ABSTRACT
Hexi Corridor is one of the most important base of vegetable producing areas in China. Livestock manure (LM) applied to agricultural field could lead to soil heavy metal (HM) pollution. Previous studies have focused on HM pollution following LM application in acidic polluted soils; however, fewer studies have been conducted in alkaline unpolluted soils. A 4-year field vegetable production experiment was conducted using pig manure (PM) and chicken manure (CM) at five application rates (0, 15, 30, 45, and 60 t ha-1) to elucidate potential risks of HMs in an alkaline unpolluted soil in the Hexi Corridor oasis agricultural area and HM uptake by Chinese cabbage. The results showed that LM application caused a significant build-up of Cu, Zn, Pb, Cd, and Ni content in topsoil by 30.6-99.7%, 11.4-51.7%, 1.4-31.3%, 5.6-44.9%, 14%-40.8%, respectively. The Cd, Cu, Zn could potentially exceed the soil threshold in next 8-65 years after 15-60 t ha-1 LM application. Under LM treatment, the soil DTPA-extractable Cu, Zn, Fe, the acid-extractable fraction of Cu, Zn, Fe, Cd, Ni, and the Oxidable fraction of Cu, Zn, Fe, Mn, Cd, Ni significantly increased, but the DTPA-extractable Pb, Cd, the acid-extractable fraction of Pb, and the reducible fraction of Cd significantly decreased. Cu and Zn could migrate to the deeper soil and relatively increase in DTPA-extracted Cu, Zn were found in 20-40 cm soil depth after LM application. The pH and SOM could influence the bioavailability of HMs in soil. The bioaccumulation factor and transfer factor (TF) values were <1 except Mn (TF > 1). HMs in leaf did not approach the threshold for HM toxicity due to the "dilution effect". Recommend the type of manure was the PM and the annual PM application rate was 30 t ha-1 to ensure a 20-year period of clean production in alkaline unpolluted Fluvo-aqiuc vegetable soils.
Subject(s)
Brassica , Metals, Heavy , Soil Pollutants , Swine , Animals , Soil/chemistry , Manure/analysis , Livestock , Cadmium , Biological Availability , Lead , Soil Pollutants/analysis , Metals, Heavy/analysis , Vegetables , China , Acids , Pentetic AcidABSTRACT
Gypsophila huashanensis Y. W. Tsui & D. Q. Lu (Caryophyllaceae) is an endemic herb species to the Qinling Mountains in China. In this study, we characterized its whole plastid genome using the Illumina sequencing platform. The complete plastid genome of G. huashanensis is 152,457 bp in length, including a large single-copy DNA region of 83,476 bp, a small single-copy DNA region of 17,345 bp, and a pair of inverted repeat DNA sequences of 25,818 bp. The genome contains 130 genes comprising 85 protein-coding genes, 37 tRNA genes, and eight rRNA genes. Evolutionary analysis showed that the non-coding regions of Caryophyllaceae exhibit a higher level of divergence than the exon regions. Gene site selection analysis suggested that 11 coding protein genes (accD, atpF, ndhA, ndhB, petB, petD, rpoCl, rpoC2, rps16, ycfl, and ycf2) have some sites under protein sequence evolution. Phylogenetic analysis showed that G. huashanensis is most closely related to the congeneric species G. oldhamiana. These results are very useful for studying phylogenetic evolution and species divergence in the family Caryophyllaceae.
