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CTLA-4 is an important co-inhibitory receptor for T cells. Here, the authors show that CTLA-4 also has a function on B-1a cells, as conditional deletion results in activation of these cells and knockout mice develop an autoimmune profile.

Purpose. This study was aimed at exploring the effect of long noncoding RNA LINC00324 (LINC00324) on gastric cancer (GC) and the potential molecular mechanisms. Methods. The expression of LINC00324 and miR-3200-5p in GC tissues and cells was detected by qRT-PCR. LINC00324 was silenced in GC cells by transfection of si-LINC00324. Then, the proliferation, migration, and invasion of GC cells were analyzed by MTT, wound healing, and transwell assays, respectively. The interactions between LINC00324 and miR-3200-5p and between miR-3200-5p and BCAT1 were determined by a dual-luciferase reporter and/or RNA pull-down assay. Results. The expression of LINC00324 was upregulated in GC cells and tissues, but miR-3200-5p was downregulated. Silencing of LINC00324 inhibited the proliferation, migration, and invasion of GC cells. LINC00324 directly targeted miR-3200-5p, and miR-3200-5p directly targeted BCAT1. si-LINC00324 negatively regulated BCAT1 expression via binding to miR-3200-5p. Furthermore, silencing of LINC00324 reversed the promoting effects of BCAT1 on the proliferation, migration, and invasion of GC cells. Conclusion. Silencing of LINC00324 inhibited the proliferation, migration, and invasion of GC cells through regulating the miR-3200-5p/BCAT1 axis.

Numerical and experimental methods were applied to investigate fatigue crack propagation behavior of reinforced concrete (RC) beams strengthened with a new type carbon fiber reinforced polymer (CFRP) named as carbon fiber laminate (CFL) subjected to hot-wet environment. J-integral of a central crack in the strengthened beam under three-point bending load was calculated by ABAQUS. In finite element model, simulation of CFL-concrete interface was based on the bilinear cohesive zone model under hot-wet environment and indoor atmosphere. And, then, fatigue crack propagation tests were carried out under high temperature and high humidity (50°C, 95% R · H) environment pretreatment and indoor atmosphere (23°C, 78% R · H) to obtain a-N curves and crack propagation rate, da/dN, of the strengthened beams. Paris-Erdogan formula was developed based on the numerical analysis and environmental fatigue tests.

Processes that define immunoglobulin repertoires are commonly presumed to be the same for all murine B cells. However, studies here that couple high-dimensional FACS sorting with large-scale quantitative IgH deep-sequencing demonstrate that B-1a IgH repertoire differs dramatically from the follicular and marginal zone B cells repertoires and is defined by distinct mechanisms. We track B-1a cells from their early appearance in neonatal spleen to their long-term residence in adult peritoneum and spleen. We show that de novo B-1a IgH rearrangement mainly occurs during the first few weeks of life, after which their repertoire continues to evolve profoundly, including convergent selection of certain V(D)J rearrangements encoding specific CDR3 peptides in all adults and progressive introduction of hypermutation and class-switching as animals age. This V(D)J selection and AID-mediated diversification operate comparably in germ-free and conventional mice, indicating these unique B-1a repertoire-defining mechanisms are driven by antigens that are not derived from microbiota.

INTRODUCTION: Ca2+ spark constitutes the elementary units of cardiac excitation-contraction (E-C) coupling in mature cardiomyocytes. Human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes are known to have electrophysiological properties similar to mature adult cardiomyocytes. However, it is unclear if they share similar calcium handling property. We hypothesized that Ca2+ sparks in human induced pluripotent stem cell (hiPSCs)-derived cardiomyocytes (hiPSC-CMs) may display unique structural and functional properties than mature adult cardiomyocytes. METHODS AND RESULTS: Ca2+ sparks in hiPSC-CMs were recorded with Ca2+ imaging assay with confocal laser scanning microscopy. Those sparks were stochastic with a tendency of repetitive occurrence at the same site. Nevertheless, the spatial-temporal properties of Ca2+ spark were analogous to that of adult CMs. Inhibition of L-type Ca2+ channels by nifedipine caused a 61% reduction in calcium spark frequency without affecting amplitude of those sparks and magnitude of caffeine releasable sarcoplasmic reticulum (SR) Ca2+ content. In contrast, high extracellular Ca2+ and ryanodine increased the frequency, full width at half maximum (FWHM) and full duration at half maximum (FDHM) of spontaneous Ca2+ sparks. CONCLUSIONS: For the first time, spontaneous Ca2+ sparks were detected in hiPSC-CMs. The Ca2+ sparks are predominately triggered by L-type Ca2+ channels mediated Ca2+ influx, which is comparable to sparks detected in adult ventricular myocytes in which cardiac E-C coupling was governed by a Ca2+-induced Ca2+ release (CICR) mechanism. However, focal repetitive sparks originated from the same intracellular organelle could reflect an immature status of the hiPSC-CMs.

