Interferon (IFN) regulatory factor 7 (IRF7) plays a key role in the production of IFN‐α in response to viral infection, and phosphorylation at IRF7 C‐terminal serine sites is prelude to its function. However, phosphatases that negatively regulate IRF7 phosphorylation and activity have not been reported. In this study, we have identified a conserved protein phosphatase 1 (PP1)‐binding motif in human and mouse IRF7 proteins, and shown that PP1 physically interacts with IRF7. Exogenous expression of PP1 subunits (PP1α, β, or γ) ablates IKKε‐stimulated IRF7 phosphorylation and dramatically attenuates IRF7 transcriptional activity. Inhibition of PP1 activity significantly increases IRF7 phosphorylation and IRF7‐mediated IFN‐α production in response to Newcastle disease virus (NDV) infection or Toll‐like receptor 7 (TLR7) challenge, leading to impaired viral replication. In addition, IFN treatment, TLR challenges and viral infection induce PP1 expression. Our findings disclose for the first time a pivotal role for PP1 in impeding IRF7‐mediated IFN‐α production in host immune responses.

No description available.

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第18461号 / 医博第3916号 / 新制||医||1005(附属図書館) / 31339 / 京都大学大学院医学研究科医学専攻 / (主査)教授 村井 俊哉, 教授 竹内 理, 教授 長田 重一 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Green open access: Authors can share their research in a variety of different ways and Elsevier has a number of green open access options available. We recommend authors see our green open access page for further information (http://elsevier.com/greenopenaccess). Authors can also self-archive their manuscripts immediately and enable public access from their institution's repository after an embargo period. This is the version that has been accepted for publication and which typically includes author-incorporated changes suggested during submission, peer review and in editor-author communications. Embargo period: For subscription articles, an appropriate amount of time is needed for journals to deliver value to subscribing customers before an article becomes freely available to the public. This is the embargo period and it begins from the date the article is formally published online in its final and fully citable form. This journal has an embargo period of 12 months. 詳細は以下のアドレスを参照https://www.elsevier.com/journals/immunology-letters/0165-2478?generatepdf=true / 京都大学 / 0048 / 新制・課程博士 / 博士(生命科学) / 甲第19870号 / 生博第351号 / 新制||生||46(附属図書館) / 32906 / 京都大学大学院生命科学研究科高次生命科学専攻 / (主査)教授 稲葉 ｶﾖ, 教授 米原 伸, 教授 杉田 昌彦 / 学位規則第4条第1項該当 / Doctor of Philosophy in Life Sciences / Kyoto University / DFAM

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第20084号 / 医博第4177号 / 新制||医||1018(附属図書館) / 33200 / 京都大学大学院医学研究科医学専攻 / (主査)教授 生田 宏一, 教授 山田 亮, 教授 椛島 健治 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

No description available.

Epstein-Barr Virus (EBV) latent infection is associated with a variety of lymphomas and carcinomas. Interferon (IFN) Regulatory Factors (IRFs) are a family of transcription factors, among which IRF7 is the “master” regulator of type I IFNs (IFN-I) that defends against invading viruses. Robust IFN-I responses require a positive feedback loop between IRF7 and IFN-I. In recent years, we have discovered that IRF7 is significantly induced and activated by the principal EBV oncoprotein--Latent Membrane Protein 1 (LMP1); however, IRF7 fails to trigger robust IFN-I responses in EBV latency. We believe this intriguing finding is critical for EBV latency and oncogenesis, yet the underlying mechanism of this paradoxical phenomenon remains unclear. It is well known that tyrosine phosphorylation of most components of the IFN-I Jak-STAT pathway is essential for its signaling transduction. Thus, we have performed phosphotyrosine proteomics. We have found that the IFN-I Jak-STAT pathway is inactive due to the attenuated STAT2 activity, whereas the IFN-II Jak-STAT pathway is constitutively active, in EBV latency. We further confirmed these results by immunoblotting. This pilot study provides valuable information for the critical question regarding how the IRF7-mediated IFN-I response is evaded by EBV in its latency, and will prompt us to elucidate the underlying mechanisms.

No description available.

