CFAP58 is involved in the sperm head shaping and flagellogenesis of cattle and mice.

CFAP58 is a testis-enriched gene that plays an important role in the sperm flagellogenesis of humans and mice. However, the effect of CFAP58 on bull semen quality and the underlying molecular mechanisms involved in spermatogenesis remain unknown. Here, we identified two single-nucleotide polymorphisms (rs110610797, A>G and rs133760846, G>T) and one indel (g.-1811_ g.-1810 ins147bp) in the promoter of CFAP58 that were significantly associated with semen quality of bulls, including sperm deformity rate and ejaculate volume. Moreover, by generating gene knockout mice, we found for the first time that the loss of Cfap58 not only causes severe defects in the sperm tail, but also affects the manchette structure, resulting in abnormal sperm head shaping. Cfap58 deficiency causes an increase in spermatozoa apoptosis. Further experiments confirmed that CFAP58 interacts with IFT88 and CCDC42. Moreover, it may be a transported cargo protein that plays a role in stabilizing other cargo proteins, such as CCDC42, in the intra-manchette transport/intra-flagellar transport pathway. Collectively, our findings reveal that CFAP58 is required for spermatogenesis and provide genetic markers for evaluating semen quality in cattle.

Predicting the target landscape of kinase inhibitors using 3D convolutional neural networks.

Many therapies in clinical trials are based on single drug-single target relationships. To further extend this concept to multi-target approaches using multi-targeted drugs, we developed a machine learning pipeline to unravel the target landscape of kinase inhibitors. This pipeline, which we call 3D-KINEssence, uses a new type of protein fingerprints (3D FP) based on the structure of kinases generated through a 3D convolutional neural network (3D-CNN). These 3D-CNN kinase fingerprints were matched to molecular Morgan fingerprints to predict the targets of each respective kinase inhibitor based on available bioactivity data. The performance of the pipeline was evaluated on two test sets: a sparse drug-target set where each drug is matched in most cases to a single target and also on a densely-covered drug-target set where each drug is matched to most if not all targets. This latter set is more challenging to train, given its non-exclusive character. Our model's root-mean-square error (RMSE) based on the two datasets was 0.68 and 0.8, respectively. These results indicate that 3D FP can predict the target landscape of kinase inhibitors at around 0.8 log units of bioactivity. Our strategy can be utilized in proteochemometric or chemogenomic workflows by consolidating the target landscape of kinase inhibitors.

Transcriptomic-based evaluation of trichloroethylene glutathione and cysteine conjugates demonstrate phenotype-dependent stress responses in a panel of human in vitro models.

Environmental or occupational exposure of humans to trichloroethylene (TCE) has been associated with different extrahepatic toxic effects, including nephrotoxicity and neurotoxicity. Bioactivation of TCE via the glutathione (GSH) conjugation pathway has been proposed as underlying mechanism, although only few mechanistic studies have used cell models of human origin. In this study, six human derived cell models were evaluated as in vitro models representing potential target tissues of TCE-conjugates: RPTEC/TERT1 (kidney), HepaRG (liver), HUVEC/TERT2 (vascular endothelial), LUHMES (neuronal, dopaminergic), human induced pluripotent stem cells (hiPSC) derived peripheral neurons (UKN5) and hiPSC-derived differentiated brain cortical cultures containing all subtypes of neurons and astrocytes (BCC42). A high throughput transcriptomic screening, utilizing mRNA templated oligo-sequencing (TempO-Seq), was used to study transcriptomic effects after exposure to TCE-conjugates. Cells were exposed to a wide range of concentrations of S-(1,2-trans-dichlorovinyl)glutathione (1,2-DCVG), S-(1,2-trans-dichlorovinyl)-L-cysteine (1,2-DCVC), S-(2,2-dichlorovinyl)glutathione (2,2-DCVG), and S-(2,2-dichlorovinyl)-L-cysteine (2,2-DCVC). 1,2-DCVC caused stress responses belonging to the Nrf2 pathway and Unfolded protein response in all the tested models but to different extents. The renal model was the most sensitive model to both 1,2-DCVC and 1,2-DCVG, with an early Nrf2-response at 3 µM and hundreds of differentially expressed genes at higher concentrations. Exposure to 2,2-DCVG and 2,2-DCVC also resulted in the upregulation of Nrf2 pathway genes in RPTEC/TERT1 although at higher concentrations. Of the three neuronal models, both the LUHMES and BCC42 showed significant Nrf2-responses and at higher concentration UPR-responses, supporting recent hypotheses that 1,2-DCVC may be involved in neurotoxic effects of TCE. The cell models with the highest expression of γ-glutamyltransferase (GGT) enzymes, showed cellular responses to both 1,2-DCVG and 1,2-DCVC. Little to no effects were found in the neuronal models from 1,2-DCVG exposure due to their low GGT-expression. This study expands our knowledge on tissue specificity of TCE S-conjugates and emphasizes the value of human cell models together with transcriptomics for such mechanistic studies.

