The ubiquitin ligase Peli1 negatively regulates T cell activation and prevents autoimmunity.

T cell activation is subject to tight regulation to avoid inappropriate responses to self antigens. Here we show that genetic deficiency in the ubiquitin ligase Peli1 caused hyperactivation of T cells and rendered T cells refractory to suppression by regulatory T cells and transforming growth factor-ß (TGF-ß). As a result, Peli1-deficient mice spontaneously developed autoimmunity characterized by multiorgan inflammation and autoantibody production. Peli1 deficiency resulted in the nuclear accumulation of c-Rel, a member of the NF-κB family of transcription factors with pivotal roles in T cell activation. Peli1 negatively regulated c-Rel by mediating its Lys48 (K48) ubiquitination. Our results identify Peli1 as a critical factor in the maintenance of peripheral T cell tolerance and demonstrate a previously unknown mechanism of c-Rel regulation.

A chromosome-scale genome sequence of sudangrass (Sorghum sudanense) highlights the genome evolution and regulation of dhurrin biosynthesis.

KEY MESSAGE: Sudangrass is more similar to US commercial sorghums than to cultivated sorghums from Africa sequence-wise and contain significantly lower dhurrin than sorghums. CYP79A1 is linked to dhurrin content in sorghum. Sudangrass [Sorghum sudanense (Piper) Stapf] is a hybrid between grain sorghum and its wild relative S. bicolor ssp. verticilliflorum and is grown as a forage crop due to its high biomass production and low dhurrin content compared to sorghum. In this study, we sequenced the sudangrass genome and showed that the assembled genome was 715.95 Mb with 35,243 protein-coding genes. Phylogenetic analysis with whole genome proteomes demonstrated that the sudangrass genome was more similar to US commercial sorghums than to its wild relatives and cultivated sorghums from Africa. We confirmed that at seedling stage, sudangrass accessions contained significantly lower dhurrin as measured by hydrocyanic acid potential (HCN-p) than cultivated sorghum accessions. Genome-wide association study identified a QTL most tightly associated with HCN-p and the linked SNPs were located in the 3' UTR of Sobic.001G012300 which encodes CYP79A1, the enzyme that catalyzes the first step of dhurrin biosynthesis. As in other grasses such as maize and rice, we also found that copia/gypsy long terminal repeat (LTR) retrotransposons were more abundant in cultivated than in wild sorghums, implying that crop domestication in the grasses was accompanied by increased copia/gypsy LTR retrotransposon insertions in the genomes.

Genome-wide association mapping identifies an SNF4 ortholog that impacts biomass and sugar yield in sorghum and sugarcane.

Sorghum is a feed/industrial crop in developed countries and a staple food elsewhere in the world. This study evaluated the sorghum mini core collection for days to 50% flowering (DF), biomass, plant height (PH), soluble solid content (SSC), and juice weight (JW), and the sorghum reference set for DF and PH, in 7-12 testing environments. We also performed genome-wide association mapping with 6 094 317 and 265 500 single nucleotide polymorphism markers in the mini core collection and the reference set, respectively. In the mini core panel we identified three quantitative trait loci for DF, two for JW, one for PH, and one for biomass. In the reference set panel we identified another quantitative trait locus for PH on chromosome 6 that was also associated with biomass, DF, JW, and SSC in the mini core panel. Transgenic studies of three genes selected from the locus revealed that Sobic.006G061100 (SbSNF4-2) increased biomass, SSC, JW, and PH when overexpressed in both sorghum and sugarcane, and delayed flowering in transgenic sorghum. SbSNF4-2 encodes a γ subunit of the evolutionarily conserved AMPK/SNF1/SnRK1 heterotrimeric complexes. SbSNF4-2 and its orthologs will be valuable in genetic enhancement of biomass and sugar yield in plants.

Fibroblast growth factor 21 attenuates salt-sensitive hypertension-induced nephropathy through anti-inflammation and anti-oxidation mechanism.

