Project description:We previously reported that a synthetic Nod1 ligand, FK565, induced coronary arteritis in mice similar to Kawasaki disease. However, the molecular mechanisms underlying this site-specific inflammation have remained elusive. In this study, we found that CD11c+MHC class II+ cells accumulated in the heart of FK565-treated mice prior to arteritis development. We used microarray analysis to detail gene expression of CD11c+MHC class II+ cells. To obtain gene expression profile of CD11c+MHC class II+ cells, we isolated these cells from hearts of FK565-treated mice. Briefly, female mice at 8weeks age were administered 500 μg of FK565 subcutaneously at day 0 and day 3. At day6, murine hearts were removed and digested with collagenase. CD11c+MHC II+ cells were sorted as PI–CD45+Ly6G–NK1.1–CD11b+CD11c+MHC II+ using FACS Aria cell sorter (BD Biosciences). Sorted cells were subjected to RNA preparation. Two independent replicates from ten mice were made.
Project description:Over expression of MHC Class l protein in skeletal muscle causes myositis. Phenotype after expression in young mice is more severe. We performed gene expression profiling on young and adult mice after over expression of self MHC class l protein in skeletal muscle Muscle from young ( early) , adult (Late) and cntrol (control) mice , n=3 each group, was used for gene expression profiling
Project description:Over expression of MHC Class l protein in skeletal muscle causes myositis. Phenotype after expression in young mice is more severe. We performed gene expression profiling on young and adult mice after over expression of self MHC class l protein in skeletal muscle Overall design: Muscle from young ( early) , adult (Late) and cntrol (control) mice , n=3 each group, was used for gene expression profiling
Project description:To further understand the early molecular events in antibody mediated chronic inflammation of the lungs, we utilized a murine Obliterative Airway Disease (OAD) model of native lungs where Abs to MHC class I induces bronchiolar obliteration and fibrosis. This study identified a unique transcriptional profile with a set of genes that were stimulated while another set of genes that were repressed in anti MHC I treated lungs compared to that of isotype treated lungs. Lung transplantation (LTx) is an accepted clinical intervention for various intractable pulmonary dysfunctions. However, de novo Abs to mismatched donor HLA (DSA) predispose lung-grafts to chronic rejection, Bronchiolitis Obliterans Syndrome (BOS). The anti-MHC induced mouse OAD model, the only available preclinical model for study of early pathogenesis following MHC ligation, produces lesions analogous to BOS. Overall design: Induction of gene expression was measured following intrabronchial administration of anti-MHC class I (H-2Kb) Ab at 200μg/mouse. Samples were collected on days 1, 2, 3 and 6 post-Ab administration. Isotype matched control Ab was administered at an equivalent dose and gene expression profile in isotype treated group served as the base line expression and was compared with that of anti-H-2Kb treated group. A total of 24 samples were processed with two treatment groups (anti-H-2Kb, isotype), four time points (days 1, 2, 3 and 6) and three biological replicates.
Project description:MHC class II (MHCII) genes are transactivated by the NOD-like receptor (NLR) factor CIITA, which is recruited to SXY regulatory modules of MHC promoters via a DNA-binding “enhanceosome” complex. NLRC5, another NLR protein, was recently found to control transcription of MHC class I (MHCI) genes. However, detailed understanding of its target-gene specificity and mechanism of action remained lacking. We therefore performed ChIP-sequencing experiments to gain comprehensive information on NLRC5-transactivated genes. In addition to classical MHCI genes, we identified novel NLRC5-targets exclusively in the H2-Q and H2-T regions of the MHCI locus, among which the best targets were found. Investigation of cells lacking the MHCII-enhanceosome factor RFX5 demonstrated its strict requirement for NLRC5 recruitment to the SXY module conserved in MHCI genes. Furthermore, patient-derived B cell lines deficient in RFX5, RFXAP, and RFXANK corroborated importance of the enhanceosome for MHCI transactivation. Although sharing similar SXY modules and common DNA-binding factors, CIITA and NLRC5 regulate distinct genes, as shown here using double-deficient Nlrc5-/-CIIta-/- mice. The identification of sequences occupied by NLRC5 in vivo allowed us to define a unique consensus motif for its recruitment, which diverges from that used by CIITA. Our results thus broaden our knowledge on transcriptional activity of NLRC5, highlighting its remarkable selectivity for genes encoding MHCI or related proteins and providing insights into the specificity of its recruitment. Analysis of Nlrc5 binding sites in T-cells