Dioxin-like polychlorinated biphenyl 126 (PCB126) disrupts gut microbiota-host metabolic dysfunction in mice via aryl hydrocarbon receptor activation.

Exposure to environmental pollutants, including dioxin-like pollutants, can cause numerous health issues. A common exposure route to pollutants is through contaminated foods, and thus the gastrointestinal system and gut microbiota are often exposed to high amounts of pollutants. Multiple studies have focused on the imbalance in intestinal microbiota composition caused by dioxin-like pollutants. Here, we examined the effects of polychlorinated biphenyl 126 (PCB126) on the composition and functions of gut microbes through metagenomic sequencing, and explored the correlations between microflora dysbiosis and aryl hydrocarbon receptor (AHR) signaling. Adult male wild-type and Ahr-/- mice with a C57BL/6 background were weekly exposed to 50 µg/kg body weight of PCB126 for 8 weeks. Results showed that PCB126 had the opposite effect on gut microbiota composition and diversity in the wild-type and Ahr-/- mice. Functional prediction found that PCB126 exposure mainly altered carbon metabolism and signal regulatory pathways in wild-type mice but impacted DNA replication and lipopolysaccharide biosynthesis in Ahr-/- mice. In wild-type mice, PCB126 exposure induced liver injury, decreased serum lipid content, and delayed gastrointestinal motility, which were significantly correlated to several specific bacterial taxa, such as Helicobacter. Following AHR knockout, however, the holistic effects of PCB126 on the host were lessened or abolished. These results suggest that PCB126 may disrupt host metabolism and gut microbiota dynamics via AHR activation. Overall, our findings provide new insight into the complex interactions between host metabolism and gut microbiota, which may contribute to grouped assessment of environmental pollutants in the future.

Genome-wide methylation and transcriptome of blood neutrophils reveal the roles of DNA methylation in affecting transcription of protein-coding genes and miRNAs in E. coli-infected mastitis cows.

BACKGROUND: Neutrophils are the first effectors of inflammatory response triggered by mastitis infection, and are important defense cells against pathogenic Escherichia coli (E. coli). DNA methylation, as a critical epigenetic mechanism for regulating gene function, is involved in bovine mastitis. RESULTS: In this study, we sequenced the blood neutrophils of healthy and E. coli-infected mastitic half-sib cows for the overall DNA methylation levels using transcriptome sequencing and reduced representation bisulfite sequencing. The methylation levels in the mastitis cows (MCs) were decreased compared with healthy cows (HCs). A total of 494 differentially methylated regions were identified, among which 61 were up-methylated and 433 were down-methylated (MCs vs. HCs). The expression levels of 1094 differentially expressed genes were up-regulated, and 245 genes were down-regulated. Twenty-nine genes were found in methylation and transcription data, among which seven genes' promoter methylation levels were negatively correlated with expression levels, and 11 genes were differentially methylated in the exon regions. The bisulfite sequencing PCR and quantitative real-time PCR validation results demonstrated that the promoter methylation of CITED2 and SLC40A1 genes affected differential expression. The methylation of LGR4 exon 5 regulated its own alternative splicing. The promoter methylation of bta-miR-15a has an indirect effect on the expression of its target gene CD163. The CITED2, SLC40A1, and LGR4 genes can be used as candidates for E. coli-induced mastitis resistance. CONCLUSIONS: This study explored the roles of DNA methylation in affecting transcription of protein-coding genes and miRNAs in E. coli-induced mastitis, thereby helping explain the function of DNA methylation in the pathogenesis of mastitis and provided new target genes and epigenetic markers for mastitis resistance breeding in dairy cattle.

In silico genome-wide miRNA-QTL-SNPs analyses identify a functional SNP associated with mastitis in Holsteins.