BACKGROUND: Patients with salt-sensitive hypertension are often accompanied with severe renal damage and accelerate to end-stage renal disease, which currently lacks effective treatment. Fibroblast growth factor 21 (FGF21) has been shown to suppress nephropathy in both type 1 and type 2 diabetes mice. Here, we aimed to investigate the therapeutic effect of FGF21 in salt-sensitive hypertension-induced nephropathy. METHODS: Changes of FGF21 expression in deoxycorticosterone acetate (DOCA)-salt-induced hypertensive mice were detected. The influence of FGF21 knockout in mice on DOCA-salt-induced nephropathy were determined. Recombinant human FGF21 (rhFGF21) was intraperitoneally injected into DOCA-salt-induced nephropathy mice, and then the inflammatory factors, oxidative stress levels and kidney injury-related indicators were observed. In vitro, human renal tubular epithelial cells (HK-2) were challenged by palmitate acid (PA) with or without FGF21, and then changes in inflammation and oxidative stress indicators were tested. RESULTS: We observed significant elevation in circulating levels and renal expression of FGF21 in DOCA-salt-induced hypertensive mice. We found that deletion of FGF21 in mice aggravated DOCA-salt-induced nephropathy. Supplementation with rhFGF21 reversed DOCA-salt-induced kidney injury. Mechanically, rhFGF21 induced AMPK activation in DOCA-salt-treated mice and PA-stimulated HK-2 cells, which inhibited NF-κB-regulated inflammation and Nrf2-mediated oxidative stress and thus, is important for rhFGF21 protection against DOCA-salt-induced nephropathy. CONCLUSION: These findings indicated that rhFGF21 could be a promising pharmacological strategy for the treatment of salt-sensitive hypertension-induced nephropathy.

Two functional subsets of FOXP3+ regulatory T cells in human thymus and periphery.

Previous studies suggest that thymus produces a homogenous population of natural regulatory T (Treg) cells that express a transcriptional factor FOXP3 and control autoimmunity through a cell-contact-dependent mechanism. We found two subsets of FOXP3+ natural Treg cells defined by the expression of the costimulatory molecule ICOS in the human thymus and periphery. Whereas the ICOS+FOXP3+ Treg cells used interleukin-10 to suppress dendritic cell function and transforming growth factor (TGF)-beta to suppress T cell function, the ICOS-FOXP3+ Treg cells used TGF-beta only. The survival and proliferation of the two subsets of Treg cells were differentially regulated by signaling through ICOS or CD28, respectively. We suggest that the selection of natural Treg cells in thymus is coupled with Treg cell differentiation into two subsets imprinted with different cytokine expression potentials and use both cell-contact-dependent and independent mechanisms for immunosuppression in periphery.

Generation of T follicular helper cells is mediated by interleukin-21 but independent of T helper 1, 2, or 17 cell lineages.

After activation, CD4(+) helper T (Th) cells differentiate into distinct effector subsets. Although chemokine (C-X-C motif) receptor 5-expressing T follicular helper (Tfh) cells are important in humoral immunity, their developmental regulation is unclear. Here we show that Tfh cells had a distinct gene expression profile and developed in vivo independently of the Th1 or Th2 cell lineages. Tfh cell generation was regulated by ICOS ligand (ICOSL) expressed on B cells and was dependent on interleukin-21 (IL-21), IL-6, and signal transducer and activator of transcription 3 (STAT3). However, unlike Th17 cells, differentiation of Tfh cells did not require transforming growth factor beta (TGF-beta) or Th17-specific orphan nuclear receptors RORalpha and RORgamma in vivo. Finally, naive T cells activated in vitro in the presence of IL-21 but not TGF-beta signaling preferentially acquired Tfh gene expression and promoted germinal-center reactions in vivo. This study thus demonstrates that Tfh is a distinct Th cell lineage.

Association mapping of germinability and seedling vigor in sorghum under controlled low-temperature conditions.

