The long noncoding RNA lncMPD2 inhibits myogenesis by targeting the miR-34a-5p/THBS1 axis.

Long noncoding RNAs (lncRNAs) participate in regulating skeletal muscle development. However, little is known about their role in regulating chicken myogenesis. In this study, we identified a novel lncRNA, lncMPD2, through transcriptome sequencing of chicken myoblasts at different developmental stages. Functionally, gain- and loss-of-function experiments showed that lncMPD2 inhibited myoblast proliferation and differentiation. Mechanistically, lncMPD2 directly bound to miR-34a-5p, and miR-34a-5p promoted myoblasts proliferation and differentiation and inhibited the mRNA and protein expression of its target gene THBS1. THBS1 inhibited myoblast proliferation and differentiation in vitro and delayed muscle regeneration in vivo. Furthermore, rescue experiments showed that lncMPD2 counteracted the inhibitory effects of miR-34a-5p on THBS1 and myogenesis-related gene mRNA and protein expression. In conclusion, lncMPD2 regulates the miR-34a-5p/THBS1 axis to inhibit the proliferation and differentiation of myoblasts and skeletal muscle regeneration. This study provides more insight into the molecular regulatory network of skeletal muscle development, identifying novel potential biomarkers for improving chicken quality and increasing chicken yield. In addition, this study provides a potential goal for breeding strategies that minimize muscle damage in chickens.

An immobilized antibody-based affinity grid strategy for on-grid purification of target proteins enables high-resolution cryo-EM.

In cryo-electron microscopy (cryo-EM), sample preparation poses a critical bottleneck, particularly for rare or fragile macromolecular assemblies and those suffering from denaturation and particle orientation distribution issues related to air-water interface. In this study, we develop and characterize an immobilized antibody-based affinity grid (IAAG) strategy based on the high-affinity PA tag/NZ-1 antibody epitope tag system. We employ Pyr-NHS as a linker to immobilize NZ-1 Fab on the graphene oxide or carbon-covered grid surface. Our results demonstrate that the IAAG grid effectively enriches PA-tagged target proteins and overcomes preferred orientation issues. Furthermore, we demonstrate the utility of our IAAG strategy for on-grid purification of low-abundance target complexes from cell lysates, enabling atomic resolution cryo-EM. This approach greatly streamlines the purification process, reduces the need for large quantities of biological samples, and addresses common challenges encountered in cryo-EM sample preparation. Collectively, our IAAG strategy provides an efficient and robust means for combined sample purification and vitrification, feasible for high-resolution cryo-EM. This approach holds potential for broader applicability in both cryo-EM and cryo-electron tomography (cryo-ET).

Divergent Regulatory Roles of Transcriptional Variants of the Chicken LDB3 Gene in Muscle Shaping.

LIM domain binding 3 (LDB3) serves as a striated muscle-specific Z-band alternatively spliced protein that plays an important role in mammalian skeletal muscle development, but its regulatory role and molecular mechanism in avian muscle development are still unclear. In this study, we reanalyzed RNA sequencing data sets of 1415 samples from 21 chicken tissues published in the NCBI GEO database. First, three variants (LDB3-X, LDB3-XN1, and LDB3-XN2) generated by alternative splicing of the LDB3 gene were identified in chicken skeletal muscle, among which LDB3-XN1 and LDB3-XN2 are novel variants. LDB3-X and LDB3-XN1 are derived from exon skipping in chicken skeletal muscle at the E18-D7 stage and share three LIM domains, but LDB3-XN2 lacks a LIM domain. Our results preliminarily suggest that the formation of three variants of LDB3 is regulated by RBM20. The three splice isomers have divergent functions in skeletal muscle according to in vitro and in vivo assays. Finally, we identified the mechanism by which different variants play different roles through interactions with IGF2BP1 and MYHC, which promote the proliferation and differentiation of chicken myoblasts, in turn regulating chicken myogenesis. In conclusion, this study revealed the divergent roles of three LDB3 variants in chicken myogenesis and muscle remodeling and demonstrated their regulatory mechanism through protein-protein interactions.

