Crosstalk among canonical Wnt and Hippo pathway members in skeletal muscle and at the neuromuscular junction

Skeletal muscles are essential for locomotion,posture,and metabolic regulation.To understand physiological processes,exercise adaptation,and muscle-related disorders,it is critical to understand the molecular pathways that underlie skeletal muscle function.The process of muscle contra ction,orchestrated by a complex interplay of molecular events,is at the core of skeletal muscle function.Muscle contraction is initiated by an action potential and neuromuscular transmission requiring a neuromuscular junction.Within muscle fibers,calcium ions play a critical role in mediating the interaction between actin and myosin filaments that generate force.Regulation of calcium release from the sarcoplasmic reticulum plays a key role in excitation-contraction coupling.The development and growth of skeletal muscle are regulated by a network of molecular pathways collectively known as myogenesis.Myogenic regulators coordinate the diffe rentiation of myoblasts into mature muscle fibers.Signaling pathways regulate muscle protein synthesis and hypertrophy in response to mechanical stimuli and nutrient availability.Seve ral muscle-related diseases,including congenital myasthenic disorders,sarcopenia,muscular dystrophies,and metabolic myopathies,are underpinned by dys regulated molecular pathways in skeletal muscle.Therapeutic interventions aimed at preserving muscle mass and function,enhancing regeneration,and improving metabolic health hold promise by targeting specific molecular pathways.Other molecular signaling pathways in skeletal muscle include the canonical Wnt signaling pathway,a critical regulator of myogenesis,muscle regeneration,and metabolic function,and the Hippo signaling pathway.In recent years,more details have been uncovered about the role of these two pathways during myogenesis and in developing and adult skeletal muscle fibers,and at the neuromuscular junction.In fact,research in the last few years now suggests that these two signaling pathways are interconnected and that they jointly control physiological and pathophysiological processes in muscle fibers.In this review,we will summarize and discuss the data on these two pathways,focusing on their concerted action next to their contribution to skeletal muscle biology.However,an in-depth discussion of the noncanonical Wnt pathway,the fibro/a dipogenic precursors,or the mechanosensory aspects of these pathways is not the focus of this review.

Expression of hepatocyte nuclear factor 4 alpha,wingless-related integration site,andβ-catenin in clinical gastric cancer

BACKGROUND Gastric cancer(GC)is the second most common cause of cancer-related deaths worldwide.Hepatocyte nuclear factor 4 alpha(HNF4α)that belongs to the nuclear hormone receptor superfamily,is overexpressed in GC tissues,and might be involved in the development of GC by regulating its downstream winglessrelated integration site(WNT)/β-catenin signaling.AIM To clarify the expression of HNF4α/WNT5a/β-catenin signaling proteins in clinical GC tissues.METHODS We immunohistochemically stained pathological blocks of GC and matched paracancerous tissues.The intensity of HNF4α,WNT5a andβ-catenin staining in the tumor cells was determined according to cell rates and staining intensity.The correlations between GC and HNF4α,WNT5a,andβ-catenin expression using chisquare and paired chi-square tests.Relationships between double-positive HNF4αand WNT5a expression and types of gastric tumor tissues were assessed using regression analysis.Correlations between HNF4αand WNT5a expression at the RNA level in GC tissues found in the TCGA database were analyzed using Pearson correlation coefficients.RESULTS We found more abundant HNF4αand WNT5a proteins in GC,especially in mucinous adenocarcinoma and mixed GC than in adjacent tissues(P<0.001).Low and high levels of cytoplasmicβ-catenin respectively expressed in GC and adjacent tissues(P<0.001)were not significantly associated with pathological parameters.CONCLUSION The expressions of HNF4αand WNT5a could serve as early diagnostic biomarkers for GC.

Expression and function of WNT4 involved in larvae development and limb regeneration in Portunus trituberculatus

The wingless-related integration site(WNT)proteins are a family of secreted glycoproteins that are evolutionarily conserved and are believed to be involved in evolution in vertebrates and invertebrates.WNT signaling pathways may be associated with limb regeneration and development in crustaceans.However,the detail mechanisms remain unclear.Therefore,the distribution of WNT4 in the hepatopancreas,muscle,hemocyte,ganglion,heart,eyestalk,gill tissue,and diff erent larvae development stages of the swimming crab(Portunus trituberculatus)were characterized using immunofl uorescence,real-time PCR,and Western blotting.Signifi cant PtWNT4 expression was detected in heart and eyestalk.In addition,PtWNT4 was expressed in all larval stages of P.trituberculatus with a dynamic expression pattern,especially in the eyestalk and other organs in the carapace area.The injection of WNT4 dsRNA into regenerative limbs signifi cantly decreased PtWNT4 mRNA levels in the eyestalk,heart,and muscle,resulting in 1.9-fold,2.2-fold,and 2.7-fold decreases compared with those detected in the group injected with crab saline(P<0.05),respectively,indicating successful gene silencing.Overall,expression analysis on the WNT4 using RNAi provides an insight to its functional mechanism during limb regeneration in P.trituberculatus.The results not only demonstrated the requirement for WNT4 in limb regeneration of Crustaceans,but also suggested its ability to promote larval development at specifi c stages.