Sorghum is one of the world's most important food, feed, and fiber crops as well as a potential feedstock for lignocellulosic bioenergy. Early-season planting extends sorghum's growing season and increases yield in temperate regions. However, sorghum's sensitivity to low soil temperatures adversely impacts seed germination. In this study, we evaluated the 242 accessions of the ICRISAT sorghum mini core collection for seed germination and seedling vigor at 12 °C as a measure of cold tolerance. Genome-wide association analysis was performed with approximately 162,177 single nucleotide polymorphism markers. Only one marker locus (Locus 7-2) was significantly associated with low-temperature germination and none with vigor. The linkage of Locus 7-2 to low-temperature germination was supported by four lines of evidence: strong association in three independent experiments, co-localization with previously mapped cold tolerance quantitative trait loci (QTL) in sorghum, a candidate gene that increases cold tolerance and germination rate when its wheat homolog is overexpressed in tobacco, and its syntenic region in rice co-localized with two cold tolerance QTL in rice. This locus may be useful in developing tools for molecular breeding of sorghums with improved low-temperature germinability.

CCAAT/enhancer-binding protein α negatively regulates IFN-γ expression in T cells.

Humoral immunity, including Ab switching and somatic hypermutation, is critically regulated by CD4(+) T cells. T follicular helper (Tfh) cells have been recently shown to be a distinct T cell subset important in germinal center reactions. The transcriptional regulation of Tfh cell development and function has not been well understood. In this study, we report that C/EBPα, a basic region/leucine zipper transcription factor, is highly expressed in Tfh cells. Cebpa-deficient CD4(+) T cells exhibit enhanced IFN-γ expression in vitro and in vivo. T cell-specific Cebpa knockout mice, although not defective in Tfh cell generation, produce significantly increased levels of IgG2a/b and IgG3 following immunization with a protein Ag. Moreover, C/EBPα binds to the Ifng gene and inhibits T-bet-driven Ifng transcription in a DNA binding-dependent manner. Our study thus demonstrates that C/EBPα restricts IFN-γ expression in T cells to allow proper class switching by B cells.

A gene in the multidrug and toxic compound extrusion (MATE) family confers aluminum tolerance in sorghum.

Crop yields are significantly reduced by aluminum toxicity on highly acidic soils, which comprise up to 50% of the world's arable land. Candidate aluminum tolerance proteins include organic acid efflux transporters, with the organic acids forming non-toxic complexes with rhizosphere aluminum. In this study, we used positional cloning to identify the gene encoding a member of the multidrug and toxic compound extrusion (MATE) family, an aluminum-activated citrate transporter, as responsible for the major sorghum (Sorghum bicolor) aluminum tolerance locus, Alt(SB). Polymorphisms in regulatory regions of Alt(SB) are likely to contribute to large allelic effects, acting to increase Alt(SB) expression in the root apex of tolerant genotypes. Furthermore, aluminum-inducible Alt(SB) expression is associated with induction of aluminum tolerance via enhanced root citrate exudation. These findings will allow us to identify superior Alt(SB) haplotypes that can be incorporated via molecular breeding and biotechnology into acid soil breeding programs, thus helping to increase crop yields in developing countries where acidic soils predominate.

Cutting edge: regulation of intestinal inflammation and barrier function by IL-17C.

In the IL-17 family of cytokines, much is known about the sources and functions of IL-17, IL-17F, and IL-25 in the host defense against infection and in inflammatory diseases; however, the physiological function of IL-17C remains poorly understood. Using mice deficient in IL-17C, we demonstrate that this cytokine is crucial for the regulation of an acute experimental colitis elicited by dextran sulfate sodium. In this model, mice lacking IL-17C exhibited exacerbated disease that was associated with increased IL-17 expression by γδ T cells and Th17 cells. Moreover, IL-17C directly regulated the expression of the tight junction molecule occludin by colonic epithelial cells. Thus, our data suggest that IL-17C plays a critical role in maintaining mucosal barrier integrity.

Profiling Gene Expression in Germinating Brassica Roots.