BACKGROUND: Single-nucleotide polymorphisms (SNPs) in microRNAs (miRNAs) and their target binding sites affect miRNA function and are involved in biological processes and diseases, including bovine mastitis, a frequent inflammatory disease. Our previous study has shown that bta-miR-2899 is significantly upregulated in the mammary gland tissue of mastitis-infected cow than that of healthy cows. RESULTS: In the present study, we used a customized miRNAQTLsnp software and identified 5252 SNPs in 691 bovine pre-miRNAs, which are also located within the quantitative trait loci (QTLs) that are associated with mastitis and udder conformation-related traits. Using luciferase assay in the bovine mammary epithelial cells, we confirmed a candidate SNP (rs109462250, g. 42,198,087 G > A) in the seed region of bta-miR-2899 located in the somatic cell score (SCS)-related QTL (Chr.18: 33.9-43.9 Mbp), which affected the interaction of bta-miR-2899 and its putative target Spi-1 proto-oncogene (SPI1), a pivotal regulator in the innate and adaptive immune systems. Quantitative real-time polymerase chain reaction results showed that the relative expression of SPI1 in the mammary gland of AA genotype cows was significantly higher than that of GG genotype cows. The SNP genotypes were associated with SCS in Holstein cows. CONCLUSIONS: Altogether, miRNA-related SNPs, which influence the susceptibility to mastitis, are one of the plausible mechanisms underlying mastitis via modulating the interaction of miRNAs and immune-related genes. These miRNA-QTL-SNPs, such as the SNP (rs109462250) of bta-miR-2899 may have implication for the mastitis resistance breeding program in Holstein cattle.

Association studies on the bovine lipoprotein lipase gene polymorphism with growth and carcass quality traits in Qinchuan cattle.

Lipoprotein lipase (LPL) is considered as an essential enzyme in lipid deposition and tissue metabolism. It has been proposed to be a lead candidate gene for genetic markers of lipid deposition and energy balance. In this paper, polymorphisms in the LPL gene were investigated in 554 Chinese Qinchuan cattle by PCR-RFLP and DNA sequencing. Seven single nucleotide polymorphisms (SNPs) were identified, which included one mutation (g.91C > T) in the 5'untranslated region (UTR), four synonymous mutations (g.17015A > G, g.18362G > A, g.18377T > C and g.19873T > C) and two mutations (g.25225A > G and g.25316T > G) in the 3'UTR. The frequencies of SNP g.18377T > C and g.25316T > G were skewed from Hardy-Weinberg equilibrium in all the samples (chi-square test, P < 0.05). An association analysis showed that five loci (except for g.91C > T and g.18377T > C) were significantly correlated with some growth and carcass quality traits. These results demonstrate that LPL might be a potential candidate gene for marker-assisted selection (MAS).

Haplotype distribution in the class I sirtuin genes and their associations with ultrasound carcass traits in Qinchuan cattle (Bos taurus).

Class I sirtuin genes including SIRT1, SIRT2 and SIRT3, are members of the nicotinamide adenine dinucleotide (NAD)-dependent family of histone deacetylases, and play essential roles in senescence, metabolism, and apoptosis. This study was conducted to detect potential polymorphisms of the bovine class I sirtuin genes and explore their relationships with ultrasound carcass traits in Qinchuan cattle. Four non-coding mutations in the 3'UTR (SIRT1: g.25751A > C, SIRT1: g.25846A > G, SIRT2: g.19676G > A and SIRT3: g. 25702C > T) and three mutations in exons (SIRT2: g.4062C > T; SIRT2: g.4406C > T and SIRT3: g.25557A > G) were identified in 468 individuals of Qinchuan cattle. Chi-square tests showed that g.25751A > C, g.19676G > A, and g.25702C > T were in Hardy-Weinberg disequilibrium (χ(2) < χ0.05(2)). The statistical analyses indicated that six SNPs were significantly associated with the ultrasound carcass traits (P < 0.05) except g.4062C > T (SIRT2) (P > 0.05). These results indicate that the variations in the class I sirtuin genes and their corresponding genotypes may be considered as molecular markers for economic traits in cattle breeding.

Expression of the SIRT2 gene and its relationship with body size traits in Qinchuan cattle (Bos taurus).