ABSTRACT
Hybridization is recognized as a major force in species evolution and biodiversity formation, generally leading to the origin and differentiation of new species. Multiple hybridization events cannot easily be reconstructed, yet they offer the potential to study a number of evolutionary processes. Here, we used nuclear expressed sequence tag-simple sequence repeat and large-scale single nucleotide polymorphism variation data, combined with niche analysis, to investigate the putative independent hybridization events in Notopterygium, a group of perennial herb plants endemic to China. Population genomic analysis indicated that the four studied species are genetically well-delimited and that N. forrestii and N. oviforme have originated by hybridization. According to Approximate Bayesian Computation, the best-fit model involved the formation of N. forrestii from the crossing of N. franchetii and N. incisum, with N. forrestii further backcrossing to N. franchetii to form N. oviforme. The niche analyses indicated that niche divergence [likely triggered by the regional climate changes, particularly the intensification of East Asian winter monsoon, and tectonic movements (affecting both Qinghai-Tibetan Plateau and Qinling Mountains)] may have promoted and maintained the reproductive isolation among hybrid species. N. forrestii shows ecological specialization with respect to their parental species, whereas N. oviforme has completely shifted its niche. These results suggested that the climate and environmental factors together triggered the two-step hybridization of the East Asia herb plants. Our study also emphasizes the power of genome-wide SNPs for investigating suspected cases of hybridization, particularly unravelling old hybridization events.
Subject(s)
Apiaceae , Hybridization, Genetic , Apiaceae/genetics , Bayes Theorem , Ecosystem , Metagenomics , PhylogenyABSTRACT
Inappropriate CD4+ T helper (Th) differentiation can compromise host immunity or promote autoimmune disease. To identify disease-relevant regulators of T cell fate, we examined mutations that modify risk for multiple sclerosis (MS), a canonical organ-specific autoimmune disease. This analysis identified a role for Zinc finger E-box-binding homeobox (ZEB1). Deletion of ZEB1 protects against experimental autoimmune encephalitis (EAE), a mouse model of multiple sclerosis (MS). Mechanistically, ZEB1 in CD4+ T cells is required for pathogenic Th1 and Th17 differentiation. Genomic analyses of paired human and mouse expression data elucidated an unexpected role for ZEB1 in JAK-STAT signaling. ZEB1 inhibits miR-101-3p that represses JAK2 expression, STAT3/STAT4 phosphorylation, and subsequent expression of interleukin-17 (IL-17) and interferon gamma (IFN-γ). Underscoring its clinical relevance, ZEB1 and JAK2 downregulation decreases pathogenic cytokines expression in T cells from MS patients. Moreover, a Food and Drug Administration (FDA)-approved JAK2 inhibitor is effective in EAE. Collectively, these findings identify a conserved, potentially targetable mechanism regulating disease-relevant inflammation.
Subject(s)
Cell Differentiation/physiology , Interleukin-17/metabolism , Multiple Sclerosis/pathology , Th17 Cells/immunology , Zinc Finger E-box-Binding Homeobox 1/metabolism , Animals , Cell Differentiation/immunology , Cytokines/metabolism , Encephalomyelitis, Autoimmune, Experimental/immunology , Humans , Interleukin-17/immunology , Mice , Multiple Sclerosis/immunology , Th1 Cells/immunology , Zinc Finger E-box-Binding Homeobox 1/genetics , Zinc Finger E-box-Binding Homeobox 1/immunologyABSTRACT
BACKGROUND: Epidermal Growth Factor Receptor (EGFR) tyrosine kinase inhibitors (TKIs) like erlotinib are effective for treating patients with EGFR mutant lung cancer; however, drug resistance inevitably emerges. Approaches to combine immunotherapies and targeted therapies to overcome or delay drug resistance have been hindered by limited knowledge of the effect of erlotinib on tumor-infiltrating immune cells. METHODS: Using mouse models, we studied the immunological profile of mutant EGFR-driven lung tumors before and after erlotinib treatment. RESULTS: We found that erlotinib triggered the recruitment of inflammatory T cells into the lungs and increased maturation of alveolar macrophages. Interestingly, this phenotype could be recapitulated by tumor regression mediated by deprivation of the EGFR oncogene indicating that tumor regression alone was sufficient for these immunostimulatory effects. We also found that further efforts to boost the function and abundance of inflammatory cells, by combining erlotinib treatment with anti-PD-1 and/or a CD40 agonist, did not improve survival in an EGFR-driven mouse model. CONCLUSIONS: Our findings lay the foundation for understanding the effects of TKIs on the tumor microenvironment and highlight the importance of investigating targeted and immuno-therapy combination strategies to treat EGFR mutant lung cancer.
