A Thpok-Directed Transcriptional Circuitry Promotes Bcl6 and Maf Expression to Orchestrate T Follicular Helper Differentiation.

The generation of high-affinity neutralizing antibodies, the objective of most vaccine strategies, occurs in B cells within germinal centers (GCs) and requires rate-limiting "help" from follicular helper CD4+ T (Tfh) cells. Although Tfh differentiation is an attribute of MHC II-restricted CD4+ T cells, the transcription factors driving Tfh differentiation, notably Bcl6, are not restricted to CD4+ T cells. Here, we identified a requirement for the CD4+-specific transcription factor Thpok during Tfh cell differentiation, GC formation, and antibody maturation. Thpok promoted Bcl6 expression and bound to a Thpok-responsive region in the first intron of Bcl6. Thpok also promoted the expression of Bcl6-independent genes, including the transcription factor Maf, which cooperated with Bcl6 to mediate the effect of Thpok on Tfh cell differentiation. Our findings identify a transcriptional program that links the CD4+ lineage with Tfh differentiation, a limiting factor for efficient B cell responses, and suggest avenues to optimize vaccine generation.

FBXO11 constitutes a major negative regulator of MHC class II through ubiquitin-dependent proteasomal degradation of CIITA.

Major histocompatibility complex (MHC) class I and II molecules play critical roles in the activation and regulation of adaptive immunity through antigen presentation to CD8+ and CD4+ T cells, respectively. Strict regulation of MHC expression is critical for proper immune responses. CIITA (MHC class II transactivator), an NLR (nucleotide-binding domain, leucine-rich-repeat containing) protein, is a master regulator of MHC class II (MHC-II) gene transcription. Although it has been known that CIITA activity is regulated at the transcriptional and protein levels, the mechanism to determine CIITA protein level has not been elucidated. Here, we show that FBXO11 is a bona fide E3 ligase of CIITA and regulates CIITA protein level through ubiquitination-mediated degradation. A nonbiased proteomic approach for CIITA-binding protein identified FBXO11, a member of the Skp1-Cullin-1-F-box E3 ligase complex, as a binding partner of CIITA but not MHC class I transactivator, NLRC5. The cycloheximide chase assay showed that the half-life of CIITA is mainly regulated by FBXO11 via the ubiquitin-proteasome system. The expression of FBXO11 led to the reduced MHC-II at the promoter activity level, transcriptional level, and surface expression level through downregulation of CIITA. Moreover, human and mouse FBXO11-deficient cells display increased levels of MHC-II and related genes. In normal and cancer tissues, FBXO11 expression level is negatively correlated with MHC-II. Interestingly, the expression of FBXO11, along with CIITA, is associated with prognosis of cancer patients. Therefore, FBXO11 is a critical regulator to determine the level of MHC-II, and its expression may serve as a biomarker for cancer.

Downmodulation of tumor suppressor p53 by T cell receptor signaling is critical for antigen-specific CD4(+) T cell responses.

Antigen specificity is critical in immune response and requires integration of antigen-specific signals with antigen-nonspecific signals such as those provided by cytokines. The mechanism integrating these pathways is incompletely understood. We report here that antigen-specific proliferative responses of CD4(+) T cells required downmodulation of tumor suppressor p53. In the absence of T cell receptor (TCR) signal, IL-2 induced sustained increase in p53 protein, which prevented proliferative responses despite strong signaling through the IL-2 receptor. In contrast, TCR signaling resulted in early termination of p53 protein expression by decreasing p53 mRNA as well as strong transcriptional induction of the p53-regulating protein Mdm2. Downmodulation of p53 in response to antigen stimulation was in fact critical for antigen-specific T cell proliferation, and preventing p53 degradation by inhibiting Mdm2 resulted in sustained p53 protein and prevented antigen-specific T cell proliferation. It is thus termination of p53 by TCR signaling that allows proliferative responses, enforcing antigen specificity.

TCR Repertoires of Thymic Conventional and Regulatory T Cells: Identification and Characterization of Both Unique and Shared TCR Sequences.