Silent information regulator 2 (SIRT2) is a member of the sirtuin family of class III NAD (nicotinamide adenine dinucleotide)-dependent protein deacetylases and may regulate senescence, metabolism and apoptosis. The aims of this study were to investigate whether the SIRT2 gene could be used as a candidate gene in the breeding of Qinchuan cattle. Real-time polymerase chain reaction (RT-PCR) results showed that among all types of tissue that were analyzed, the highest mRNA expression levels of the gene were found in subcutaneous fat. DNA sequencing of 468 individual Qinchuan cattle identified two novel, single nucleotide polymorphisms (g.19501 C > T and g.19518 C > T) in the 3' untranslated region (3'UTR) of the SIRT2 gene. The frequencies of SNP g.19501 C > T and g.19518 C > T were in Hardy-Weinberg disequilibrium in all the samples (chi-square test, χ2 < χ0.052). An association analysis showed that the two loci were significantly correlated with some body size traits and the H2H2 (-CT-CT-) diplotypes performed better than other combinations. These results indicated that the variations in the SIRT2 gene and their corresponding genotypes may be considered as molecular markers for economic traits in cattle breeding.

Molecular Characterization of Bovine SMO Gene and Effects of Its Genetic Variations on Body Size Traits in Qinchuan Cattle (Bos taurus).

Smoothened (Smo)-mediated Hedgehog (Hh) signaling pathway governs the patterning, morphogenesis and growth of many different regions within animal body plans. This study evaluated the effects of genetic variations of the bovine SMO gene on economically important body size traits in Chinese Qinchuan cattle. Altogether, eight single nucleotide polymorphisms (SNPs: 1-8) were identified and genotyped via direct sequencing covering most of the coding region and 3'UTR of the bovine SMO gene. Both the p.698Ser.>Ser. synonymous mutation resulted from SNP1 and the p.700Ser.>Pro. non-synonymous mutation caused by SNP2 mapped to the intracellular C-terminal tail of bovine Smo protein; the other six SNPs were non-coding variants located in the 3'UTR. The linkage disequilibrium was analyzed, and five haplotypes were discovered in 520 Qinchuan cattle. Association analyses showed that SNP2, SNP3/5, SNP4 and SNP6/7 were significantly associated with some body size traits (p < 0.05) except SNP1/8 (p > 0.05). Meanwhile, cattle with wild-type combined haplotype Hap1/Hap1 had significantly (p < 0.05) greater body length than those with Hap2/Hap2. Our results indicate that variations in the SMO gene could affect body size traits of Qinchuan cattle, and the wild-type haplotype Hap1 together with the wild-type alleles of these detected SNPs in the SMO gene could be used to breed cattle with superior body size traits. Therefore, our results could be helpful for marker-assisted selection in beef cattle breeding programs.

Molecular characterization, expression profiles, and analysis of Qinchuan cattle SIRT1 gene association with meat quality and body measurement traits (Bos taurus).

Silent information regulator (SIRT1), was closely associated with senescence, metabolism, and apoptosis. The objective of this study was to investigate whether SIRT1 gene could be used as a candidate gene in the breeding process of Qinchuan cattle. Via sequencing technology conducted in 453 individuals of Qinchuan cattle, single nucleotide polymorphisms (G25764A, A25846G, and T25868C) with 5 haplotypes and 6 combined genotypes in 3' untranslated region of SIRT1 gene were identified. In addition, three loci were significantly associated with some of the body measurements and meat quality traits in Qinchuan cattle (P < 0.05), and the H2H2 (GG-AA-CC) diplotypes had better performance than other combinations in Qinchuan cattle. These results suggest that the SIRT1 gene could be used in marker assisted selection to improve the production traits of Qinchuan cattle.

Ultra-sensitive detection of SARS-CoV-2 S1 protein by coupling rolling circle amplification with poly(N-isopropylacrylamide)-based sandwich-type assay.

In the past few years, the COVID-19 pandemic, caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) seriously threatens global public health security due to its high contagiousness. It remains of vital importance to develop a rapid and sensitive assay for SARS-CoV-2. In this work, we proposed a sandwich-type assay based on poly(N-isopropylacrylamide) (PNIPAM), allowing efficient detection of the SARS-CoV-2 S1 protein in the homogeneous solution. Firstly, a direct sandwich-type assay was established with a linear range of 0.2-2 µg/mL and a limit of detection (LOD) of 0.11 µg/mL, which could realize rapid detection in about 1 h. Furthermore, the sandwich-type assay coupled with rolling circle amplification (RCA) obtained an increase in sensitivity of 5.9 × 104 folds with a wide linear range of 0.01 - 100 ng/mL and a LOD of 1.88 pg/mL. The average recoveries in unpretreated saliva were 90 %-113.0 %, indicating the potential of the developed method for application in practical samples. Given the high selectivity and sensitivity of the developed method, it has a significant potential for rapid and early detection of SARS-CoV-2.