Subject(s)
Antineoplastic Agents/therapeutic use , ErbB Receptors/genetics , ErbB Receptors/immunology , Erlotinib Hydrochloride/therapeutic use , Lung Neoplasms , Protein Kinase Inhibitors/therapeutic use , Animals , Lung Neoplasms/drug therapy , Lung Neoplasms/genetics , Lung Neoplasms/immunology , Mice, Transgenic , Mutation , Oncogenes , T-Lymphocytes/drug effects , T-Lymphocytes/immunology , Tumor Microenvironment/drug effects , Tumor Microenvironment/immunologyABSTRACT
Long-term immunity depends partly on the establishment of memory CD8+ T cells. We identified a counterregulatory network between the homologous transcription factors ZEB1 and ZEB2 and the miR-200 microRNA family, which modulates effector CD8+ T cell fates. Unexpectedly, Zeb1 and Zeb2 had reciprocal expression patterns and were functionally uncoupled in CD8+ T cells. ZEB2 promoted terminal differentiation, whereas ZEB1 was critical for memory T cell survival and function. Interestingly, the transforming growth factor ß (TGF-ß) and miR-200 family members, which counterregulate the coordinated expression of Zeb1 and Zeb2 during the epithelial-to-mesenchymal transition, inversely regulated Zeb1 and Zeb2 expression in CD8+ T cells. TGF-ß induced and sustained Zeb1 expression in maturing memory CD8+ T cells. Meanwhile, both TGF-ß and miR-200 family members selectively inhibited Zeb2. Additionally, the miR-200 family was necessary for optimal memory CD8+ T cell formation. These data outline a previously unknown genetic pathway in CD8+ T cells that controls effector and memory cell fate decisions.
Subject(s)
CD8-Positive T-Lymphocytes/cytology , Cell Lineage , MicroRNAs/metabolism , Zinc Finger E-box Binding Homeobox 2/metabolism , Zinc Finger E-box-Binding Homeobox 1/metabolism , Animals , Cell Survival , Homeostasis , Immunity , Immunologic Memory , Lymphocytic choriomeningitis virus/physiology , Mice, Inbred C57BL , RNA, Messenger/genetics , RNA, Messenger/metabolism , Transforming Growth Factor beta/metabolism , Zinc Finger E-box Binding Homeobox 2/genetics , Zinc Finger E-box-Binding Homeobox 1/geneticsABSTRACT
Understanding immunological memory formation depends on elucidating how multipotent memory precursor (MP) cells maintain developmental plasticity and longevity to provide long-term immunity while other effector cells develop into terminally differentiated effector (TE) cells with limited survival. Profiling active (H3K27ac) and repressed (H3K27me3) chromatin in naive, MP, and TE CD8+ T cells during viral infection revealed increased H3K27me3 deposition at numerous pro-memory and pro-survival genes in TE relative to MP cells, indicative of fate restriction, but permissive chromatin at both pro-memory and pro-effector genes in MP cells, indicative of multipotency. Polycomb repressive complex 2 deficiency impaired clonal expansion and TE cell differentiation, but minimally impacted CD8+ memory T cell maturation. Abundant H3K27me3 deposition at pro-memory genes occurred late during TE cell development, probably from diminished transcription factor FOXO1 expression. These results outline a temporal model for loss of memory cell potential through selective epigenetic silencing of pro-memory genes in effector T cells.