Whole transcriptome profiling reveals a lncMDP1 that regulates myogenesis by adsorbing miR-301a-5p targeting CHAC1.

Myoblast proliferation and differentiation are essential for skeletal muscle development. In this study, we generated the expression profiles of mRNAs, long noncoding RNAs (lncRNAs), and microRNAs (miRNAs) in different developmental stages of chicken primary myoblasts (CPMs) using RNA sequencing (RNA-seq) technology. The dual luciferase reporter system was performed using chicken embryonic fibroblast cells (DF-1), and functional studies quantitative real-time polymerase chain reaction (qPCR), cell counting kit-8 (CCK-8), 5-Ethynyl-2'-deoxyuridine (EdU), flow cytometry cycle, RNA fluorescence in situ hybridization (RNA-FISH), immunofluorescence, and western blotting assay. Our research demonstrated that miR-301a-5p had a targeted binding ability to lncMDP1 and ChaC glutathione-specific gamma-glutamylcyclotransferase 1 (CHAC1). The results revealed that lncMDP1 regulated the proliferation and differentiation of myoblasts via regulating the miR-301a-5p/CHAC1 axis, and CHAC1 promotes muscle regeneration. This study fulfilled the molecular regulatory network of skeletal muscle development and providing an important theoretical reference for the future improvement of chicken meat performance and meat quality.

Incorporating Neural Networks into the AMOEBA Polarizable Force Field.

Neural network potentials (NNPs) offer significant promise to bridge the gap between the accuracy of quantum mechanics and the efficiency of molecular mechanics in molecular simulation. Most NNPs rely on the locality assumption that ensures the model's transferability and scalability and thus lack the treatment of long-range interactions, which are essential for molecular systems in the condensed phase. Here we present an integrated hybrid model, AMOEBA+NN, which combines the AMOEBA potential for the short- and long-range noncovalent atomic interactions and an NNP to capture the remaining local covalent contributions. The AMOEBA+NN model was trained on the conformational energy of the ANI-1x data set and tested on several external data sets ranging from small molecules to tetrapeptides. The hybrid model demonstrated substantial improvements over the baseline models in term of accuracy as the molecule size increased, suggesting its potential as a next-generation approach for chemically accurate molecular simulations.

RRM2 promotes the proliferation of chicken myoblasts, inhibits their differentiation and muscle regeneration.

During myogenesis and regeneration, the proliferation and differentiation of myoblasts play key regulatory roles and may be regulated by many genes. In this study, we analyzed the transcriptomic data of chicken primary myoblasts at different periods of proliferation and differentiation with protein‒protein interaction network, and the results indicated that there was an interaction between cyclin-dependent kinase 1 (CDK1) and ribonucleotide reductase regulatory subunit M2 (RRM2). Previous studies in mammals have a role for RRM2 in skeletal muscle development as well as cell growth, but the role of RRM2 in chicken is unclear. In this study, we investigated the effects of RRM2 on skeletal muscle development and regeneration in chickens in vitro and in vivo. The interaction between RRM2 and CDK1 was initially identified by co-immunoprecipitation and mass spectrometry. Through a dual luciferase reporter assay and quantitative real-time PCR, we identified the core promoter region of RRM2, which is regulated by the SP1 transcription factor. In this study, through cell counting kit-8 assays, 5-ethynyl-2'-deoxyuridine incorporation assays, flow cytometry, immunofluorescence staining, and Western blot analysis, we demonstrated that RRM2 promoted the proliferation and inhibited the differentiation of myoblasts. In vivo studies showed that RRM2 reduced the diameter of muscle fibers and slowed skeletal muscle regeneration. In conclusion, these data provide preliminary insights into the biological functions of RRM2 in chicken muscle development and skeletal muscle regeneration.