VIS Atlas:A Database of Virus Integration Sites in Human Genome from NGS Data to Explore Integration Patterns

Integration of oncogenic DNA viruses into the human genome is a key step in most virusinduced carcinogenesis.Here,we constructed a virus integration site(VIS)Atlas database,an extensive collection of integration breakpoints for three most prevalent oncoviruses,human papillomavirus,hepatitis B virus,and Epstein-Barr virus based on the next-generation sequencing(NGS)data,literature,and experimental data.There are 63,179 breakpoints and 47,411 junctional sequences with full annotations deposited in the VIS Atlas database,comprising 47 virus genotypes and 17 disease types.The VIS Atlas database provides(1)a genome browser for NGS breakpoint quality check,visualization of VISs,and the local genomic context;(2)a novel platform to discover integration patterns;and(3)a statistics interface for a comprehensive investigation of genotypespecific integration features.Data collected in the VIS Atlas aid to provide insights into virus pathogenic mechanisms and the development of novel antitumor drugs.

Identification of neutral genome integration sites with high expression and high integration efficiency in Fusarium venenatum TB01

CRISPR/Cas9-mediated homology-directed recombination is an efficient method to express target genes.Based on the above method,providing ideal neutral integration sites can ensure the reliable,stable,and high expression of target genes.In this study,we obtained a fluorescent transformant with neutral integration and high expression of the GFP expression cassette from the constructed GFP expression library and named strain FS.The integration site mapped at 4886 bp upstream of the gene FVRRES_00686 was identified in strain FS based on a Y-shaped adaptor-dependent extension,and the sequence containing 600 bp upstream and downstream of this site was selected as the candidate region for designing sgRNAs(Sites)for CRISPR/Cas9-mediated homology-directed recombination.PCR analysis showed that the integration efficiency of CRISPR/Cas9-mediated integration of target genes in designed sites reached 100%.Further expression stability and applicability analysis revealed that the integration of the target gene into the above designed sites can be stably inherited and expressed and has no negative effect on the growth of F.venenatum TB01.These results indicate the above designed neutral sites have the potential to accelerate the development of F.venenatum TB01 through overexpression of target genes in metabolic engineering.

Determination of the chromosomal position effects for plug-and-play application in the Myxococcus xanthus chassis cells

The chromosomal position effect can significantly affect the transgene expression,which may provide an efficient strategy for the inauguration of alien genes in new hosts,but has been less explored rationally.The bacterium Myxococcus xanthus harbors a large circular high-GC genome,and the position effect in this chassis may result in a thousand-fold expression variation of alien natural products.In this study,we conducted transposon insertion at TA sites on the M.xanthus genome,and used enrichment and dilution indexes to respectively appraise high and low expression potentials of alien genes at insertion sites.The enrichment sites are characteristically distributed along the genome,and the dilution sites are overlapped well with the horizontal transfer genes.We experimentally demonstrated the enrichment sites as high expression integration sites(HEISs),and the dilution sites unsuitable for gene integration expression.This work highlights that HEISs are the plug-and-play sites for efficient expression of integrated genes.

The integration characteristics of the exogenous growth hormone gene in a transgenic common carp(Cyprinus carpio L.) with fast-growth performance

The genetic stability and expression efficiency of exogenous genes in transgenic animals are closely related to integration site and copy number. In our laboratory, by transgenic manipulation and subsequent test crosses, we established an ‘‘all-fish'' growth hormone(GH)transgenic common carp family that exhibits fast growth.In this present study, genome walking, real-time quantitative polymerase chain reaction, and fluorescence in situ hybridization techniques were applied to identify the integration characteristics of the exogenous grass carp GH gene in the transgenic common carp. The exogenous GH genes, in the form of two complete and one incomplete tandem repeats, were found to have integrated into an ATrich region near the end of a chromosome pair. We hypothesize that the high efficiency of exogenous GH gene expression might be due to the low copy number in the genome and the AT-rich integration site.

Heterogeneity of HIV-1 latent reservoirs

Antiretroviral therapy(ART)can effectively inhibit human immunodeficiency virus-1(HIV-1)replication,but is not curative due to the existence of a stable viral latent reservoir harboring replication-competent proviruses.In order to reduce or eliminate the HIV-1 latent reservoir,characteristics of the latently infected cells need to be intensively studied,and a comprehensive understanding of the heterogenous nature of the latent reservoir will be critical to develop novel therapeutic strategies.Here,we discuss the different cell types and mechanisms contributing to the complexity and heterogeneity of HIV-1 latent reservoirs,and summarize the key challenges to the development of cure strategies for acquired immunodeficiency syndrome(AIDS).