Subject(s)
CD8-Positive T-Lymphocytes/immunology , Cell Differentiation/immunology , Chromatin/immunology , Polycomb Repressive Complex 2/immunology , Animals , CD8-Positive T-Lymphocytes/metabolism , Cell Differentiation/genetics , Chromatin/genetics , Chromatin/metabolism , Enhancer of Zeste Homolog 2 Protein/genetics , Enhancer of Zeste Homolog 2 Protein/immunology , Enhancer of Zeste Homolog 2 Protein/metabolism , Flow Cytometry , Forkhead Box Protein O1/genetics , Forkhead Box Protein O1/immunology , Forkhead Box Protein O1/metabolism , Gene Expression/immunology , Histones/immunology , Histones/metabolism , Immunoblotting , Immunologic Memory/genetics , Immunologic Memory/immunology , Lysine/immunology , Lysine/metabolism , Methylation , Mice, Inbred C57BL , Mice, Knockout , Mice, Transgenic , Models, Immunological , Multipotent Stem Cells/immunology , Multipotent Stem Cells/metabolism , Polycomb Repressive Complex 2/genetics , Polycomb Repressive Complex 2/metabolism , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
T cell responses are guided by cytokines that induce transcriptional regulators, which ultimately control differentiation of effector and memory T cells. However, it is unknown how the activities of these molecular regulators are coordinated and integrated during the differentiation process. Using genetic approaches and transcriptional profiling of antigen-specific CD8(+) T cells, we reveal a common program of effector differentiation that is regulated by IL-2 and IL-12 signaling and the combined activities of the transcriptional regulators Blimp-1 and T-bet. The loss of both T-bet and Blimp-1 leads to abrogated cytotoxic function and ectopic IL-17 production in CD8(+) T cells. Overall, our data reveal two major overlapping pathways of effector differentiation governed by the availability of Blimp-1 and T-bet and suggest a model for cytokine-induced transcriptional changes that combine, quantitatively and qualitatively, to promote robust effector CD8(+) T cell differentiation.
Subject(s)
CD8-Positive T-Lymphocytes/immunology , Cell Differentiation/immunology , Interleukin-12/immunology , Interleukin-2/immunology , T-Box Domain Proteins/immunology , Transcription Factors/immunology , Animals , Arenaviridae Infections/immunology , Chromatin Immunoprecipitation , Cytokines/immunology , Flow Cytometry , Gene Expression Profiling , Influenza A Virus, H1N1 Subtype , Interleukin-17/immunology , Lymphocytic choriomeningitis virus , Mice , Orthomyxoviridae Infections/immunology , Positive Regulatory Domain I-Binding Factor 1 , Real-Time Polymerase Chain Reaction , STAT4 Transcription Factor/immunology , STAT5 Transcription Factor/immunology , Sequence Analysis, RNA , Signal TransductionABSTRACT
The transcription factor T-bet is critical for cytotoxic T lymphocyte (CTL) differentiation, but it is unclear how it operates in a graded manner in the formation of both terminal effector and memory precursor cells during viral infection. We find that, at high concentrations, T-bet induced expression of Zeb2 mRNA, which then triggered CTLs to adopt terminally differentiated states. ZEB2 and T-bet cooperate to switch on a terminal CTL differentiation program, while simultaneously repressing genes necessary for central memory CTL development. Chromatin immunoprecipitation sequencing showed that a large proportion of these genes were bound by T-bet, and this binding was altered by ZEB2 deficiency. Furthermore, T-bet overexpression could not fully bypass ZEB2 function. Thus, the coordinated actions of T-bet and ZEB2 outline a novel genetic pathway that forces commitment of CTLs to terminal differentiation, thereby restricting their memory cell potential.
