Structural Basis for the Secretion of EvpC: A Key Type VI Secretion System Protein from Edwardsiella tarda

The recently identified type VI secretion system (T6SS) is implicated in the virulence of many Gram-negative bacteria. Edwardsiella tarda is an important cause of hemorrhagic septicemia in fish and also gastro- and extra-intestinal infections in humans. The E. tarda virulent protein (EVP) gene cluster encodes a conserved T6SS which contains 16 open reading frames. EvpC is one of the three major EVP secreted proteins and shares high sequence similarity with Hcp1, a key T6SS virulence factor from Pseudomonas aeruginosa. EvpC contributes to the virulence of E. tarda by playing an essential role in functional T6SS. Here, we report the crystal structure of EvpC from E. tarda PPD130/91 at a 2.8 Å resolution, along with functional studies of the protein. EvpC has a β-barrel domain with extended loops. The β-barrel consists of 11 anti-parallel β-strands with an α-helix located on one side. In solution, EvpC exists as a dimer at low concentration and as a hexamer at higher concentration. In the crystal, the symmetry related EvpC molecules form hexameric rings which stack together to form a tube similar to Hcp1. Structure based mutagenesis revealed that N-terminal negatively charged residues, Asp4, Glu15 and Glu26, and C-terminal positively charged residues, Lys161, Lys162 and Lys163, played crucial roles in the secretion of EvpC. Moreover, the localization study indicates the presence of wild type EvpC in cytoplasm, periplasm and secreted fractions, whereas the N-terminal and C-terminal mutants were found mostly in the periplasmic region and was completely absent in the secreted fraction. Results reported here provide insight into the structure, assembly and function of EvpC. Further, these findings can be extended to other EvpC homologs for understanding the mechanism of T6SS and targeting T6SS mediated virulence in Gram-negative pathogens

Abnormalities in circulating plasmacytoid dendritic cells in patients with systemic lupus erythematosus

Introduction: Dendritic cells (DCs) are capable of inducing immunity or tolerance. Previous studies have suggested plasmacytoid DCs (pDCs) are pathogenic in systemic lupus erythematosus (SLE). However, the functional characteristics of directly isolated peripheral circulating blood pDCs in SLE have not been evaluated previously.Methods: Peripheral blood pDCs from 62 healthy subjects and 58 SLE patients were treated with apoptotic cells derived from polymorphonuclear cells (PMNs). Antigen loaded or unloaded pDCs were then co-cultured with autologous or allogenous T cells. Changes in T cell proliferation, cell surface CD25 expression, intracellular Foxp3 expression and cytokine production were evaluated. pDCs that had captured apoptotic PMNs (pDCs + apoPMNs were also studied for their cytokine production (interferon (IFN)-alpha, interleukin (IL)-6, IL-10, IL-18) and toll like receptor (TLR) expression.Results: Circulating pDCs from SLE patients had an increased ability to stimulate T cells when compared with control pDCs. Using allogenous T cells as responder cells, SLE pDCs induced T cell proliferation even in the absence of apoptotic PMNs. In addition, healthy pDCs + apoPMNs induced suppressive T regulatory cell features with increased Foxp3 expression in CD4 + CD25 + cells while SLE pDCs + apoPMNs did not. There were differences in the cytokine profile of pDCs that had captured apoptotic PMNs between healthy subjects and patients with SLE. Healthy pDCs + apoPMNs showed decreased production of IL-6 but no significant changes in IL-10 and IL-18. These pDCs + apoPMNs also showed increased mRNA transcription of TLR9. On the other hand, while SLE pDCs + apoPMNs also had decreased IL-6, there was decreased IL-18 mRNA expression and persistent IL-10 protein synthesis. In addition, SLE pDCs lacked TLR9 recruitment.Conclusions: We have demonstrated that peripheral circulating pDCs in patients with SLE were functionally abnormal. They lacked TLR9 expression, were less capable of inducing regulatory T cell differentiation and had persistent IL-10 mRNA expression following the capture of apoptotic PMNs. We suggest circulating pDCs may be pathogenically relevant in SLE. © 2010 Jin et al.; licensee BioMed Central Ltd.published_or_final_versio