Based on previously developed solid-phase gene extraction (SPGE) we examined the mRNA profile in primary roots of Brassica rapa seedlings for highly expressed genes like ACT7 (actin7), TUB (tubulin1), UBQ (ubiquitin), and low expressed GLK (glucokinase) during the first day post-germination. The assessment was based on the mRNA load of the SPGE probe of about 2.1 ng. The number of copies of the investigated genes changed spatially along the length of primary roots. The expression level of all genes differed significantly at each sample position. Among the examined genes ACT7 expression was most even along the root. UBQ was highest at the tip and root-shoot junction (RS). TUB and GLK showed a basipetal gradient. The temporal expression of UBQ was highest in the MZ 9 h after primary root emergence and higher than at any other sample position. Expressions of GLK in EZ and RS increased gradually over time. SPGE extraction is the result of oligo-dT and oligo-dA hybridization and the results illustrate that SPGE can be used for gene expression profiling at high spatial and temporal resolution. SPGE needles can be used within two weeks when stored at 4 °C. Our data indicate that gene expression studies that are based on the entire root miss important differences in gene expression that SPGE is able to resolve for example growth adjustments during gravitropism.

Genome-wide association study of plant color in Sorghum bicolor.

Introduction: Sorghum plant color is the leaf sheath/leaf color and is associated with seed color, tannin and phenol content, head blight disease incidence, and phytoalexin production. Results: In this study, we evaluated plant color of the sorghum mini core collection by scoring leaf sheath/leaf color at maturity as tan, red, or purple across three testing environments and performed genome-wide association mapping (GWAS) with 6,094,317 SNPs markers. Results and Discussion: Eight loci, one each on chromosomes 1, 2, 4, and 6 and two on chromosomes 5 and 9, were mapped. All loci contained one to three candidate genes. In qPC5-1, Sobic.005G165632 and Sobic.005G165700 were located in the same linkage disequilibrium (LD) block. In qPC6, Sobic.006G149650 and Sobic.006G149700 were located in the different LD block. The single peak in qPC6 covered one gene, Sobic.006G149700, which was a senescence regulator. We found a loose correlation between the degree of linkage and tissue/organ expression of the underlying genes possibly related to the plant color phenotype. Allele analysis indicated that none of the linked SNPs can differentiate between red and purple accessions whereas all linked SNPs can differentiate tan from red/purple accessions. The candidate genes and SNP markers may facilitate the elucidation of plant color development as well as molecular plant breeding.

The protective effects of icariin against testicular dysfunction in type 1 diabetic mice Via AMPK-mediated Nrf2 activation and NF-κB p65 inhibition.

BACKGROUND: Owing to the early suffering age and the rising incidence of type 1 diabetes (T1D), the resulting male reproductive dysfunction and fertility decline have become a disturbing reality worldwide, with no effective strategy being available. Icariin (ICA), a flavonoid extracted from Herba Epimedium, has been proved its promising application in improving diabetes-related complications including diabetic nephropathy, endothelial dysfunction and erectile dysfunction. Ensuring the future reproductive health of children and adolescents with T1D is crucial to improve global fertility. However, its roles in the treatment of T1D-induced testicular dysfunction and the potential mechanisms remain elusive. PURPOSE: The purpose of this present study was to investigate whether ICA ameliorates T1D-induced testicular dysfunction as well as its potential mechanisms. METHODS: T1D murine model was established by intraperitoneal injection of STZ with or without treated with ICA for eleven weeks. Morphological, pathological and serological experiments were used to determine the efficacy of ICA on male reproductive function of T1D mice. Western blotting, Immunohistochemistry analysis, qRT-PCR and kit determination were performed to investigated the underlying mechanisms. RESULTS: We found that replenishment of ICA alleviated testicular damage, promoted testosterone production and spermatogenesis, ameliorated apoptosis and blood testis barrier impairment in streptozotocin-induced T1D mice. Functionally, ICA treatment triggered adenosine monophosphate protein kinase (AMPK) activation, which in turn inhibited the nuclear translocation of nuclear factor kappa B p65 (NF-κB p65) to reduce inflammatory responses in the testis and activated nuclear factor erythroid 2-related factor 2(Nrf2), thereby enhancing testicular antioxidant capacity. Further studies revealed that supplementation with the AMPK antagonist Compound C or depletion of Nrf2 weakened the beneficial effects of ICA on testicular dysfunction of T1D mice. CONCLUSION: Collectively, these results demonstrate the feasibility of ICA in the treatment of T1D-induced testicular dysfunction, and reveal the important role of AMPK-mediated Nrf2 activation and NF-κB p65 inhibition in ICA-associated testicular protection during T1D.