Over the past decades, information has accumulated concerning the mechanism how an exterior signal induced by ligand on neighboring cells is transmitted to the nucleus through the Notch receptor and the cellular effects of Notch signaling on the regulation of differentiation, proliferation and apoptosis in many cell types. However, the function and the mechanism of Notch signaling in peripheral T cells still remains to be addressed. Therefore, we asked whether Notch1 is involved in CD8+ cytolytic effector T cell (CTLs) maturation and effector functions and how Notch1 exerts its cellular function in the nucleus and in the cytoplasm. The maturation of naïve CD8+ T cells into CTLs is a critical feature of a functional adaptive immune system. Development of CTLs depends, in part, upon the expression of the transcriptional regulator, Eomesodermin (EOMES), which is thought to regulate the expression of two key effector molecules, perforin and granzyme B. In addition, the data from previous studies in our lab showed that Notch signaling results in the activation of NF-κB, IFN-γ secretion and cell proliferation both in CD4+ and CD8+ T cells. Therefore, we hypothesized that Notch1 may be involved in CD8+ T cell maturation and effector function. We observed that Notch1 regulates the expression of EOMES, perforin and granzyme B through direct binding to the promoters of these crucial effector molecules. By abrogating Notch signaling, both biochemically as well as genetically, we conclude that Notch activity mediates CTL development through direct regulation of EOMES, perforin and granzyme B. We further investigated the molecular steps leading to the formation of intracellular Notch1 (N1ICD)/CSL (also known as CBF1/RBP-Jκ in mammals; Suppressor of Hairless in Drosophila; and Lag-1 in C. elegans) with other co-factors in target promoters of Notch1 signaling. We proposed that the association of two nuclear complexes with N1ICD controls the transcription of genes, allowing the development of effector CTL in the immune system. Recent studies proposed a model where Notch1 colocalizes with CD4, a component of the immune synapse, upon T cell stimulation and directly associates with p56Lck and CD28, as well as PI3K. However, the link between Notch and the TCR signalosome needed further investigation. We found that Notch1 functions as a scaffold, associated with the cytosolic components, Carma1, Bcl10, PKCθ and the IKK complex upon TCR stimulation, leading to the activation of NF-κB and IL-2 production. We further showed that the N-terminal region of N1ICD is essential for interaction with Carma1 and that deficiency of Notch1 abolishes the nuclear binding of NF-κB on the il- 2 promoter, leading to reduced IL-2 production.

Background: The p53-up-regulated modulator of apoptosis (PUMA) protein is a pro-apoptotic, BH3-only member of the BCL2 family of effector proteins responsible for promoting organized cell death. PUMA is required for resolution of pneumococcal pneumonia in mice, as mice deficient of PUMA exhibit greater numbers of S. pneumoniae CFU within tissues and higher mortality rates than observed in Puma+/+ mice. Methods: Puma+/+ and Puma-/- mice were intranasally challenged with TIGR4 pneumococcus and sacrificed 24 h post-infection. Differences in cytokine levels from blood and whole lung tissue were detected by MILLIPLEX MAP Mouse Cytokine/Chemokine Magnetic Bead Panel. Lung transcriptomes from Puma+/+ and Puma-/- mice were prepared from total lung RNA using NEBNext Poly(A) mRNA Magnetic Isolation Module and NEBNext Ultra RNA Library Prep Kit for Illumina. Libraries were read by Illumina NovaSeq and transcript reads were referenced to Mus musculus. Results: Puma-/- mice exhibited significant differences in G-CSF, GM-CSF, IFN-gamma, IL-1-alpha and -beta, -6, -9, -10, -12 (p40 and p70), -13, and -17, IP-10, KC, MCP-1, MIP- iv 1alpha and -beta, MIP-2, RANTES, and TNF-alpha compared to Puma+/+ mice. Puma-/- lungs exhibited higher levels of IL-12, IFN-gamma, and IP-10. Loss of PUMA also resulted in expression of the pro-angiogenic genes Adam19 and Neurexin2. Additionally, Puma+/+ and Puma-/- mice displayed similar levels of colonization, but Puma-/- mice were more susceptible to subsequent dissemination to the lungs and blood. Conclusion: Polymorphonuclear cells (PMNs) were previously demonstrated to be one of the innate cell types responsible for Puma-dependent resolution of pneumococcal pneumonia in mice. Observations reported here suggest that this resolution is propelled by suppressing the inflammatory response via the inhibition of IL-12/IFN-gamma/IP-10 pro-inflammatory axis. Pulmonary tissue transcriptomic analysis also suggests PUMA-dependent positive regulation of homeostatic control of pulmonary vasculature, smooth muscle innervation, and maintenance of the interstitium. Gene ontological analysis further demonstrated Puma's modulatory role in Type I and II IFN signaling. For the first time, we report Puma's regulatory effects on pro-inflammatory cytokine signaling and gene expression during pneumococcal pneumonia.