Structure of GrlR and the Implication of Its EDED Motif in Mediating the Regulation of Type III Secretion System in EHEC

Enterohemorrhagic Escherichia coli (EHEC) is a common cause of severe hemorrhagic colitis. EHEC's virulence is dependent upon a type III secretion system (TTSS) encoded by 41 genes. These genes are organized in several operons clustered in the locus of enterocyte effacement. Most of the locus of enterocyte effacement genes, including grlA and grlR, are positively regulated by Ler, and Ler expression is positively and negatively modulated by GrlA and GrlR, respectively. However, the molecular basis for the GrlA and GrlR activity is still elusive. We have determined the crystal structure of GrlR at 1.9 Å resolution. It consists of a typical β-barrel fold with eight β-strands containing an internal hydrophobic cavity and a plug-like loop on one side of the barrel. Strong hydrophobic interactions between the two β-barrels maintain the dimeric architecture of GrlR. Furthermore, a unique surface-exposed EDED (Glu-Asp-Glu-Asp) motif is identified to be critical for GrlA–GrlR interaction and for the repressive activity of GrlR. This study contributes a novel molecular insight into the mechanism of GrlR function

ITGAM is associated with disease susceptibility and renal nephritis of systemic lupus erythematosus in Hong Kong Chinese and Thai

ITGAM was recently found to be associated with systemic lupus erythematosus (SLE) in populations of not only European ancestry, but also in Hispanic- and African-Americans, Mexicans and Colombians. The risk alleles in the gene, however, were found to be monomorphic in two Asian populations examined: Japanese and Korean. In this study, using a collection of 910 SLE patients and 2360 controls from Chinese living in Hong Kong, analyzed by both genome-wide association and direct sequencing, we confirmed the association of the same risk alleles in ITGAM with the disease. These findings were further replicated in the Thai population with 278 patients and 383 ethnicity- and geography-matched controls. Subphenotype stratification analyses showed significantly more involvement of the gene in patients with renal nephritis and neurological disorders. Although our results support a pivotal role by rs1143679 (R77H) in disease association, our data also suggests an additional contribution from rs1143683, another non-synonymous polymorphism in this gene (A858V). Therefore, despite the low-allele frequencies of the risk alleles of the gene in our two Asian populations, ITGAM was confirmed to be a risk factor related to disease susceptibility and probably severe manifestations of SLE

Comparison of performance of the Assessment of Spondyloarthritis International Society, the European Spondyloarthropathy Study Group and the modified New York criteria in a cohort of Chinese patients with spondyloarthritis

Early diagnosis of spondyloarthritis (SpA) is essential as anti-tumor necrosis factor therapy can achieve significant symptomatic relief and control of disease activity. This study aims to compare the clinical characteristics, disease activity, and functional status of a Chinese cohort of SpA patients who were re-classified into ankylosing spondylitis (AS) patients fulfilling the modified New York (MNY) criteria, those with undifferentiated SpA (USpA) fulfilling the European Spondyloarthropathy Study Group (ESSG) classification criteria only (USpA/ESSG) and those who fulfill Assessment of SpondyloArthritis International Society (ASAS) only (USpA/ASAS). Disease activity was evaluated by Bath Ankylosing Spondylitis Disease Activity Index (BASDAI), severity of morning stiffness, patient global assessment, and C-reactive protein. Functional status was evaluated by Bath Ankylosing Spondylitis Functional Index (BASFI), modified Schober index, and dimension of chest expansion. One hundred and twenty-eight patients with disease duration of 16.3 ± 10.4 years were recruited. Patients in USpA/ESSG and USpA/ASAS were significantly younger (p = 0.01), had shorter disease duration (p < 0.01), and lower BASFI (p = 0.03) than established AS patients. All three groups have active disease with comparable BASDAI >3. BASFI correlated inversely with dimension of chest expansion and negatively modified Schober index in AS patients (p < 0.01) and modestly with BASDAI (r = 0.25, p < 0.01). BASFI correlated moderately with BASDAI in USpA/ESSG (r = 0.61, p < 0.01) but not with chest expansion or modified Schober index. Compared with established AS patients recognized by MNY criteria, patients fulfilling USpA defined by ESSG or ASAS criteria had earlier disease, as active disease and less irreversible functional deficit