Interleukin 25 promotes the initiation of proallergic type 2 responses.

The molecular mechanisms underlying the initiation of innate and adaptive proallergic type 2 responses are not understood. Interleukin (IL) 25, a member of the IL-17 cytokine family, was recently reported (Owyang, A.M., C. Zaph, E.H. Wilson, K.J. Guild, T. McClanahan, H.R. Miller, D.J. Cua, M. Goldschmidt, C.A. Hunter, R.A. Kastelein, and D. Artis. 2006. J. Exp. Med. 203:843-849; Fallon, P.G., S.J. Ballantyne, N.E. Mangan, J.L. Barlow, A. Dasvarma, D.R. Hewett, A. McIlgorm, H.E. Jolin, and A.N. McKenzie. 2006. J. Exp. Med. 203:1105-1116) to be important in Th2 cell-mediated immunity to parasitic infection. However, the cellular source and targets of IL-25 are not well understood. We show that mouse IL-25 is expressed by lung epithelial cells as a result of innate immune responses to allergens. Transgenic overexpression of IL-25 by these cells leads to mucus production and airway infiltration of macrophages and eosinophils, whereas blockade of IL-25 conversely reduces the airway inflammation and Th2 cytokine production in an allergen-induced asthma model. In addition, IL-25, with a receptor more highly expressed in Th2 than other effector T cells, promotes Th2 cell differentiation in an IL-4- and signal transducer and activator of transcription 6-dependent manner. During early T cell activation, IL-25 potentiates expression of the nuclear factor of activated T cells c1 and JunB transcription factors, which possibly results in increased levels of initial IL-4 production, up-regulation of GATA-3 expression, and enhanced Th2 cell differentiation. Thus, IL-25 is a critical factor regulating the initiation of innate and adaptive proallergic responses.

PACSIN1 regulates the TLR7/9-mediated type I interferon response in plasmacytoid dendritic cells.

Plasmacytoid dendritic cells (pDCs) are the professional interferon (IFN)-producing cells of the immune system. pDCs specifically express Toll-like receptor (TLR)7 and TLR9 molecules and produce massive amounts of type I IFN by sensing microbial nucleic acids via TLR7 and TLR9. Here we report that protein kinase C and casein kinase substrate in neurons (PACSIN) 1, is specifically expressed in human and mouse pDCs. Knockdown of PACSIN1 by short hairpin RNA (shRNA) in a human pDC cell line significantly inhibited the type I IFN response of the pDCs to TLR9 ligand. PACSIN1-deficient mice exhibited normal levels of conventional DCs and pDCs, demonstrating that development of pDCs was intact although PACSIN1-deficient pDCs showed reduced levels of IFN-α production in response to both cytosine guanine dinucleotide (CpG)-oligonucleotide (ODN) and virus. In contrast, the production of proinflammatory cytokines in response to those ligands was not affected in PACSIN1-deficient pDCs, suggesting that PACSIN1 represents a pDC-specific adaptor molecule that plays a specific role in the type I IFN signaling cascade.

Identification of genetic markers linked to anthracnose resistance in sorghum using association analysis.

Anthracnose in sorghum caused by Colletotrichum sublineolum is one of the most destructive diseases affecting sorghum production under warm and humid conditions. Markers and genes linked to resistance to the disease are important for plant breeding. Using 14,739 SNP markers, we have mapped eight loci linked to resistance in sorghum through association analysis of a sorghum mini-core collection consisting of 242 diverse accessions evaluated for anthracnose resistance for 2 years in the field. The mini-core was representative of the International Crops Research Institute for the Semi-Arid Tropics' world-wide sorghum landrace collection. Eight marker loci were associated with anthracnose resistance in both years. Except locus 8, disease resistance-related genes were found in all loci based on their physical distance from linked SNP markers. These include two NB-ARC class of R genes on chromosome 10 that were partially homologous to the rice blast resistance gene Pib, two hypersensitive response-related genes: autophagy-related protein 3 on chromosome 1 and 4 harpin-induced 1 (Hin1) homologs on chromosome 8, a RAV transcription factor that is also part of R gene pathway, an oxysterol-binding protein that functions in the non-specific host resistance, and homologs of menthone:neomenthol reductase (MNR) that catalyzes a menthone reduction to produce the antimicrobial neomenthol. These genes and markers may be developed into molecular tools for genetic improvement of anthracnose resistance in sorghum.