Subject(s)
Cell Differentiation/immunology , Homeodomain Proteins/immunology , Lymphocytic Choriomeningitis/immunology , Repressor Proteins/immunology , T-Box Domain Proteins/immunology , T-Lymphocytes, Cytotoxic/immunology , Animals , CD8-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/metabolism , Cell Differentiation/genetics , Cluster Analysis , Flow Cytometry , Homeodomain Proteins/genetics , Host-Pathogen Interactions/immunology , Lectins, C-Type , Lymphocytic Choriomeningitis/genetics , Lymphocytic Choriomeningitis/virology , Lymphocytic choriomeningitis virus/immunology , Lymphocytic choriomeningitis virus/physiology , Mice, Inbred C57BL , Mice, Knockout , Mice, Transgenic , Oligonucleotide Array Sequence Analysis , Protein Binding/immunology , Receptors, Immunologic/immunology , Receptors, Immunologic/metabolism , Repressor Proteins/deficiency , Repressor Proteins/genetics , Reverse Transcriptase Polymerase Chain Reaction , T-Box Domain Proteins/genetics , T-Box Domain Proteins/metabolism , T-Lymphocytes, Cytotoxic/metabolism , Transcriptome/genetics , Transcriptome/immunology , Zinc Finger E-box Binding Homeobox 2ABSTRACT
Memory CD8(+) T cells are critical for host defense upon reexposure to intracellular pathogens. We found that interleukin 10 (IL-10) derived from CD4(+) regulatory T cells (Treg cells) was necessary for the maturation of memory CD8(+) T cells following acute infection with lymphocytic choriomeningitis virus (LCMV). Treg cell-derived IL-10 was most important during the resolution phase, calming inflammation and the activation state of dendritic cells. Adoptive transfer of IL-10-sufficient Treg cells during the resolution phase 'restored' the maturation of memory CD8(+) T cells in IL-10-deficient mice. Our data indicate that Treg cell-derived IL-10 is needed to insulate CD8(+) T cells from inflammatory signals, and reveal that the resolution phase of infection is a critical period that influences the quality and function of developing memory CD8(+) T cells.
Subject(s)
CD8-Positive T-Lymphocytes/immunology , Interleukin-10/immunology , Lymphocytic Choriomeningitis/immunology , Lymphocytic choriomeningitis virus/immunology , T-Lymphocytes, Regulatory/immunology , Adoptive Transfer , Animals , CD8-Positive T-Lymphocytes/metabolism , Dendritic Cells/immunology , Dendritic Cells/metabolism , Flow Cytometry , Gene Expression Profiling , Host-Pathogen Interactions/immunology , Immunologic Memory/immunology , Inflammation/genetics , Inflammation/immunology , Inflammation/metabolism , Interleukin-10/genetics , Interleukin-10/metabolism , Lymphocytic Choriomeningitis/genetics , Lymphocytic Choriomeningitis/virology , Lymphocytic choriomeningitis virus/physiology , Mice, Inbred C57BL , Mice, Knockout , Reverse Transcriptase Polymerase Chain Reaction , T-Lymphocytes, Regulatory/metabolism , T-Lymphocytes, Regulatory/transplantationABSTRACT
Tissue-resident memory T (Trm) cells provide enhanced protection against infection at mucosal sites. Here we found that CD4(+) T cells are important for the formation of functional lung-resident CD8(+) T cells after influenza virus infection. In the absence of CD4(+) T cells, CD8(+) T cells displayed reduced expression of CD103 (Itgae), were mislocalized away from airway epithelia, and demonstrated an impaired ability to recruit CD8(+) T cells to the lung airways upon heterosubtypic challenge. CD4(+) T cell-derived interferon-γ was necessary for generating lung-resident CD103(+) CD8(+) Trm cells. Furthermore, expression of the transcription factor T-bet was increased in "unhelped" lung Trm cells, and a reduction in T-bet rescued CD103 expression in the absence of CD4(+) T cell help. Thus, CD4(+) T cell-dependent signals are important to limit expression of T-bet and allow for the development of CD103(+) CD8(+) Trm cells in the lung airways following respiratory infection.