Meta-analysis Followed by Replication Identifies Loci in or near CDKN1B, TET3, CD80, DRAM1, and ARID5B as Associated with Systemic Lupus Erythematosus in Asians

Systemic lupus erythematosus (SLE) is a prototype autoimmune disease with a strong genetic involvement and ethnic differences. Susceptibility genes identified so far only explain a small portion of the genetic heritability of SLE, suggesting that many more loci are yet to be uncovered for this disease. In this study, we performed a meta-analysis of genome-wide association studies on SLE in Chinese Han populations and followed up the findings by replication in four additional Asian cohorts with a total of 5,365 cases and 10,054 corresponding controls. We identified genetic variants in or near CDKN1B, TET3, CD80, DRAM1, and ARID5B as associated with the disease. These findings point to potential roles of cell-cycle regulation, autophagy, and DNA demethylation in SLE pathogenesis. For the region involving TET3 and that involving CDKN1B, multiple independent SNPs were identified, highlighting a phenomenon that might partially explain the missing heritability of complex diseases

Genome-Wide Association Study in Asian Populations Identifies Variants in ETS1 and WDFY4 Associated with Systemic Lupus Erythematosus

Systemic lupus erythematosus is a complex and potentially fatal autoimmune disease, characterized by autoantibody production and multi-organ damage. By a genome-wide association study (320 patients and 1,500 controls) and subsequent replication altogether involving a total of 3,300 Asian SLE patients from Hong Kong, Mainland China, and Thailand, as well as 4,200 ethnically and geographically matched controls, genetic variants in ETS1 and WDFY4 were found to be associated with SLE (ETS1: rs1128334, P = 2.33×10−11, OR = 1.29; WDFY4: rs7097397, P = 8.15×10−12, OR = 1.30). ETS1 encodes for a transcription factor known to be involved in a wide range of immune functions, including Th17 cell development and terminal differentiation of B lymphocytes. SNP rs1128334 is located in the 3′-UTR of ETS1, and allelic expression analysis from peripheral blood mononuclear cells showed significantly lower expression level from the risk allele. WDFY4 is a conserved protein with unknown function, but is predominantly expressed in primary and secondary immune tissues, and rs7097397 in WDFY4 changes an arginine residue to glutamine (R1816Q) in this protein. Our study also confirmed association of the HLA locus, STAT4, TNFSF4, BLK, BANK1, IRF5, and TNFAIP3 with SLE in Asians. These new genetic findings may help us to gain a better understanding of the disease and the functions of the genes involved

Oral health of Chinese people with systemic sclerosis

The aim was to study oral health status, salivary function, and oral features of Chinese people with Systemic Sclerosis (SSc). Chinese people with SSc attending a university specialist clinic were invited for a questionnaire survey and a clinical examination. Ethics approval was sought (UW 08-305). Gender- and age-matched individuals without SSc who attended a university dental hospital were recruited for comparison. Forty-two SSc patients with a mean age of 54.0 ± 12.2 were examined. This study found no Chinese people with systemic sclerosis were periodontally healthy and many (76%) had periodontal pockets despite most of them (93%) practiced daily tooth-brushing. They all had caries experience (DMFT = 10.5) and many (65%) had untreated decay. Mucosal telangiectasia was a common oral feature (80%). They had lower resting salivary flow rates (0.18 ± 0.17 ml/min vs. 0.31 ± 0.21 ml/min; p = 0.003) and pH values (6.90 ± 0.40 vs. 7.28 ± 0.31; p < 0.001) and reduced maximal mouth opening (40.1 ± 6.5 mm vs. 43.6 ± 7.0 mm) than people without SSc

Endothelial dysfunction and vascular disease in systemic sclerosis

published_or_final_versionMedicineMasterDoctor of Medicin