Thymic regulatory T cells (tTreg) are critical in the maintenance of normal T cell immunity and tolerance. The role of TCR in tTreg selection remains incompletely understood. In this study, we assessed TCRα and TCRß sequences of mouse tTreg and thymic conventional CD4+ T cells (Tconv) by high-throughput sequencing. We identified αß TCR sequences that were unique to either tTreg or Tconv and found that these were distinct as recognized by machine learning algorithm and by preferentially used amino acid trimers in αß CDR3 of tTreg. In addition, a proportion of αß TCR sequences expressed by tTreg were also found in Tconv, and machine learning classified the great majority of these shared αß TCR sequences as characteristic of Tconv and not tTreg. These findings identify two populations of tTreg, one in which the regulatory T cell fate is associated with unique properties of the TCR and another with TCR properties characteristic of Tconv for which tTreg fate is determined by factors beyond TCR sequence.

Neighborhood-level socioeconomic factors moderate the association between physical activity and relative age effect: a cross-sectional survey study with Japanese adolescents.

BACKGROUND: Relative age effect is defined as a phenomenon where children born early generally perform better than children born later in the same cohort. Physical activity is an important factor that might be influenced by the relative age effect. Socioeconomic factors (e.g., parent's income, education level) are also associated with the adolescent's physical activity. However, no existing study has examined whether socioeconomic factors moderate the relative age effect on the adolescent's physical activity. This study aims to clarify whether and how birth month and socioeconomic factors relate to organized sports and physical activity among adolescents in Japan. METHODS: We conducted a questionnaire survey targeting 21,491 adolescents who live in a widespread neighborhood. We included 8102 adolescents (4087 males and 4015 females: mean age 13.1 ± 1.4) in the analysis. Based on the participants' birth months, we divided them into four groups (April to June, July to September, October to December, January to March). We asked participants to report their organized sports participation. Using the International Physical Activity Questionnaire for Japanese Early Adolescents, we identified their moderate to vigorous physical activity (MVPA). Neighborhood-level socioeconomic factors (areal deprivation, average annual income, education level) were analyzed based on national surveys, such as the population census. We performed multilevel logistic and linear regression analysis for organized sports participation and MVPA, respectively. Moreover, a simple slope analysis was implemented if the interaction between birth month and socioeconomic factor was significant in the multilevel linear regression analysis. RESULTS: Among males, relatively younger adolescents (adolescents who were born later in the same grade) were less likely to participate in organized sports activites (OR=0.90, 95% CI 0.82-0.97, p<0.05), while both males and females engaged in less MVPA (b=-0.54, b=-0.25, p< 0.01, respectively). We observed an interaction between birth month and socioeconomic factors. Among males in low-income neighborhoods, and females in more deprived neighborhoods, relatively younger adolescents engaged in less MVPA. CONCLUSIONS: Socioeconomic factors moderate the relative age effect on adolescents' physical activity. The relative age effect on adolescents' physical activity might be more likely to appear among adolescents from socioeconomically disadvantaged neighborhoods.

Association Between Age of Achieving Gross Motor Development Milestones During Infancy and Body Fat Percentage at 6 to 7 Years of Age.

OBJECTIVES: The later achievement of gross motor milestones during infancy is associated with adiposity in early childhood. However, the associations between gross motor development and adiposity after entering primary school are unclear. This study examined the associations between the ages at which six gross motor milestones were achieved and adiposity during early school years. METHODS: This retrospective study was conducted in 2012 and 2013. Data were collected from 225 first-grade primary school children (mean age, 6.9 years; 39% girls). Adiposity was assessed using dual-energy X-ray absorptiometry and expressed as body fat percentage. Data describing the ages of achieving six gross motor milestones (holding head up, sitting, crawling, standing supported, walking supported, and independent walking) were obtained from the Maternal and Child Health Handbooks. RESULTS: Mean body fat percentage was 21.7%. Multiple linear regression analyses revealed that later ages of achieving crawling (p < .001 [95% confidence interval: 0.33-1.16]), standing supported (p < .001 [95% confidence interval: 0.64-1.65]), and walking supported [p = .013 (95% confidence interval: 0.13-1.07)] were associated with increased fat. However, the ages of achieving holding head up (p = .053), sitting (p = .175), and independent walking (p = .736) were not statistically significant. CONCLUSIONS: Achieving crawling, standing supported, and walking supported later predict increased body fat when aged 6-7 years. The practice of observing gross motor milestone achievements may allow early targeted interventions to optimize body composition before beginning school and thereby, potentially prevent childhood obesity.

