The ERK inhibitor GDC-0994 selectively inhibits growth of BRAF mutant cancer cells.

BRAF or RAS mutation-induced aberrant activation of the mitogen-activated protein kinase (MAPK) pathway is frequently observed in human cancers. As the key downstream node of MAPK pathway, ERK1/2 is as an important therapeutic target. GDC-0994 (ravoxertinib), an orally bioavailable, highly selective small-molecule inhibitor of ERK1/2, showed acceptable safety and pharmacodynamic profile in a recent phase I clinical trial. In this study, we investigated dependence of the anti-tumor effect of ERK inhibitor GDC-0994 on genetic alterations in the MAPK pathway. The results showed that GDC-0994 sharply inhibited cell proliferation and colony formation and induced remarkable G1 phase cell-cycle arrest in cancer cells harboring BRAF mutation but had little effect on cell behaviors in most RAS mutant or wild-type cell lines. The expression of a large number of genes, particularly the genes in the cell cycle pathway, were significantly changed after GDC-0994 treatment in BRAF mutant cells, while no remarkable expression change of such genes was observed in wild-type cells. Moreover, GDC-0994 selectively inhibited tumor growth in a BRAF mutant xenograft mice model. Our findings demonstrate a BRAF mutation-dependent anti-tumor effect of GDC-0994 and provide a rational strategy for patient selection for ERK1/2 inhibitor treatment.

Therapeutic targeting of PLK1 in TERT promoter-mutant hepatocellular carcinoma.

BACKGROUND: Hotspot mutations in the promoter of telomerase reverse transcriptase (TERT) gene are the most common genetic variants in hepatocellular carcinoma (HCC) and associated with poor prognosis of the disease. However, no drug was currently approved for treating TERT promoter mutation positive HCC patients. Here, we aim to explore the potential therapeutic strategy for targeting TERT promoter mutation in HCC. METHODS: The Liver Cancer Model Repository database was used for screening potential drugs to selectively suppress the growth of TERT promoter mutant HCC cells. RNA-seq, CRISPR-Cas9 technology and siRNA transfection were performed for mechanistic studies. Cell counting kit-8 (CCK8) assay and the xenograft tumour models were used for cell growth detection in vitro and in vivo, respectively. Cell apoptosis and cell cycle arrest were analysed by Annexin V-FITC staining and/or propidium iodide staining. RESULTS: PLK1 inhibitors were remarkably more sensitive to HCC cells harbouring TERT promoter mutation than wild-type cells in vitro and in vivo, which were diminished after TERT promoter mutation was edited to the wild-type nucleotide. Comparing the HCC cells with wild-type promoter of TERT, PLK1 inhibitors specifically downregulated Smad3 to regulate TERT for inducing apoptosis and G2/M arrest in TERT mutant HCC cells. Moreover, knockout of Smad3 counteracted the effects of PLK1 inhibitors in TERT mutant HCC cells. Finally, a cooperative effect of PLK1 and Smad3 inhibition was observed in TERT mutant cells. CONCLUSIONS: PLK1 inhibition selectively suppressed the growth of TERT mutant HCC cells through Smad3, thus contributed to discover a novel therapeutic strategy to treat HCC patients harbouring TERT promoter mutations. KEY POINTS: TERT promoter mutation confers sensitivity to PLK1 inhibitors in HCC. The selective growth inhibition of TERT mutant HCC cells induced by PLK1 inhibitor was mediated by Smad3. Combined inhibition of PLK1 and Smad3 showed a cooperative anti-tumor effect in TERT mutant HCC cells.

Risk stratification by combining common genetic mutations and TERT promoter methylation in papillary thyroid cancer.