A mechanoluminescent material, ZnS:Mn,Li, with enhanced brightness for visualizing dental occlusion.

Mechanoluminescent materials are characterized by high luminescence intensity, high repeatability, no external voltage activation, and a good linear relationship between stress and mechanoluminescence intensity within a certain range. Therefore, mechanoluminescent materials have attracted increasing attention from researchers in the fields of stress sensing, encryption and anti-counterfeiting, structural health monitoring, energy-saving lighting, intelligent wearable devices, and other fields. In this study, ZnS:Mn powders with different Mn2+ ratios and different ion doping were synthesized by a high-temperature solid-phase reaction, and the synthesis of various materials was characterized. Then, the optimal mechanoluminescence effect of the ZnS:1%Mn,1%Li material was obtained. The photoluminescence intensity of ZnS:1%Mn,1%Li was 16.7 times higher than that of the sample without doping with Li+, and the mechanoluminescence intensity was 1.64 times higher. Finally, polyethylene terephthalate (PET) film was combined with ZnS:Mn,Li mechanoluminescent powders to prepare flexible three-layer composite film. Based on this, a feasible strategy for the detection of temporomandibular disorders was proposed. The composite film is easy to use, economical, and safe, and has good mechanoluminescent performance, which has potential application value in the field of occlusal force detection and visualization.

Effects of SLC45A2 and GPNMB on Melanin Deposition Based on Transcriptome Sequencing in Chicken Feather Follicles.

As an essential genetic and economic trait, chicken feather color has long been an important research topic. To further understand the mechanism of melanin deposition associated with coloration in chicken feathers, we selected feather follicle tissues from the neck and wings of chickens with differently colored feathers (yellow, sub-Columbian, and silver) for transcriptome analysis. We focused on genes that were expressed in both the wings and neck and were expressed with the same trends in breeds with two different plumage colors, specifically, SLC45A2, GPNMB, MLPH, TYR, KIT, WNT11, and FZD1. GO and KEGG enrichment analyses showed the DEGs were enriched in melanin-related pathways, such as tyrosine metabolic pathway and melanogenesis, and PPI analysis highlighted the genes SLC45A2 and GPNMB as associated with melanin deposition. Verification experiments in chicken melanocytes demonstrated that these two genes promote melanocyte melanin deposition. These data enrich our knowledge of the mechanisms that regulate chicken feather color.

Transcriptome-Based Identification of the Muscle Tissue-Specific Expression Gene CKM and Its Regulation of Proliferation, Apoptosis and Differentiation in Chicken Primary Myoblasts.

Skeletal muscle is an essential tissue in meat-producing animals, and meat-producing traits have been a hot topic in chicken genetic breeding research. Current research shows that creatine kinase M-type-like (CKM) is one of the most abundant proteins in skeletal muscle and plays an important role in the growth and development of skeletal muscle, but its role in the development of chicken skeletal muscle is still unclear. Via RNA sequencing (RNA-seq), we found that CKM was highly expressed in chicken breast muscle tissue. In this study, the expression profile of CKM was examined by quantitative real-time PCR (qPCR), and overexpression and RNA interference techniques were used to explore the functions of CKM in the proliferation, apoptosis and differentiation of chicken primary myoblasts (CPMs). It was shown that CKM was specifically highly expressed in breast muscle and leg muscle and was highly expressed in stage 16 embryonic muscle, while CKM inhibited proliferation, promoted the apoptosis and differentiation of CPMs and was involved in regulating chicken myogenesis. Transcriptome sequencing was used to identify genes that were differentially expressed in CPMs after CKM disruption, and bioinformatics analysis showed that CKM was involved in regulating chicken myogenesis. In summary, CKM plays an important role in skeletal muscle development during chicken growth and development.

Nd-Mn Molecular Cluster with Searched Targets for Oral Cancer Imaging.

