angiogenesis during vein formation.

developmental biology↗

bioRxiv2022

Characterization, Comparison, and Optimization of Lattice Light Sheets

Lattice light sheet microscopy excels at the non-invasive imaging of three-dimensional (3D) dynamic processes at high spatiotemporal resolution within cells and developing embryos. Recently, several papers have called into question the performance of lattice light sheets relative to the Gaussian sheets most common in light sheet microscopy. Here we undertake a comprehensive theoretical and experimental analysis of various forms of light sheet microscopy which both demonstrates and explains why lattice light sheets provide significant improvements in resolution and photobleaching reduction. The analysis provides a procedure to select the correct light sheet for a desired experiment and specifies the processing that maximizes the use of all fluorescence generated within the light sheet excitation envelope for optimal resolution while minimizing image artifacts and photodamage. Development of a new type of "harmonic balanced" lattice light sheet is shown to improve performance at all spatial frequencies within its 3D resolution limits and maintains this performance over lengthened propagation distances allowing for expanded fields of view. Significance StatementDespite its rapidly growing use, several misconceptions remain concerning the physics of image formation and its optimization in light sheet microscopy, particularly in high resolution variants tailored for subcellular imaging. These include the role of excitation sidelobes, the significance of out-of-focus fluorescence, the importance and optimization of deconvolution, and the perceived advantages of Gaussian beams. Here we attempt to shatter these misconceptions by showing that the professed tradeoffs between axial resolution and background haze, photobleaching rate, phototoxicity, and propensity for image artifacts do not exist for well-crafted lattice light sheets whose data is acquired and processed rigorously. The framework we provide should enable others to optimize light sheets and extract the most information at the lowest cost in their experiments.

cell biology↗

bioRxiv2022

Gene expression and RNA splicing explain large proportions of the heritability for complex traits in cattle

Many quantitative trait loci (QTL) are located in non-coding genomic regions. Therefore, QTL are assumed to affect gene regulation. Gene expression and RNA splicing are primary steps of transcription so QTL changing gene expression (eQTL) or RNA splicing (sQTL) are expected to significantly contribute to phenotypic variations. Here, we quantify the contribution of eQTL and sQTL detected from 16 tissues (N~5,000) to 37 complex traits of ~120k cattle. Using Bayesian methods, we show that including more regulatory variants in the model explains larger proportions of heritability. Across traits, cis and trans eQTL and sQTL detected from 16 tissues jointly explain ~70% (SE=0.5%) of heritability, 44% more than expected from the same number of random variants, where trans e/sQTL contribute 24% (14% more than expected). Multi-tissue cis and trans e/sQTL also explain 71% (SE=0.3%) of heritability for the metabolome, demonstrating the essential role of proximal and distal regulatory variants in shaping mammalian phenotypes.

genomics↗

bioRxiv2022

Impaired hepatic autophagy exacerbates xenobiotics induced liver injury

Xenobiotics can activate the hepatic survival pathway, but it is not clear how impaired hepatic survival pathways may affect xenobiotic-induced liver injury. We investigated the role of hepatic autophagy, a cellular survival pathway, in cholestatic liver injury driven by a xenobiotic. Here we demonstrate that DDC diet impaired autophagic flux, resulting in the accumulation of p62-Ub-intrahyaline bodies (IHBs) but not the Mallory Denk-Bodies (MDBs). Impaired autophagic flux was linked to a deregulated hepatic protein-chaperonin system and a significant decline in Rab family proteins. In addition, we demonstrate that heterozygous deletion of Atg7, a key autophagy gene, aggravated the p62-Ub-IHB accumulation and cholestatic liver injury. Moreover, we showed that p62-Ub-IHB accumulation did not activate the proteostasis-related ER stress signaling pathway, but rather activated the NRF2 pathway and suppressed the FXR nuclear receptor, resulting in cholestatic liver injury. Conclusion: Impaired autophagy exacerbates xenobiotic-induced cholestatic liver injury. Promotion of autophagy may represent a new therapeutic approach for xenobiotic-induced liver injury.

pharmacology and toxicology↗

bioRxiv2022

Two-Stage Multivariate Mendelian Randomization on Multiple Outcomes with Mixed Distributions