Does earlier acquisition of motor competence promote pubertal physical activity in Japanese elementary school children: A 4-year follow-up study.

This 4-year follow-up study investigated which profile of motor competence (MC) in early childhood more strongly correlated with moderate-to-vigorous physical activity (MVPA) in early pubescence. A total of 247 first-grade children (152 boys) aged 6 to 7 years old participated in the baseline measurement. MVPA was measured using an accelerometer annually until the fifth grade. Body fat percentage was assessed using dual-energy X-ray absorptiometry at baseline. MC included fundamental movement skills (FMS) and motor performance, which were assessed in terms of locomotor and object control. FMS was assessed by the Test of Gross Motor Development-2, while motor performance (locomotor: run, hop, jump, and side-step, object control: throw, kick, and hand dribble) was assessed by Japanese Physical Fitness and Motor Abilities Test at baseline. Multiple regression analysis showed that locomotor and object control performance were significant predictors of MVPA in later grades in boys but not in girls. These associations between locomotor performance and MVPA mainly remained significant after controlling for body fat and MVPA at baseline. FMS in both genders was not a significant predictor of MVPA in early pubescence. This study suggests that appropriate strategies for increasing MVPA from prepubescence to early pubescence may differ between boys and girls.

Measuring Thymic Clonal Deletion at the Population Level.

Clonal deletion of T cells specific for self-antigens in the thymus has been widely studied, primarily by approaches that focus on a single receptor (using TCR transgenes) or a single specificity (using peptide-MHC tetramers). However, less is known about clonal deletion at the population level. In this article, we report an assay that measures cleaved caspase 3 to define clonal deletion at the population level. This assay distinguishes clonal deletion from apoptotic events caused by neglect and approximates the anatomic site of deletion using CCR7. This approach showed that 78% of clonal deletion events occur in the cortex in mice. Medullary deletion events were detected at both the semimature and mature stages, although mature events were associated with failed regulatory T cell induction. Using this assay, we showed that bone marrow-derived APC drive approximately half of deletion events at both stages. We also found that both cortical and medullary deletion rely heavily on CD28 costimulation. These findings demonstrate a useful strategy for studying clonal deletion within the polyclonal repertoire.

Changes in disease characteristics of primary biliary cholangitis: An observational retrospective study from 1982 to 2016.

AIM: Disease characteristics of primary biliary cholangitis have changed recently. However, detailed studies on the subject have been limited. Therefore, we aimed to clarify disease characteristics of patients with recent primary biliary cholangitis using the cohort from Niigata University and 21 affiliated hospitals. METHODS: Overall, 508 patients were enrolled in this study from 1982 to 2016, divided into three cohorts according to their year of diagnosis: ≤1999, 2000-2009 and ≥2010. We compared differences in clinical characteristics, response to ursodeoxycholic acid and prognosis. RESULTS: The male-to-female ratio increased incrementally from 1:16.4 (≤1999) to 1:3.8 (≥2010) (P < 0.001). In women, the median age at diagnosis increased incrementally from 54.0 years (≤1999) to 60.5 years (≥2010) (P < 0.001) and serum albumin decreased gradually (P = 0.001), which might have affected the increase in the Fibrosis-4 Index and albumin-bilirubin score. The ursodeoxycholic acid response rate according to the Barcelona criteria increased incrementally from 26.7% (≤1999) to 78.4% (≥2010) (P < 0.010), and those according to other criteria (Paris-I, Rotterdam and Toronto) were approximately ≥80% in all cohorts. Ten-year survival rate in the ≤1999 and 2000-2009 cohorts were 98.6% and 95.6%, respectively. These earlier cohorts were also characterized by a higher rate of asymptomatic state and mild histology (83.5% [≤1999] and 84.7% [2000-2009], and 93.6% [≤1999] and 91.1% [2000-2009]). CONCLUSIONS: Patients with primary biliary cholangitis were characterized by older age at diagnosis and an increase in male to female ratio as well as higher response rates of ursodeoxycholic acid and longer survival, resulting from the early recognition of primary biliary cholangitis.

