Attenuation of regulatory T cell function by type I IFN signaling in an MDA5 gain-of-function mutant mouse model

Melanoma differentiation-associated gene 5 (MDA5) is an essential viral double-stranded RNA sensor to trigger antiviral immune responses, including type I interferon (IFN) induction. Aberrant activation of this viral sensor is known to cause autoimmune diseases designated as type I interferonopathies. However, the cell types responsible for these diseases and the molecular mechanisms behind their onset and development are still largely unknown. In this study, we revealed the attenuation of regulatory T cell (Treg) function by type I IFN signaling in a mouse model expressing a gain-of-function MDA5 G821S mutant. We found that experimental colitis induced by adoptive transfer of naïve T cells in Rag2⁻/⁻ mice was rescued by simultaneous transfer of Tregs from wild-type but not from the MDA5 mutant mice. Type I IFN receptor deficiency in the MDA5 mutant mice recovered the suppressive function of MDA5 mutant Tregs. These results suggest that constitutive MDA5 and type I IFN signaling in Tregs decreases the suppressive function of Tregs, potentially contributing to the onset and exacerbation of autoimmune disorders in interferonopathies

The hierarchical structure of the perichromosomal layer comprises Ki67, ribosomal RNAs, and nucleolar proteins

The perichromosomal layer (PCL) is a structure that surrounds mitotic chromosomes, found in both animal and plant cells. It comprises various proteins and RNAs, mainly derived from the nucleolus. Several functions for the PCL have been suggested; however, the mechanism of PCL organization during mitosis remains unclear. The localization of several nucleolar proteins to the PCL is reportedly dependent on pre-ribosomal RNAs and the marker of proliferation, Ki67, which is a major PCL-localized protein. Here we demonstrate that, although the removal of pre-ribosomal RNAs from the PCL causes PCL delocalization of several nucleolar proteins, it does not affect the localization of Ki67. Conversely, Ki67 depletion results in the dissociation of both pre-ribosomal RNAs and nucleolar proteins from the PCL, which indicates that Ki67 is required for the PCL accumulation of pre-ribosomal RNAs, to which several nucleolar proteins are associated. Given these findings, we propose a model for PCL organization that comprises three essential layers: the scaffolding protein Ki67, pre-ribosomal RNAs for linkage, and outer nucleolar proteins

Phos-tag-based approach to study protein phosphorylation in the thylakoid membrane

Protein phosphorylation is a fundamental post-translational modification in all organisms. In photoautotrophic organisms, protein phosphorylation is essential for the fine-tuning of photosynthesis. The reversible phosphorylation of the photosystem II (PSII) core and the light-harvesting complex of PSII (LHCII) contribute to the regulation of photosynthetic activities. Besides the phosphorylation of these major proteins, recent phosphoproteomic analyses have revealed that several proteins are phosphorylated in the thylakoid membrane. In this study, we utilized the Phos-tag technology for a comprehensive assessment of protein phosphorylation in the thylakoid membrane of Arabidopsis. Phos-tag SDS-PAGE enables the mobility shift of phosphorylated proteins compared with their non-phosphorylated isoform, thus differentiating phosphorylated proteins from their non-phosphorylated isoforms. We extrapolated this technique to two-dimensional (2D) SDS-PAGE for detecting protein phosphorylation in the thylakoid membrane. Thylakoid proteins were separated in the first dimension by conventional SDS-PAGE and in the second dimension by Phos-tag SDS-PAGE. In addition to the isolation of major phosphorylated photosynthesis-related proteins, 2D Phos-tag SDS-PAGE enabled the detection of several minor phosphorylated proteins in the thylakoid membrane. The analysis of the thylakoid kinase mutants demonstrated that light-dependent protein phosphorylation was mainly restricted to the phosphorylation of the PSII core and LHCII proteins. Furthermore, we assessed the phosphorylation states of the structural domains of the thylakoid membrane, grana core, grana margin, and stroma lamella. Overall, these results demonstrated that Phos-tag SDS-PAGE is a useful biochemical tool for studying in vivo protein phosphorylation in the thylakoid membrane protein

