Connexin 36 (Cx36) is a channel-forming protein found in the membranes of apposed cells, forming the hexameric hemichannels of intercellular gap junction channels. It localizes to certain neurons in various regions of the brain including the retina. We characterized the expression pattern of neuronal Cx36 in the guinea pig retina by immunocytochemistry using specific antisera against Cx36 and green/red cone opsin or recoverin. Strong Cx36 immunoreactivity was visible in the ON sublamina of the inner plexiform layer and in the outer plexiform layer, as punctate labelling patterns. Double-labelling experiments with antibody directed against Cx36 and green/red cone opsin or recoverin showed that strong clustered Cx36 immunoreactivity localized to the axon terminals of cone or close to rod photoreceptors. By electron microscopy, Cx36 immunoreactivity was visible in the gap junctions as well as in the cytoplasmic matrices of both sides of cone photoreceptors. In the gap junctions between cone and rod photoreceptors, Cx36 immunoreactivity was only visible in the cytoplasmic matrices of cone photoreceptors. These results clearly indicate that Cx36 forms homologous gap junctions between neighbouring cone photoreceptors, and forms heterologous gap junctions between cone and rod photoreceptors in guinea pig retina. This focal location of Cx36 at the terminals of the photoreceptor suggests that rod photoreceptors can transmit rod signals to the pedicle of a neighbouring cone photoreceptor via Cx36, and that the cone in turn signals to corresponding ganglion cells via ON and OFF cone bipolar cells.

Phage display technology provides a powerful tool to screen a library for a binding molecule via an enrichment process. It has been adopted as a critical technology in the development of therapeutic antibodies. However, a major drawback of phage display technology is that because the degree of the enrichment cannot be controlled during the bio-panning process, it frequently results in a limited number of clones. In this study, we applied next-generation sequencing (NGS) to screen clones from a library and determine whether a greater number of clones can be identified using NGS than using conventional methods. Three chicken immune single-chain variable fragment (scFv) libraries were subjected to bio-panning on prostate-specific antigen (PSA). Phagemid DNA prepared from the original libraries as well as from the Escherichia coli pool after each round of bio-panning was analyzed using NGS, and the heavy chain complementarity-determining region 3 (HCDR3) sequences of the scFv clones were determined. Subsequently, through two-step linker PCR and cloning, the entire scFv gene was retrieved and analyzed for its reactivity to PSA in a phage enzyme immunoassay. After four rounds of bio-panning, the conventional colony screening method was performed for comparison. The scFv clones retrieved from NGS analysis included all clones identified by the conventional colony screening method as well as many additional clones. The enrichment of the HCDR3 sequence throughout the bio-panning process was a positive predictive factor for the selection of PSA-reactive scFv clones.

Purpose To investigate the diagnostic accuracy of radiographic signs for complete discoid lateral meniscus and whether a predictive model combining the radiographic signs can improve its diagnostic accuracy in adults. Methods A total of adult 119 knees with complete discoid lateral meniscus conirmed by arthroscopy and 119 age-and sex-matched knees with normal meniscus were included. The radiographic signs of lateral joint space, ibular head height, lateral tibial spine height, lateral tibial plateau obliquity, lateral femoral condyle squaring, lateral tibial plateau cupping, lateral femoral condyle notching, and the condylar cut-of sign were evaluated. The receiver-operating characteristic (ROC) curves and area under the curve (AUC) were evaluated for best accuracy. A prediction model was developed by multivariable regression with generalized estimating models, and was validated using data from 111 knees of children with complete discoid lateral meniscus and 111 normal controls. ResultsThe ibular head height, lateral joint space, lateral tibial plateau obliquity, and the condylar cut-of sign were signiicantly diferent between the complete discoid lateral meniscus and the normal groups (p < 0.05). Among the four radiographic signs, the ibular head height showed the highest accuracy with 78.9% sensitivity and 57.3% speciicity. The prediction models developed by logistic regression showed signiicantly improved accuracy for complete discoid lateral meniscus compared to the ibular head height (sensitivity: 69.8%, speciicity: 82.9%, p = 0.001). For validation, the AUC of children seemed to be larger than that of adults, which indicated that the prediction models could be applied for children to detect complete discoid lateral meniscus. Conclusion Among several radiographic signs, the ibular head height can be used as a screening tool for complete discoid lateral meniscus. The prediction models combined with lateral joint space, ibular head height, lateral tibial plateau obliquity, and/or the condylar cut-of sign yielded a much higher diagnostic value than each radiographic sign. Therefore, ibular head height and prediction models combined with radiographic signs can provide improved diagnostic value for complete discoid lateral meniscus. Level of evidence III.

Post-translational modifications of core histones affect various cellular processes, primarily through transcription. However, their relationship with the termination of transcription has remained largely unknown. In this study, we show that DNA damage-activated AKT phosphorylates threonine 45 of core histone H3 (H3-T45). By genome-wide chromatin immunoprecipitation sequencing (ChIP-seq) analysis, H3-T45 phosphorylation was distributed throughout DNA damage-responsive gene loci, particularly immediately after the transcription termination site. H3-T45 phosphorylation pattern showed close-resemblance to that of RNA polymerase II C-terminal domain (CTD) serine 2 phosphorylation, which establishes the transcription termination signal. AKT1 was more effective than AKT2 in phosphorylating H3-T45. Blocking H3-T45 phosphorylation by inhibiting AKT or through amino acid substitution limited RNA decay downstream of mRNA cleavage sites and decreased RNA polymerase II release from chromatin. Our findings suggest that AKT-mediated phosphorylation of H3-T45 regulates the processing of the 3′ end of DNA damage-activated genes to facilitate transcriptional termination.