The adaptation of bumblebees to extremely high elevation associated with their gut microbiota.

Bumblebees are among the most abundant and important pollinators for sub-alpine and alpine flowering plant species in the Northern Hemisphere, but little is known about their adaptations to high elevations. In this article, we focused on two bumblebee species, Bombus friseanus and Bombus prshewalskyi, and their respective gut microbiota. The two species, distributed through the Hengduan Mountains of southwestern China, show species replacement at different elevations. We performed genome sequencing based on 20 worker bee samples of each species. Applying evolutionary population genetics and metagenomic approaches, we detected genes under selection and analyzed functional pathways between bumblebees and their gut microbes. We found clear genetic differentiation between the two host species and significant differences in their microbiota. Species replacement occurred in both hosts and their bacteria (Snodgrassella) with an increase in elevation. These extremely high-elevation bumblebees show evidence of positive selection related to diverse biological processes. Positively selected genes involved in host immune systems probably contributed to gut microbiota changes, while the butyrate generated by gut microbiota may influence both host energy metabolism and immune systems. This suggests a close association between the genomes of the host species and their microbiomes based on some degree of natural selection.IMPORTANCETwo closely related and dominant bumblebee species, distributed at different elevations through the Hengduan Mountains of southwestern China, showed a clear genomic signature of adaptation to elevation at the molecular level and significant differences in their respective microbiota. Species replacement occurred in both hosts and their bacteria (Snodgrassella) with an increase in elevation. Bumblebees' adaptations to higher elevations are closely associated with their gut microbiota through two biological processes: energy metabolism and immune response. Information allowing us to understand the adaptive mechanisms of species to extreme conditions is implicit if we are to conserve them as their environments change.

A simple, rapid and sensitive sandwich immunoassay based on poly(N-isopropylacrylamide) for the detection of alpha-fetoprotein.

Alpha-fetoprotein (AFP), as a tumor marker, plays a vital role in the diagnosis of liver cancer. In this work, a novel sandwich immunoassay based on a thermosensitive polymer, poly(N-isopropylacrylamide) (PNIPAM), was developed for the detection of AFP. This immunoassay could realize one-step rapid reaction within 1 h, and facilitate the separation of the target molecules by incorporating PNIPAM. In this method, a conjugate of PNIPAM and capture antibody (Ab1) was successfully synthesized as a capture probe and the synthetic method of PNIPAM-Ab1 was simple, while the detection antibody (Ab2) was labeled with fluorescein isothiocyanate (FITC) to form a fluorescent detection probe. By employing a sandwich immunoassay, the method achieved quantitative determination of AFP, exhibiting a wide linear range from 5 ng/mL to 200 ng/mL and a low detection limit of 2.44 ng/mL. Furthermore, it was successfully applied to the analysis of spiked human serum samples and the screening of patients with hepatic diseases in clinical samples, indicating its potential application prospect in the diagnosis of liver cancer.

Non-Invasive Detection of Early-Stage Fatty Liver Disease via an On-Skin Impedance Sensor and Attention-Based Deep Learning.

Early-stage nonalcoholic fatty liver disease (NAFLD) is a silent condition, with most cases going undiagnosed, potentially progressing to liver cirrhosis and cancer. A non-invasive and cost-effective detection method for early-stage NAFLD detection is a public health priority but challenging. In this study, an adhesive, soft on-skin sensor with low electrode-skin contact impedance for early-stage NAFLD detection is fabricated. A method is developed to synthesize platinum nanoparticles and reduced graphene quantum dots onto the on-skin sensor to reduce electrode-skin contact impedance by increasing double-layer capacitance, thereby enhancing detection accuracy. Furthermore, an attention-based deep learning algorithm is introduced to differentiate impedance signals associated with early-stage NAFLD in high-fat-diet-fed low-density lipoprotein receptor knockout (Ldlr-/-) mice compared to healthy controls. The integration of an adhesive, soft on-skin sensor with low electrode-skin contact impedance and the attention-based deep learning algorithm significantly enhances the detection accuracy for early-stage NAFLD, achieving a rate above 97.5% with an area under the receiver operating characteristic curve (AUC) of 1.0. The findings present a non-invasive approach for early-stage NAFLD detection and display a strategy for improved early detection through on-skin electronics and deep learning.