Subject(s)
CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/immunology , Immunologic Memory , Influenza A Virus, H3N2 Subtype/immunology , Lung/immunology , Orthomyxoviridae Infections/immunology , T-Box Domain Proteins/biosynthesis , Animals , Antigens, CD/immunology , Integrin alpha Chains/immunology , Interferon-gamma/immunology , Lung/cytology , Lymphocyte Activation/immunology , Mice , Mice, Inbred C57BL , Mice, Knockout , Mucous Membrane/cytology , Mucous Membrane/immunologyABSTRACT
The transcription factor Blimp-1 regulates the overall accumulation of virus-specific CD8⺠T cells during acute viral infections. We found that increased proliferation and survival of Blimp-1-deficient CD8⺠T cells resulted from sustained expression of CD25 and CD27 and persistent cytokine responsiveness. Silencing of Il2ra and Cd27 reduced the Blimp-1-deficient CD8⺠T cell response. Genome-wide chromatin immunoprecipitation (ChIP) sequencing analysis identified Il2ra and Cd27 as direct targets of Blimp-1. At the peak of the antiviral response, but not earlier, Blimp-1 recruited the histone-modifying enzymes G9a and HDAC2 to the Il2ra and Cd27 loci, thereby repressing expression of these genes. In the absence of Blimp-1, Il2ra and Cd27 exhibited enhanced histone H3 acetylation and reduced histone H3K9 trimethylation. These data elucidate a central mechanism by which Blimp-1 acts as an epigenetic regulator and enhances the numbers of short-lived effector cells while suppressing the development of memory-precursor CD8⺠T cells.
Subject(s)
CD8-Positive T-Lymphocytes/immunology , Epigenesis, Genetic/immunology , Lymphocytic Choriomeningitis/genetics , Lymphocytic choriomeningitis virus/immunology , Transcription Factors/genetics , Amino Acid Sequence , Animals , CD8-Positive T-Lymphocytes/pathology , CD8-Positive T-Lymphocytes/virology , Disease Progression , Histone Deacetylase 2/genetics , Histone Deacetylase 2/immunology , Histone-Lysine N-Methyltransferase/genetics , Histone-Lysine N-Methyltransferase/immunology , Histones/genetics , Histones/immunology , Humans , Immunologic Memory , Interleukin-2 Receptor alpha Subunit/antagonists & inhibitors , Interleukin-2 Receptor alpha Subunit/genetics , Interleukin-2 Receptor alpha Subunit/immunology , Lymphocytic Choriomeningitis/immunology , Lymphocytic Choriomeningitis/pathology , Lymphocytic Choriomeningitis/virology , Methylation , Mice , Mice, Transgenic , Molecular Sequence Data , Positive Regulatory Domain I-Binding Factor 1 , RNA, Small Interfering/genetics , RNA, Small Interfering/immunology , Signal Transduction , Transcription Factors/immunology , Tumor Necrosis Factor Receptor Superfamily, Member 7/antagonists & inhibitors , Tumor Necrosis Factor Receptor Superfamily, Member 7/genetics , Tumor Necrosis Factor Receptor Superfamily, Member 7/immunologyABSTRACT
Host antibody response is a crucial defense against pathogenic infection. Here, we report a novel technique allowing quantitative measurement of polyclonal antibody response in vivo. This involves expression of a combinatorial library of target proteins from a candidate pathogen on the surface of yeast Saccharomyces cerevisiae. After mixing with serum/plasma from infected or immunized subjects, positive yeast clones were isolated via fluorescence-activated cell sorting (FACS). Using this technique, we have studied mouse immunized serum with recombinant hemagglutinin (HA) protein from a human influenza H5N1 strain (A/Anhui/1/2005) and convalescent plasma from an infected human in China. Our technique has identified novel antigenic domains targeted by serum/plasma and allowed calculation of the relative proportion of the antibody response against each domain. We believe such systematic measurement of an antibody response is unprecedented, and applying this method to different pathogens will improve understanding of protective immunity and guide development of vaccines and therapeutics.