Project description:A mutation in the Signal Transducer and Activator of Transcription 3 (STAT3) has been linked to incidence of Autosomal Dominant Hyper Immunoglobulin E Syndrome (AD-HIES), a disease characterized by elevated serum IgE antibody. However, how this genetic mutation leads to the phenotype has not been fully understood. We investigated the specific role of STAT3 in the germinal center (GC) B cells and plasma cells for IgE class switching. Through the use of STAT3 conditional knockout mice in a Th2-type immunization model, we demonstrated that CD2-Cre driven STAT3 cKO mice showed elevated IgE and decreased IgG1 in the serum, and a reduction in GC formation. Within the GC, IgG1+ GC B cells were decreased while IgE+ GC B cells were more prevalent. Additionally, these mice exhibited reduced IgG1 and elevated IgE populations of antibody-producing plasma cells. Subsequent experiments using a CD19-Cre B-cell specific cKO mouse established this effect to be B-cell intrinsic. Transcription factors critical for GC and plasma cell differentiation, including Bcl-6 and Aicda, were shown to function as downstream signals of STAT3 regulation. Further ChIP-seq analysis revealed that many genes including Bcl3 and Crtc2 were among the direct STAT3 regulated targets. Mice with STAT3 deficiency in B cells also demonstrated an increase in lung inflammation when used in an asthma-like disease model. This model suggests a negative role for STAT3 in regulating class switching of the GC B cells from the IgG1 to the IgE producing state, which may serve as a therapeutic target for treatment of AD-HIES and other immune disorders.
Project description:In vitro, both 5PP-InsP5 (IP7) and 1,5(PP)2InsP4 (IP8) can pyrophosphorylate peptides and proteins. It is unclear, which inositol pyrophosphate species is responsible for installing endogenous pyrophosphorylation in cells. In order to test this, we used HCT116 cells deficient in IP8 synthesis. Both enzymes that convert IP7 to IP8 are knocked out in this cell line. Importantly, this cell line does overaccumulates IP7, complicating the analysis.
We found no obvious difference in the number of observable pyrophosphorylation sites between the knockout and wildtype, suggesting that either IP7 or both molecules are responsible for creating pyrophosphoproteins in cells.
Project description:In situ synthesized oligo arrays, U74Av2, from Affymetrix were used to measure differential gene expression in RNA samples generated from the liver of Nrf2 knockout and wildtype mice at 5 month age. Total RNAs from two Nrf2 knockout or wildtype littermates were analyzed separately. There are two replicates (GSM 13431, 13435) for the female Nrf2 wildtype group, two replicates (GSM 13439, 13441) for the male Nrf2 wildtype group, two replicates (GSM 13436, 13437) for the female Nrf2 knockout group, and two replicates (GSM 13438, 13440) for the male Nrf2 knockout group. Keywords: parallel sample
Project description:In situ synthesized oligo arrays, U74Av2, from Affymetrix were used to measure differential gene expression in RNA samples generated from the spleen of Nrf2 knockout and wildtype female mice at 5 month age. Total RNAs from two Nrf2 knockout or wildtype littermates were analyzed separately. There are two replicates (GSM 13470, 13472) for the female Nrf2 wildtype group and two replicates (GSM 13467, 13469) for the female Nrf2 knockout group. Keywords: parallel sample
Project description:We report here the deep sequencing of the mRNA from peritoneal exudate cells (macrophages) purified from wildtype or Ptpn1 (PTP1B) knockout mice, either treated or untreated with IL-10. In periotenal macrophages IL-10 activates the transcription factor STAT3 to execute and anti-inflammatory gene expression programme. The tyrosine phosphatase PTPN1 targets STAT3 for dephosphorylation and leads to the deactivation of STAT3. In this study we examined the role of PTP1B in controlling the normal homeostatic level phosphorylation of STAT3 by comparing the IL-10/STAT3-mediated anti-inflammatory gene expression programme. We find that loss of PTP1B leads to an up-regulation of the activity of STAT3, both at the level of phosphorylation and also in enhanced expression of anti-inflammatory gene products. RNA-seq of wildtype and Ptpn1 (PTP1B) knockout mouse peritoneal macrophages, treated or untreated with IL-10
Project description:In this study we have explored the effects of ERdj5 chaperone protein deletion in the proteome of 129SV mouse submandibular salivary glands. This knockout mouse model recapitulates many aspects of Sjögren’s Syndrome, including salivary gland inflammatory infiltrations, anti-nuclear autoantibodies and others (Apostolou et al., Front.Immunol.2019;10,506). Additionally, for 12SV wildtype animals we explore the proteome profile of the extensive sexual dimorphism observed in the submanidular glands of mice.