Association mapping of maturity and plant height using SNP markers with the sorghum mini core collection.

Plant height and maturity are two critical traits in sorghum breeding. To develop molecular tools and to identify genes underlying the traits for molecular breeding, we developed 14,739 SNP markers used to genotype the complete sorghum [Sorghum bicolor (L.) Moench] mini core collection. The collection was evaluated in four rainy and three post-rainy season environments for plant height and maturity. Association analysis identified six marker loci linked to height and ten to maturity in at least two environments with at least two SNPs in each locus. Of these, 14 were in close proximity to previously mapped height/maturity QTL in sorghum. Candidate genes for maturity or plant height close to the marker loci include a sugar transporter (SbSUC9), an auxin response factor (SbARF3), an FLC and FT regulator (SbMED12), and a photoperiod response gene (SbPPR1) for maturity and peroxidase 53, and an auxin transporter (SbLAX4) for plant height. Linkage disequilibrium analysis showed that SbPPR1 and SbARF3 were in regions with reduced sequence variation among early-maturing accessions, suggestive of past purifying selection. We also found a linkage disequilibrium block that existed only among the accessions with short plant height in rainy season environments. The block contains a gene homologous to the Arabidopsis flowering time gene, LUMINIDEPENDENS (LD). Functional LD promotes early maturity while mutation delays maturity, affecting plant height. Previous studies also found reduced sequence variations within this gene. These newly-mapped SNP markers will facilitate further efforts to identify plant height or maturity genes in sorghum.

Mapping and candidate genes associated with saccharification yield in sorghum.

Sorghum (Sorghum bicolor (L.) Moench) is a high-yielding, stress tolerant energy crop for lignocellulosic-based biofuel production. Saccharification is a process by which hydrolytic enzymes break down lignocellulosic materials to fermentable sugars for biofuel production, and mapping and identifying genes underlying saccharification yield is an important first step to genetically improve the plant for higher biofuel productivity. In this study, we used the ICRISAT sorghum mini core germplasm collection and 14 739 single nucleotide polymorphism markers to map saccharification yield. Seven marker loci were associated with saccharification yield and five of these loci were syntenic with regions in the maize genome that contain quantitative trait loci underlying saccharification yield and cell wall component traits. Candidate genes from the seven loci were identified but must be validated, with the most promising candidates being ß-tubulin, which determines the orientation of cellulose microfibrils in plant secondary cell walls, and NST1, a master transcription factor controlling secondary cell wall biosynthesis in fibers. Other candidate genes underlying the different saccharification loci included genes that play a role in vascular development and suberin deposition in plants. The identified loci and candidate genes provide information into the factors controlling saccharification yield and may facilitate increasing biofuel production in sorghum.

Plasmacytoid dendritic cells sense self-DNA coupled with antimicrobial peptide.

Plasmacytoid dendritic cells (pDCs) sense viral and microbial DNA through endosomal Toll-like receptors to produce type 1 interferons. pDCs do not normally respond to self-DNA, but this restriction seems to break down in human autoimmune disease by an as yet poorly understood mechanism. Here we identify the antimicrobial peptide LL37 (also known as CAMP) as the key factor that mediates pDC activation in psoriasis, a common autoimmune disease of the skin. LL37 converts inert self-DNA into a potent trigger of interferon production by binding the DNA to form aggregated and condensed structures that are delivered to and retained within early endocytic compartments in pDCs to trigger Toll-like receptor 9. Thus, our data uncover a fundamental role of an endogenous antimicrobial peptide in breaking innate tolerance to self-DNA and suggest that this pathway may drive autoimmunity in psoriasis.