Characterization key genes of Arabidopsis seedlings in response to ß-caryophyllene, eugenol using combined transcriptome and WGCN analysis.

Weeds present a significant challenge to high crop yield and quality. In our study, we investigated the phytotoxic activity of ß-caryophyllene (BCP) and eugenol, which are natural allelopathic chemical compounds, on Arabidopsis seedlings. We found that these compounds inhibited the growth of Arabidopsis thaliana plants. When either BCP or eugenol was applied, it led to decrease in the content of cell wall components such as lignin, cellulose, hemicellulose, and pectin; and increase in the levels of endogenous hormones like ETH, ABA, SA, and JA in the seedlings. Through transcriptome profiling, we identified 7181 differentially expressed genes (DEGs) in the roots and shoots that were induced by BCP or eugenol. The genes involved in the synthesis of lignin, cellulose, hemicellulose, and pectin were down-regulated, whereas genes related to synthesis and signal transduction of ABA, ETH, SA, and JA were up-regulated. However, genes related to IAA synthesis and signal transduction were found to be down-regulated. Furthermore, we characterized 24 hub genes using Weighted Correlation Network Analysis (WGCNA). Among them, the identified 16 genes in response to BCP was primarily associated with hypoxia stress, while 8 genes induced by eugenol were linked to inhibition of cell division. Our results suggested that BCP and eugenol had ability to target multiple genes to inhibit growth and development of Arabidopsis plants. Therefore, they can serve as excellent candidates for natural biological herbicides.

Signatures of selection in indigenous Chinese cattle genomes reveal adaptive genes and genetic variations to cold climate.

Cold climate shapes the genome of animals and drives them to carry sufficient genetic variations to adapt to changes in temperature. However, limited information is available about the genome-wide pattern of adaptations to cold environments in cattle. In the present study, we used 777K SNP genotyping (15 Chinese cattle breeds, 198 individuals) and whole genome resequencing data (54 cattle breeds of the world, 432 individuals) to disentangle divergent selection signatures, especially between the cold-adapted (annual average temperature of habitat, 6.24 °C to 10.3 °C) and heat-adapted (20.2 °C to 24.73 °C) Chinese native cattle breeds. Genomic analyses revealed a set of candidate genes (e.g., UQCR11, DNAJC18, EGR1, and STING1) were functionally associated with thermogenesis and energy metabolism. We also characterized the adaptive loci of cattle exposed to cold temperatures. Our study finds new candidate genes and provides new insights into adaptations to cold climates in cattle.

Cold climates can affect cattle performance, survival, and health. Local cattle breeds have been adapted to the local environments including extremely cold temperatures after a long period of natural and artificial selection. Selection and local adaptation are shaping populations. However, identifying loci associated with cold adaptation has been a major challenge. We used high-density SNP arrays and resequencing data to comprehensively analyze and compare the genomic selection signatures of Chinese northern and southern cattle, and elucidated several adaptive genes and alleles involved in cold adaptation. The complexity of genetic adaptation mechanism among different low-temperature adapted cattle breeds was also emphasized.

Expression of Tryptophan Metabolism Enzymes in Patients with Diffuse Large B-cell Lymphoma and NK/T-cell Lymphoma.