Ubiquitination is one of the most important post-translational modifications in eukaryotes. The ubiquitination cascade includes ubiquitin-activating enzymes (E1), ubiquitin-conjugating enzymes (E2), and ubiquitin ligases (E3). The E3 ligases, responsible for substrate recognition, are the most abundant and varied proteins in the cascade and the most studied. SKP1-CUL1-F-Box (SCF)-type E3 ubiquitin ligases are multi-subunit RING (Really Interesting New Gene) E3 ubiquitin ligases, composed of CUL1 (Cullin 1), RBX1 (RING BOX 1), SKP1 (S-phase Kinase-associated Protein 1), and F-box proteins. In vitro ubiquitination assays, used for studying the specific recognition of substrate proteins by E3 ubiquitin ligases, require the purification of all components involved in the cascade, and for assays with SCF-type E3 ligases, additional proteins (several SCF complex subunits). Here, the Duet expression system was used to co-express E1, E2, ubiquitin, ubiquitylation target (substrate), and the four subunits of a SCF-type E3 ligase in E. coli. When these proteins co-exist in bacterial cells, ubiquitination occurs and can be detected by Western Blot. The effectiveness of this bacterial system for detecting ubiquitination cascade activity was demonstrated by replicating both AtSCFTIR1-mediated and human SCFFBXO28-mediated ubiquitylation in bacteria. This system provides a basic but adaptable platform for the study of SCF-type E3 ubiquitin ligases. [ABSTRACT FROM AUTHOR]

Auxin regulates plant growth and development through downstream signaling pathways, including the best-known SCFTIR1/AFB-Aux/IAA-ARF pathway and several other less characterized "noncanonical" pathways. Recently, one SCFTIR1/AFB-independent noncanonical pathway, mediated by Transmembrane Kinase 1 (TMK1), was discovered through the analyses of its functions in Arabidopsis apical hook development. Asymmetric accumulation of auxin on the concave side of the apical hook triggers DAR1-catalyzed release of the C-terminal of TMK1, which migrates into the nucleus, where it phosphorylates and stabilizes IAA32/34 to inhibit cell elongation, which is essential for full apical hook formation. However, the molecular factors mediating IAA32/34 degradation have not been identified. Here, we show that proteins in the CYTOKININ INDUCED ROOT WAVING 1 (CKRW1)/WAVY GROWTH 3 (WAV3) subfamily act as E3 ubiquitin ligases to target IAA32/34 for ubiquitination and degradation, which is inhibited by TMK1c-mediated phosphorylation. This antagonistic interaction between TMK1c and CKRW1/WAV3 subfamily E3 ubiquitin ligases regulates IAA32/34 levels to control differential cell elongation along opposite sides of the apical hook. [ABSTRACT FROM AUTHOR]

The purpose of this experiment was to investigate the effects of garlic straw on the serum biochemistry, antioxidant capacity, growth performance, and nutrient apparent digestibility of Qingshan sheep. Forty 5-month-old male Qingshan sheep were randomly divided into four groups, with 10 replicates per group and one sheep per replicate. The control group was fed with barley straw, and Group 1, Group 2, and Group 3 were fed with garlic straw instead of 10%, 20%, and 30% barley straw, respectively. The pre-test period was seven days, and the formal test period was 30 days. The results showed that compared with the control group, the average daily gain of Group 1, Group 2 and Group 3 increased by 17.7%, 29.7%, and 16.5% (P<0.05), and the feed to gain ratio of Group 2 decreased by 13.1% (P<0.05). The apparent digestibility of dry matter (DM), crude protein (CP), neutral detergent fiber (NDF), and acid detergent fiber (ADF) in Group 1, Group 2, and Group 3 increased significantly (P<0.05). The total protein content in serum of Group 1, Group 2, and Group 3 increased by 3.7%, 4.1%, and 3.4% respectively (P<0.05), the albumin content increased by 6.5%, 5.2%, and 2.3% (P<0.05), the globulin content increased by 4.1%, 3.5%, and 3.0% (P<0.05), serum glucose content decreased by 7.6%, 10.2%, and 6.4% (P<0.05), and the content of triglyceride decreased by 14.3%, 23.8%, and 21.4%. The activities of superoxide dismutase (SOD), catalase (CAT), and total antioxidant capacity (T-AOC) in serum were significantly higher than those in control group (P<0.05), the activity of glutathione peroxidase (GSH-Px) in serum was significantly higher than those in control group and Group 1 (P<0.05), and the serum malondialdehyde (MDA) in Group 1, Group 2, and Group 3 was significantly lower than those in control group (P<0.05). The study indicates that garlic straw has a good effect on feeding Qingshan sheep instead of part of barley straw, and the best effect is to replace 20% barley straw. [ABSTRACT FROM AUTHOR]