In this research, we designed a novel NIR II luminescence imaging probe with targeting effect to accurately track oral squamous cell carcinoma (OSCC) cells. Massive gene expression data were processed by weighted gene co-expression network analysis to establish a network of relationships between genes. After clustering, correlation of clinical information, and gene functional enrichment analysis, MMP1 was predicted to be a biomarker/therapeutic target for OSCC cells. To obtain rare-earth probes with better luminescence in the NIR II region, we adjusted the doping ratio of the rare-earth element (Nd, Gd, Er, and Yb) fraction of the Nd-Mn molecular cluster to optimize its luminescence properties. The results of in vitro targeting experiments showed that Nd-Mn-MMP1Ab can target Cal-27 cells, demonstrating at the cellular level that the MMP1 gene is a biomarker for oral cancer, which also proves that the cancer targets predicted by the bioinformatics approach are correct.

Pathway and mechanism of tubulin folding mediated by TRiC/CCT along its ATPase cycle revealed using cryo-EM.

The eukaryotic chaperonin TRiC/CCT assists the folding of about 10% of cytosolic proteins through an ATP-driven conformational cycle, and the essential cytoskeleton protein tubulin is the obligate substrate of TRiC. Here, we present an ensemble of cryo-EM structures of endogenous human TRiC throughout its ATPase cycle, with three of them revealing endogenously engaged tubulin in different folding stages. The open-state TRiC-tubulin-S1 and -S2 maps show extra density corresponding to tubulin in the cis-ring chamber of TRiC. Our structural and XL-MS analyses suggest a gradual upward translocation and stabilization of tubulin within the TRiC chamber accompanying TRiC ring closure. In the closed TRiC-tubulin-S3 map, we capture a near-natively folded tubulin-with the tubulin engaging through its N and C domains mainly with the A and I domains of the CCT3/6/8 subunits through electrostatic and hydrophilic interactions. Moreover, we also show the potential role of TRiC C-terminal tails in substrate stabilization and folding. Our study delineates the pathway and molecular mechanism of TRiC-mediated folding of tubulin along the ATPase cycle of TRiC, and may also inform the design of therapeutic agents targeting TRiC-tubulin interactions.

Research Note: Combined analysis of BSA-seq based mapping and RNA-seq reveals candidate genes associated with sub-Columbian plumage in H line chickens.

Columbian coloration patterns in plumage are widespread phenomena in several standard breeds of poultry, such as the Columbian Plymouth Rock chicken. H line chicken plumage is generally a pure white except in the hackle, wing, and tail plumage, which coloration is very similar to the Columbian plumage pattern, but with the barring substituting for the black vertical striping. Thus, we refer to this plumage coloration as "sub-Columbian" pattern. However, the genetic basis of this phenotype remains unknown. Here, a F3 cross population between yellow plumage roosters and sub-Columbian plumage hens was constructed, for verifying sub-Columbian plumage was sex-linked dominant inheritance. To identify the candidate regions, F2 generation sub-Columbian plumage hens and yellow plumage hens with their parental lines were used for BSA-seq, and sub-Columbian plumage genes were mapped to a 10.46 Mb interval on chromosome Z. Remarkably, by transcriptome analysis of the neck and wing tip follicle tissues of the 2 plumage colors, we demonstrated that within the interval, only 1 gene, SLC45A2 expressed significant differently (P < 0.05). Through KASP, we identified L347M and A10331272T in solute carrier family 45 member 2 (SLC45A2), and B2 haplotype of cyclin-dependent kinase inhibitor 2A (CDKN2A), showed near complete association with the phenotype. Eventually, we designed a hybridization experiment for verifying the locus of sub-Columbian plumage, which is inherited through Z-linked dominant inheritance and is controlled by SLC45A2 and CDKN2A.

Structural basis of plp2-mediated cytoskeletal protein folding by TRiC/CCT.