BACKGROUND: Metabolites of tryptophan (Trp) metabolism in the tumor microenvironment play crucial immunosuppressive roles in various cancers. However, the role of Trp metabolism in diffuse large B-cell lymphoma (DLBCL) or natural killer/T-cell lymphoma (NK/TCL) remains unelucidated. METHODS: We investigated the potential role of Trp metabolism in a cohort of 43 patients with DLBCL and 23 with NK/TCL. We constructed tissue microarrays and performed in situ staining of Trp-catabolizing enzymes and PD-L1 using immunohistochemistry (IHC). RESULTS: We observed 14.0% positive staining of IDO1 in DCBCL and 60.9% in NK/TCL; 55.8% of IDO2 in DCBCL and 95.7% in NK/TCL; 79.1% of TDO2 in DCBCL and 43.5% in NK/TCL; 29.7% of IL4I1 in DCBCL and 39.1% in NK/TCL. However, IDO1, IDO2, TDO2, and IL4I1 positivity did not significantly differ between PD-L1+ and PD-L1- biopsy tissue samples of NK/TCL; nonetheless, a positive correlation of IDO1 (r = 0.87, p < 0.001), IDO2 (r = 0.70, p < 0.001), TDO2 (r = 0.63, p < 0.001), and IL4I1 (r = 0.53, p < 0.05) with PD-L1 expression was observed in the TCGA-DLBCL dataset. Finally, immunohistochemical (IHC) analysis revealed the lack of superior prognostic effect with higher expression of Trp enzymes in DLBCL and NK/TCL. Furthermore, IDO1, IDO2, TDO2, and IL4I1 expression, as well as survival rates, did not significantly differ across all groups in the TCGA-DLBCL cohort. CONCLUSION: Collectively, our findings provide novel insights into the enzymes involved in Trp metabolism in DLBCL and NK/TCL and their association with PD-L1 expression, which offers potential strategies to combine Trp-metabolism enzyme inhibitors with anti-PD-L1 or other immunotherapeutic strategies in clinical DLBCL or NK/TCL treatment.

Coordinated alternation of DNA methylation and alternative splicing of PBRM1 affect bovine sperm structure and motility.

DNA methylation and gene alternative splicing drive spermatogenesis. In screening DNA methylation markers and transcripts related to sperm motility, semen from three pairs of full-sibling Holstein bulls with high and low motility was subjected to reduced representation bisulphite sequencing. A total of 948 DMRs were found in 874 genes (gDMRs). Approximately 89% of gDMR-related genes harboured alternative splicing events, including SMAD2, KIF17, and PBRM1. One DMR in exon 29 of PBRM1 with the highest 5mC ratio was found, and hypermethylation in this region was related to bull sperm motility. Furthermore, alternative splicing events at exon 29 of PBRM1 were found in bull testis, including PBRM1-complete, PBRM1-SV1 (exon 28 deletion), and PBRM1-SV2 (exons 28-29 deletion). PBRM1-SV2 exhibited significantly higher expression in adult bull testes than in newborn bull testes. In addition, PBRM1 was localized to the redundant nuclear membrane of bull sperm, which might be related to sperm motility caused by sperm tail breakage. Therefore, the hypermethylation of exon 29 may be associated with the production of PBRM1-SV2 in spermatogenesis. These findings indicated that DNA methylation alteration at specific loci could regulate gene splicing and expression and synergistically alter sperm structure and motility.

Exploring the cellular and molecular differences between ovarian clear cell carcinoma and high-grade serous carcinoma using single-cell RNA sequencing and GEO gene expression signatures.

The two most prevalent subtypes of epithelial ovarian carcinoma (EOC) are ovarian clear cell carcinoma (OCCC) and high-grade serous ovarian carcinoma (HGSC). Patients with OCCC have a poor prognosis than those with HGSC due to chemoresistance, implying the need for novel treatment target. In this study, we applied single-cell RNA sequencing (scRNA-seq) together with bulk RNA-seq data from the GEO (Gene Expression Omnibus) database (the GSE189553 dataset) to characterize and compare tumor heterogeneity and cell-level evolution between OCCC and HGSC samples. To begin, we found that the smaller proportion of an epithelial OCCC cell subset in the G2/M phase might explain OCCC chemoresistance. Second, we identified a possible pathogenic OCCC epithelial cell subcluster that overexpresses LEFTY1. Third, novel biomarkers separating OCCC from HGSC were discovered and subsequently validated on a wide scale using immunohistochemistry. Amine oxidase copper containing 1 (AOC1) was preferentially expressed in OCCC over HGSC, while S100 calcium-binding protein A2 (S100A2) was detected less frequently in OCCC than in HGSC. In addition, we discovered that metabolic pathways were enriched in the epithelial compartment of the OCCC samples. In vitro experiments verified that inhibition of oxidative phosphorylation or glycolysis pathways exerted direct antitumor effects on both OCCC and HGSC cells, while targeting glutamine metabolism or ferroptosis greatly attenuated chemosensitivity only in OCCC cells. Finally, to determine whether there were any variations in immune cell subsets between OCCC and HGSC, data from scRNA-seq and mass cytometry were pooled for analysis. In summary, our work provides the first holistic insights into the cellular and molecular distinctions between OCCC and HGSC and is a valuable source for discovering new targets to leverage in clinical treatments to improve the poor prognosis of patients with OCCC.