PURPOSE: Risk stratification based on somatic mutations in TERT promoter and BRAF/RAS has been well established for papillary thyroid cancer (PTC), and there is emerging evidence showed that TERT promoter methylation was frequently observed in thyroid cancer patients with adverse features. This study was aimed to comprehensive explore the prognostic value of BRAF/RAS mutations, TERT promoter mutations, and TERT promoter methylation in PTC. METHODS: The relationships of BRAF/RAS mutations, TERT promoter mutations, and TERT promoter methylation with clinical characteristics and outcomes of PTC were analyzed in 382 patients with PTC. RESULTS: TERT promoter mutation and hypermethylation were collectively observed in 52 (13.6%) samples and associated with BRAF/RAS mutation, aggressive clinical characteristics, and poor clinical outcomes of PTC. Coexistence of BRAF/RAS and TERT alterations was found in 45 of 382 (11.8%) PTC patients and strongly associated with old patient age, extrathyroidal extension, advanced pathologic T stage and metastasis. Importantly, patients with both BRAF/RAS and TERT alterations had higher rates of tumor recurrence (13.6% vs 1.5%, P = 0.042) and disease progression (24.4% vs 3.3%, P < 0.001) than patients without any alterations, and cox regression analysis revealed that the coexistence of BRAF/RAS and TERT alterations, but not BRAF/RAS or TERT alterations alone, increased the risk of progression-free interval with an adjusted HR of 10.35 (95% CI: 1.79-59.81, P = 0.009). CONCLUSIONS: This study suggested that comprehensively analysis of BRAF/RAS mutations, TERT promoter mutation and methylation is an effective strategy to identify high-risk patients with PTC.

Towards precision oncology discovery: four less known genes and their unknown interactions as highest-performed biomarkers for colorectal cancer.

The goal of this study was to use a new interpretable machine-learning framework based on max-logistic competing risk factor models to identify a parsimonious set of differentially expressed genes (DEGs) that play a pivotal role in the development of colorectal cancer (CRC). Transcriptome data from nine public datasets were analyzed, and a new Chinese cohort was collected to validate the findings. The study discovered a set of four critical DEGs - CXCL8, PSMC2, APP, and SLC20A1 - that exhibit the highest accuracy in detecting CRC in diverse populations and ethnicities. Notably, PSMC2 and CXCL8 appear to play a central role in CRC, and CXCL8 alone could potentially serve as an early-stage marker for CRC. This work represents a pioneering effort in applying the max-logistic competing risk factor model to identify critical genes for human malignancies, and the interpretability and reproducibility of the results across diverse populations suggests that the four DEGs identified can provide a comprehensive description of the transcriptomic features of CRC. The practical implications of this research include the potential for personalized risk assessment and precision diagnosis and tailored treatment plans for patients.

Integration of risk variants from GWAS with SARS-CoV-2 RNA interactome prioritizes FUBP1 and RAB2A as risk genes for COVID-19.

The role of host genetic factors in COVID-19 outcomes remains unclear despite various genome-wide association studies (GWAS). We annotate all significant variants and those variants in high LD (R2 > 0.8) from the COVID-19 host genetics initiative (HGI) and identify risk genes by recognizing genes intolerant nonsynonymous mutations in coding regions and genes associated with cis-expression quantitative trait loci (cis-eQTL) in non-coding regions. These genes are enriched in the immune response pathway and viral life cycle. It has been found that host RNA binding proteins (RBPs) participate in different phases of the SARS-CoV-2 life cycle. We collect 503 RBPs that interact with SARS-CoV-2 RNA concluded from in vitro studies. Combining risk genes from the HGI with RBPs, we identify two COVID-19 risk loci that regulate the expression levels of FUBP1 and RAB2A in the lung. Due to the risk allele, COVID-19 patients show downregulation of FUBP1 and upregulation of RAB2A. Using single-cell RNA sequencing data, we show that FUBP1 and RAB2A are expressed in SARS-CoV-2-infected upper respiratory tract epithelial cells. We further identify NC_000001.11:g.77984833C>A and NC_000008.11:g.60559280T>C as functional variants by surveying allele-specific transcription factor sites and cis-regulatory elements and performing motif analysis. To sum up, our research, which associates human genetics with expression levels of RBPs, identifies FUBP1 and RAB2A as two risk genes for COVID-19 and reveals the anti-viral role of FUBP1 and the pro-viral role of RAB2A in the infection of SARS-CoV-2.

Exploring the Impact of Insertion/Deletion in FTO and PLIN1 Genes on Morphometric Traits in Sheep.