Role of intracellular zinc in molecular and cellular function in allergic inflammatory diseases.

Zinc is an essential micronutrient in human body and a vital cofactor for the function of numerous proteins encoded by the human genome. Zinc has a critical role in maintaining many biochemical and physiological processes at the molecular, cellular, and multiple organ and systemic levels. The alteration of zinc homeostasis causes dysfunction of many organs and systems. In the immune system, zinc regulates the differentiation, proliferation and function of inflammatory cells, including T cells, eosinophils, and B cells, by modifying several signaling pathways such as NFκB signaling pathways and TCR signals. An adequate zinc level is essential for proper immune responses and decreased zinc levels were reported in many allergic inflammatory diseases, including atopic dermatitis, bronchial asthma, and chronic rhinosinusitis. Decreased zinc levels often enhance inflammatory activation. On the other hand, the inflammatory conditions alter the intracellular homeostasis of zinc, often decreasing zinc levels. These findings implied that there could be a vicious cycle between zinc deficiency and inflammatory conditions. In this review, we present recent evidence on the involvement of zinc in atopic dermatitis, bronchial asthma, and chronic rhinosinusitis, with insights into the involvement of zinc in the underlying molecular and cellular mechanisms related to these allergic inflammatory diseases.

Management for a patient of moyamoya disease presenting with ischemic stroke in the first trimester of pregnancy.

We report an extremely rare case of a 27-year-old woman presenting with ischemic stroke as an initial manifestation of moyamoya disease in the first trimester of pregnancy. We conducted an artificial abortion when her neurological symptoms rapidly became refractory to optimal antithrombotic treatments. The progression of neurologic deficits stopped immediately after abortion, resulting in recovery to independence, with slight motor aphasia and right hemiparesis due to improved cerebral flow. We highlight rapid artificial abortion combined with antithrombotic treatment for patients of moyamoya disease with pregnancy-associated ischemic stroke as an appropriate treatment to correct hemodynamic instability and suppress the progression of neurological symptoms.

Preparation of Wrinkled Nanoribbons Consisting of Langmuir-Blodgett Films.

Langmuir-Blodgett films are normally deposited on hard substrates, such as mica plates. In this study, soft substrates consisting of a poly(dimethylsiloxane) elastomer were used for film deposition. A docosanoic acid monolayer was transferred onto the substrate by moving it in an upward direction while immersed in a subphase CoCl2 solution. Interestingly, the obtained deposited layer showed a ribbon-like morphology that oriented itself parallel to the water surface. Subsequently, by shrinking the substrate, wrinkled ribbons were obtained. This novel wrinkling method is unique to soft substrates because hard substrates cannot be shrunk. Such a wrinkling method can be useful to control the morphology of the LB films, and thus improve its properties as functional materials.

p53 represses the transcription of snRNA genes by preventing the formation of little elongation complex.

The regulation of transcription by RNA polymerase II (Pol II) is important for a variety of cellular functions. ELL/EAF-containing little elongation complex (LEC) was found to be required for transcription of Pol II-dependent small nuclear RNA (snRNA) genes. It was shown that the tumor suppressor p53 interacts with ELL and inhibits transcription elongation activity of ELL. Here, we show that p53 inhibits interaction between ELL/EAF and ICE1 in LEC and thereby p53 represses transcription of Pol II-dependent snRNA genes through inhibiting LEC function. Furthermore, induction of p53 expression by ultraviolet (UV) irradiation decreases the occupancy of ICE1 at Pol II-dependent snRNA genes. Consistent with the results, knockdown of p53 increased both the expression of snRNA genes and the occupancy of Pol II and components of LEC at snRNA genes. Our results indicate that p53 interferes with the interaction between ELL/EAF and ICE1 and represses transcription of snRNA genes by Pol II.