The cytoskeletal proteins tubulin and actin are the obligate substrates of TCP-1 ring complex/Chaperonin containing TCP-1 (TRiC/CCT), and their folding involves co-chaperone. Through cryo-electron microscopy analysis, we present a more complete picture of TRiC-assisted tubulin/actin folding along TRiC adenosine triphosphatase cycle, under the coordination of co-chaperone plp2. In the open S1/S2 states, plp2 and tubulin/actin engaged within opposite TRiC chambers. Notably, we captured an unprecedented TRiC-plp2-tubulin complex in the closed S3 state, engaged with a folded full-length ß-tubulin and loaded with a guanosine triphosphate, and a plp2 occupying opposite rings. Another closed S4 state revealed an actin in the intermediate folding state and a plp2. Accompanying TRiC ring closure, plp2 translocation could coordinate substrate translocation on the CCT6 hemisphere, facilitating substrate stabilization and folding. Our findings reveal the folding mechanism of the major cytoskeletal proteins tubulin/actin under the coordination of the biogenesis machinery TRiC and plp2 and extend our understanding of the links between cytoskeletal proteostasis and related human diseases.

Thermally activated upconversion luminescence and ratiometric temperature sensing under 1064 nm/808 nm excitation.

In this research, a thermally activated upconversion luminescence (UCL) probe with ratiometric temperature sensing under 1064 nm and 808 nm excitation was designed. Especially, Nd3+, Tm3+and Ce3+were doped in rare earth nanoparticles (RENPs) as UCL modulators. By optimizing the elements and ratios, the excitation wavelength is successfully modulated to 1064 nm excitation with UCL intensity enhanced. Additionally, the prepared RENPs have a significant temperature response at 1064 nm excitation and can be used for thermochromic coatings. The intensity ratio of three-photon UCL (1064 nm excitation) to two-photon UCL (808 nm excitation) as an exponential function of temperature can be used as a ratiometric temperature detector. Therefore, this designed thermochromic coatings may enable new applications in optoelectronic device and industrial sensor.

Molecular Characterization, Expression Profile, and A 21-bp Indel within the ASB9 Gene and Its Associations with Chicken Production Traits.

A growing number of studies have shown that members of the ankyrin repeat and suppressors of cytokine signaling (SOCS) box-containing protein (ASB) family are extensively involved in biological processes such as cell growth, tissue development, insulin signaling, ubiquitination, protein degradation, and skeletal muscle membrane protein formation, while the specific biological role of ankyrin-repeat and SOCS box protein 9 (ASB9) remains unclear. In this study, a 21 bp indel in the intron of ASB9 was identified for the first time in 2641 individuals from 11 different breeds and an F2 resource population, and differences were observed among individuals with different genotypes (II, ID, and DD). An association study of a cross-designed F2 resource population revealed that the 21-bp indel was significantly related to growth and carcass traits. The significantly associated growth traits were body weight (BW) at 4, 6, 8, 10, and 12 weeks of age; sternal length (SL) at 4, 8, and 12 weeks of age; body slope length (BSL) at 4, 8, and 12 weeks of age; shank girth (SG) at 4 and 12 weeks of age; tibia length (TL) at 12 weeks of age; and pelvic width (PW) at 4 weeks of age (p < 0.05). This indel was also significantly correlated with carcass traits including semievisceration weight (SEW), evisceration weight (EW), claw weight (CLW), breast muscle weight (BMW), leg weight (LeW), leg muscle weight (LMW), claw rate (CLR), and shedding weight (ShW) (p < 0.05). In commercial broilers, the II genotype was the dominant genotype and underwent extensive selection. Interestingly, the ASB9 gene was expressed at significantly higher levels in the leg muscles of Arbor Acres broilers than those of Lushi chickens, while the opposite was true for the breast muscles. In summary, the 21-bp indel in the ASB9 gene significantly influenced the expression of the ASB9 gene in muscle tissue and was associated with multiple growth and carcass traits in the F2 resource population. These findings suggested that the 21-bp indel within the ASB9 gene could be used in marker-assisted selection breeding for traits related to chicken growth.