SO_SCPLOWUMMARYC_SCPLOWIn clinical research, it is of importance to study whether certain clinical factors or exposures have causal effects on clinical and patient reported outcomes like toxicities, quality of life, and self-reported symptoms, which can help improve patient care. Usually, such outcomes are recorded as multiple variables with different distributions. Mendelian randomization is a commonly used technique for causal inference with the help of genetic instrumental variables to deal with observed and unobserved confounders. Nevertheless, the current methodology of Mendelian randomization on multiple outcomes only focuses on one outcome at a time, meaning that it does not consider the correlation structure of multiple outcomes, which may lead to loss of statistical power. In situations with multiple outcomes of interest, especially when there are mixed correlated outcomes with multiple distributions, it is much more desirable to jointly analyze them with a multivariate approach. Some multivariate methods have been proposed to model mixed outcomes, however, they do not incorporate instrumental variables and cannot handle unmeasured confounders. To overcome the above challenges, we propose a two-stage multivariate Mendelian Randomization Method (MRMO), that can perform multivariate analysis on mixed outcomes using instrumental variables. We demonstrate that our proposed MRMO algorithm can gain power over the existing univariate method through simulation studies and a clinical application on a randomized Phase III clinical trial study on colorectal cancer patients.

genetics↗

bioRxiv2022

Within-host genotypic and phenotypic diversity of contemporaneous carbapenem-resistant Klebsiella pneumoniae from blood cultures of patients with bacteremia

Carbapenem-resistant Klebsiella pneumoniae (CRKP) are major pathogens globally. It is unknown whether bloodstream infections (BSIs) by CRKP and other bacteria are commonly caused by single organisms or mixed microbial populations. We hypothesized that contemporaneous CRKP from blood cultures of individual patients are genetically and phenotypically distinct. We determined short-read whole genome sequences of 10 strains from single colonies from CRKP-positive blood cultures in each of 6 patients (Illumina HiSeq). All strains were sequence type (ST)-258 K. pneumoniae that were unique by core genome single nucleotide polymorphism phylogeny, antibiotic resistance and virulence genes, capsular polysaccharide (CPS) gene mutations, and/or plasmid loss. Strains from each of 3 patients that differed in antibiotic resistance, virulence and/or CPS gene content underwent long-read sequencing for genome completion (Oxford Nanopore), and were tested for phenotypes in vitro and pathogenicity during mouse BSIs. Genetically distinct strains within individual patients exhibited significant differences in carbapenem, beta-lactam/beta-lactamase inhibitor and other antibiotic responses, CPS production, mucoviscosity, and susceptibility to serum killing. In 2 patients, strains differed significantly in their ability to infect organs and cause mortality in mice. In conclusion, we identified genotypic and phenotypic variant ST258 K. pneumoniae strains from blood cultures of individual patients, which were not detected by the clinical laboratory at time of BSI diagnosis. The data support a new paradigm of CRKP population diversity during BSIs. If validated for other BSIs, within-host bacterial diversity may have profound implications for medical, microbiology laboratory and infection prevention practices, and for understanding emergence of antibiotic resistance and pathogenesis. IMPORTANCEIn processing positive microbiologic cultures, standard clinical laboratory practice is to test a single bacterial strain from each morphologically distinct colony. We performed comprehensive whole genome sequence analyses on 10 carbapenem-resistant Klebsiella pneumoniae (CRKP) strains from positive blood cultures from each of 6 patients. Our findings that all strains were genetically unique and that genetic variants manifested differences in phenotypes like antibiotic responsiveness and virulence suggest that CRKP bloodstream infections may be commonly caused by mixed bacterial populations. Results raise questions about laboratory protocols and treatment decisions that are directed against a single strain. The observation that pan-genome analyses revealed inter-strain differences that were not evident by studying core genomes has important implications for investigating nosocomial outbreaks and transmission. Data also suggest a model of pathogenesis of CRKP infections, in which environmental pressures in vivo may select for outgrowth of variants that manifest antibiotic resistance, tolerance or specific virulence attributes.

microbiology↗

bioRxiv2022

Dendritic cell Piezo1 integrating mechanical stiffness and inflammatory signals directs the differentiation of TH1 and Treg cells in cancer

Dendritic cells (DCs) play an important role in anti-tumor immunity by inducing T cell differentiation. Herein, we found that the mechanical sensor Piezo1 expressed by DCs integrates innate inflammatory stimuli and stiffness signals and directs the reciprocal differentiation of TH1 and regulatory T (Treg) cells in cancer. Genetic deletion of Piezo1 in DCs inhibited the generation of TH1 cells while driving the development of Treg cells in promoting cancer growth. Mechanistically, Piezo1-deficient DCs regulated the secretion of the polarizing cytokines TGF{beta}1 and IL-12, leading to increased TGF{beta}R2-p-Smad3 activity and decreased IL-12R{beta}2-p-STAT4 activity while inducing the reciprocal differentiation of Treg and TH1 cells. In addition, Piezo1 integrated the SIRT1-hypoxia-inducible factor-1 alpha (HIF1)-dependent metabolic pathway and calcium-calcineurin-NFAT signaling pathway to orchestrate reciprocal TH1 and Treg lineage commitment through DC-derived IL-12 and TGF{beta}1. Our studies provide critical insight for understanding the role of the DC-based mechanical regulation of immunopathology in directing T cell lineage commitment in tumor microenvironments.

immunology↗