Sez6l2 regulates phosphorylation of ADD and neuritogenesis.

Increasing evidence shows that immune-mediated mechanisms may contribute to the pathogenesis of central nervous system disorders including cerebellar ataxias, as indicated by the aberrant production of neuronal surface antibodies. We previously reported a patient with cerebellar ataxia associated with production of a new anti-neuronal antibody, anti-seizure-related 6 homolog like 2 (Sez6l2). Sez6l2 is a type 1 membrane protein that is highly expressed in the hippocampus and cerebellar cortex and mice lacking Sez6l2 protein family members develop ataxia. Here we used a proteomics-based approach to show that serum derived from this patient recognizes the extracellular domain of Sez6l2 and that Sez6l2 protein binds to both adducin (ADD) and glutamate receptor 1 (GluR1). Our results indicate that Sez6l2 is one of the auxiliary subunits of the AMPA receptor and acts as a scaffolding protein to link GluR1 to ADD. Furthermore, Sez6l2 overexpression upregulates ADD phosphorylation, whereas siRNA-mediated downregulation of Sez612 prevents ADD phosphorylation, suggesting that Sez6l2 modulates AMPA-ADD signal transduction.

TRIM39 negatively regulates the NFκB-mediated signaling pathway through stabilization of Cactin.

NFκB is one of the central regulators of cell survival, immunity, inflammation, carcinogenesis and organogenesis. The activation of NFκB is strictly regulated by several posttranslational modifications including phosphorylation, neddylation and ubiquitination. Several types of ubiquitination play important roles in multi-step regulations of the NFκB pathway. Some of the tripartite motif-containing (TRIM) proteins functioning as E3 ubiquitin ligases are known to regulate various biological processes such as inflammatory signaling pathways. One of the TRIM family proteins, TRIM39, for which the gene has single nucleotide polymorphisms, has been identified as one of the genetic factors in Behcet's disease. However, the role of TRIM39 in inflammatory signaling had not been fully elucidated. In this study, to elucidate the function of TRIM39 in inflammatory signaling, we performed yeast two-hybrid screening using TRIM39 as a bait and identified Cactin, which has been reported to inhibit NFκB- and TLR-mediated transcriptions. We show that TRIM39 stabilizes Cactin protein and that Cactin is upregulated after TNFα stimulation. TRIM39 knockdown also causes activation of the NFκB signal. These findings suggest that TRIM39 negatively regulates the NFκB signal in collaboration with Cactin induced by inflammatory stimulants such as TNFα.

The novel heart-specific RING finger protein 207 is involved in energy metabolism in cardiomyocytes.

A failing heart shows severe energy insufficiency, and it is presumed that this energy shortage plays a critical role in the development of cardiac dysfunction. However, little is known about the mechanisms that cause energy metabolic alterations in the failing heart. Here, we show that the novel RING-finger protein 207 (RNF207), which is specifically expressed in the heart, plays a role in cardiac energy metabolism. Depletion of RNF207 in neonatal rat cardiomyocytes (NRCs) leads to a reduced cellular concentration of adenosine triphosphate (ATP) and mitochondrial dysfunction. Consistent with this result, we observed here that the expression of RNF207 was significantly reduced in mice with common cardiac diseases including heart failure. Intriguingly, proteomic approaches revealed that RNF207 interacts with the voltage-dependent anion channel (VDAC), which is considered to be a key regulator of mitochondria function, as an RNF207-interacting protein. Our findings indicate that RNF207 is involved in ATP production by cardiomyocytes, suggesting that RNF207 plays an important role in the development of heart failure.