Prognostic value of RKIP and p-ERK in gastric cancer

<p>Abstract</p> <p>Background</p> <p>The mitogen-activated protein kinase (MAPK) signaling pathway participates in several steps of tumour development and is considered a prominent therapeutic target for the design of chemotherapeutic agents. We evaluated the expressions of extracellular signal-regulated kinase (ERK), mitogen-activated protein kinase (MEK), an upstream regulator of ERK, and Raf kinase inhibitor protein (RKIP), and investigated correlations of these expressions with clinicopathological features and outcomes in gastric cancer.</p> <p>Methods</p> <p>Tumour samples were obtained from 105 patients with gastric adenocarcinomas who underwent radical gastrectomy. The expressions of phosphorylated ERK (p-ERK), phosphorylated MEK (p-MEK), and RKIP were analysed by immunohistochemical staining.</p> <p>Results</p> <p>Expression of RKIP, p-MEK, and p-ERK was found in 69 (66%), 54 (51%), and 64 (61%) of all tumours, respectively. RKIP expression negatively correlated with the depth of invasion (p < 0.001), lymph node involvement (p = 0.028), and Union for International Cancer Control (UICC) stage (p = 0.007). RKIP expression was associated with significantly longer relapse-free survival (RFS) (p = 0.0033), whereas p-MEK was not (p = 0.79). Patients with p-ERK expression had slightly, but not significantly shorter RFS than those without such expression (p = 0.054). Patients with positive p-ERK and negative RKIP expression had significantly shorter RFS than the other patients (p < 0.001). The combination of RKIP and p-ERK expression was an independent prognostic factor (hazard ratio, 2.4; 95% confidence interval, 1.3 - 4.6; p = 0.008).</p> <p>Conclusions</p> <p>Our results demonstrated that loss of RKIP was associated with tumour progression and poor survival. Negative RKIP expression combined with positive p-ERK expression was an independent predictor of poor outcomes in patients with gastric cancer.</p

Organellar Glue: A Molecular Tool to Artificially Control Chloroplast–Chloroplast Interactions

細胞小器官を接着する新技術「オルガネラグルー」を開発 --オルガネラ間コミュニケーションの操作に期待--. 京都大学プレスリリース. 2022-09-30.Organelles can physically interact to facilitate various cellular processes such as metabolite exchange. Artificially regulating these interactions represents a promising approach for synthetic biology. Here, we artificially controlled chloroplast–chloroplast interactions in living plant cells with our organelle glue (ORGL) technique, which is based on reconstitution of a split fluorescent protein. We simultaneously targeted N-terminal and C-terminal fragments of a fluorescent protein to the chloroplast outer envelope membrane or cytosol, respectively, which induced chloroplast–chloroplast interactions. The cytosolic C-terminal fragment likely functions as a bridge between two N-terminal fragments, thereby bringing the chloroplasts in close proximity to interact. We modulated the frequency of chloroplast–chloroplast interactions by altering the ratio of N- and C-terminal fragments. We conclude that the ORGL technique can successfully control chloroplast–chloroplast interactions in plants, providing a proof of concept for the artificial regulation of organelle interactions in living cells

Surgical Treatment for Atlanto-Occipital Subluxation due to Destructive Spondyloarthropathy in a Patient Undergoing Long-Term Hemodialysis

Destructive spondyloarthropathy (DSA) has been reported in patients undergoing long-term hemodialysis. Cervical spinal lesions, including those of the upper cervical spine, are reported to be some of the most common. To our knowledge, we report for the first time, a case of atlanto-occipital subluxation requiring surgical treatment due to severe myelopathy and nuchal pain in a patient undergoing long-term hemodialysis. The patient was a 66-year-old woman who had undergone hemodialysis for 40 years. She visited our hospital due to an acute progression of gait disturbance and severe nuchal pain. Computed tomography showed posterior subluxation of the atlanto-occipital joints. DSA was also observed in the lower cervical spine. Magnetic resonance imaging showed spinal canal stenosis at both the upper and lower cervical levels. We performed Oc-C7 fixation, C1 laminectomy, and C3-C7 laminoplasty. We first recognized that the atlanto-occipital subluxation was caused by the extremely long-term, in this case, 40 years, hemodialysis

Surgical Outcome for Hemodialysis-Related Upper Cervical Lesions

Study DesignA retrospective study.PurposeTo investigate the surgical outcome for hemodialysis-related upper cervical lesions.Overview of LiteratureSurgical outcome of lower cervical lesions in patients undergoing hemodialysis has been reported. However, surgical outcome for upper cervical lesions in hemodialysis patients is unclear.MethodsUpper cervical lesions in nine patients undergoing hemodialysis were surgically treated. Mean age at surgery was 61.6 years (range, 52-68 years), and the mean follow-up period was 45.4 months (range, 2-98 months). Patients had undergone hemodialysis for an average of 25.3 years (range, 16-40 years) at surgery. Seven patients with destructive spondyloarthropathy (DSA) of the upper cervical spine were treated with atlantoaxial or occipitocervical fixation. Two patients with retro-odontoid pseudotumors were treated with C1 posterior arch resection alone. Japanese Orthopedic Association (JOA) scores for cervical myelopathy, postoperative complications, postoperative radiography, and preoperative and postoperative occipital pain were evaluated.ResultsMean preoperative and postoperative JOA score was 3.7 and 8.1, respectively. The seven patients with DSA had severe preoperative occipital pain that disappeared postoperatively. Postoperative radiography showed solid bone union in DSA cases and no instability in pseudotumor cases.ConclusionsSatisfactory surgical outcome was observed for hemodialysis-related upper cervical lesions