Comment on the Marshall-Hoare-Henssge model for estimating the time since death.

Timewise temperature variations in objects that are undergoing unsteady heating or cooling is a commonly encountered problem in the thermal sciences. One particular area of application is the cooling of a body post-death and the use of body temperatures to estimate the time of death. Here, a new approach based on the theory of transient heat transfer is formulated to allow efficient calculation of unsteady conduction problems. The theoretically derived unsteady temperature models are compared with experimentally based correlations (the Marshall-Hoare-Henssge model). The two approaches are found to agree very well. With this new theoretically based approach, timewise temperature variation can be calculated for both large and small Biot number transient problems.

Current Trends and Challenges in Drug-Likeness Prediction: Are They Generalizable and Interpretable?

Importance: Drug-likeness of a compound is an overall assessment of its potential to succeed in clinical trials, and is essential for economizing research expenditures by filtering compounds with unfavorable properties and poor development potential. To this end, a robust drug-likeness prediction method is indispensable. Various approaches, including discriminative rules, statistical models, and machine learning models, have been developed to predict drug-likeness based on physiochemical properties and structural features. Notably, recent advancements in novel deep learning techniques have significantly advanced drug-likeness prediction, especially in classification performance. Highlights: In this review, we addressed the evolving landscape of drug-likeness prediction, with emphasis on methods employing novel deep learning techniques, and highlighted the current challenges in drug-likeness prediction, specifically regarding the aspects of generalization and interpretability. Moreover, we explored potential remedies and outlined promising avenues for future research. Conclusion: Despite the hurdles of generalization and interpretability, novel deep learning techniques have great potential in drug-likeness prediction and are worthy of further research efforts.

An Efficient Approach to Large-Scale Ab Initio Conformational Energy Profiles of Small Molecules.

Accurate conformational energetics of molecules are of great significance to understand maby chemical properties. They are also fundamental for high-quality parameterization of force fields. Traditionally, accurate conformational profiles are obtained with density functional theory (DFT) methods. However, obtaining a reliable energy profile can be time-consuming when the molecular sizes are relatively large or when there are many molecules of interest. Furthermore, incorporation of data-driven deep learning methods into force field development has great requirements for high-quality geometry and energy data. To this end, we compared several possible alternatives to the traditional DFT methods for conformational scans, including the semi-empirical method GFN2-xTB and the neural network potential ANI-2x. It was found that a sequential protocol of geometry optimization with the semi-empirical method and single-point energy calculation with high-level DFT methods can provide satisfactory conformational energy profiles hundreds of times faster in terms of optimization.

Rare earth nanoparticles for sprayed and intravenous NIR II imaging and photodynamic therapy of tongue cancer.

In this research, rare earth nanoparticles coupled with dihydroartemisinin (DHA) and a targeted antibody (RENP-DHA-Cap) for sprayed NIR II imaging and photodynamic therapy (PDT) of tongue cancer were designed. Genetic algorithms combined with combinatorial chemistry were proposed and successfully achieved in a single optimized luminescent phosphor with enhanced NIR II and high upconversion luminescence (UCL) under a NIR laser of wavelength 980 nm or/and 808 nm. In particular, T1 magnetic resonance imaging (MRI) signals can be adjusted with the Gd ion concentration. In combination with the targeted antibody of capmatinib (Cap), precise NIR II imaging for in situ tongue cancer by a simple spray method can be achieved. Most importantly, NIR II imaging and PDT treatment can be realized with RENP-DHA-capmatinib injected intravenously. This orthogonal theranostic mode with precise diagnosis under 808 nm and targeted effective treatment under 980 nm may promote tongue cancer theranostics.