This study aimed to identify InDels from the FTO and PLIN1 genes and to analyze their association with morphometric traits in Hu sheep (HS), Dupor sheep (DS), and Small Tail Han sheep (STHS). The FTO and PLIN1 genes were genotyped using the insertion/deletion (InDel) method. A one-way ANOVA with SPSS 26.0 software (IBM Corp, Armonk, NY, USA) was used to assess the effect of the InDel FTO and PLIN1 genes on morphometric traits. The results revealed significant associations between certain InDels and the morphometric traits in different breeds of sheep. Specifically, FTO-2 was significantly associated with cannon circumference (CaC) in HS rams and body height (BoH) in HS ewes (p < 0.05). FTO-2 was also significantly associated with chest width (ChW), CaC, head length (HeL), and coccyx length (CoL) in the STHS breed (p < 0.05). FTO-3 showed significant associations with BoH in HS rams and BoH, back height (BaH), ChW, and chest depth (ChD) in HS ewes (p < 0.05). FTO-3 was also significantly associated with ChW in the DS and STHS breeds (p < 0.05). FTO-5 was significantly associated with body weight (BoW) in the DS breed and BoH in the STHS breed (p < 0.05). Furthermore, PLIN1 was significantly related to BoW in the DS breed and was significantly associated with CoL and forehead width (FoW) in the STHS breed (p < 0.05). In conclusion, the study suggested that InDels in the FTO and PLIN1 genes could provide practical information to improve morphometric traits in sheep breeding.

Methylation-Mediated Silencing of ATF3 Promotes Thyroid Cancer Progression by Regulating Prognostic Genes in the MAPK and PI3K/AKT Pathways.

Background: Aberrant expression of oncogenes and/or tumor suppressor genes (TSGs) drives the tumorigenesis and development of thyroid cancer. We investigated the expression and function of a member of the activating transcription factor (ATF)/cAMP-responsive element-binding protein (CREB) transcription factor (TF) family, ATF3, in thyroid cancer. Methods: Data from 80 patients with papillary thyroid cancer (PTC) in the First Affiliated Hospital of Sun Yat-sen University and 510 PTC samples in The Cancer Genome Atlas thyroid cancer database were utilized for gene expression and prognosis analyses. The survival data were analyzed by Kaplan-Meier curves and Cox regression with adjustment for age, sex, multilocality, extrathyroidal extension, lymph metastases, and history of neoadjuvant treatment. DNA methylation was analyzed by methylation-specific polymerase chain reaction (PCR) and bisulfite sequencing PCR. TFs binding to ATF3 promoter were identified by DNA pull-down combined with mass spectrum assay, and confirmed by quantitative PCR (qPCR), luciferase reporter assay, and chromatin immunoprecipitation (ChIP)-qPCR. We conducted functional assays in vitro and in a xenograft mouse model to evaluate the function of ATF3 in thyroid cancer. Integrated analyses based on RNA sequencing, ChIP-seq, and CUT&Tag assays were performed to explore the mechanisms underlying the function of ATF3. Results: ATF3 was significantly downregulated in PTC and patients with low ATF3 expression had reduced progression-free survival (adjusted hazard ratio = 0.50 [CI 0.26-0.98], p = 0.043). DNA hypermethylation in ATF3 promoter disrupted the binding of SP1 and MYC-MAX, leading to inactivation of the gene. ATF3 functioned as a TSG by inhibiting the proliferation and mobility of thyroid cancer cells. And ATF3 regulated the expression of a number of genes by binding to the regulatory elements of them, particularly for genes in MAPK and PI3K/AKT pathways. Among these target genes, filamin C was positively regulated by ATF3 and associated with a more favorable thyroid cancer prognosis, while dual specificity phosphatase 10, fibronectin-1, tenascin C, and CREB5 were negatively regulated by ATF3 and associated with a poorer prognosis. Conclusions: We observed that the promoter DNA hypermethylation decreased the expression of ATF3, which in turn promoted the progression of thyroid cancer, at least partially, by directly regulating prognosis-related genes in the MAPK and PI3K/AKT pathways.

Identified eleven exon variants in PKD1 and PKD2 genes that altered RNA splicing by minigene assay.

BACKGROUND: Autosomal dominant polycystic kidney disease (ADPKD) is a common monogenic multisystem disease caused primarily by mutations in the PKD1 gene or PKD2 gene. There is increasing evidence that some of these variants, which are described as missense, synonymous or nonsense mutations in the literature or databases, may be deleterious by affecting the pre-mRNA splicing process. RESULTS: This study aimed to determine the effect of these PKD1 and PKD2 variants on exon splicing combined with predictive bioinformatics tools and minigene assay. As a result, among the 19 candidate single nucleotide alterations, 11 variants distributed in PKD1 (c.7866C > A, c.7960A > G, c.7979A > T, c.7987C > T, c.11248C > G, c.11251C > T, c.11257C > G, c.11257C > T, c.11346C > T, and c.11393C > G) and PKD2 (c.1480G > T) were identified to result in exon skipping. CONCLUSIONS: We confirmed that 11 variants in the gene of PKD1 and PKD2 affect normal splicing by interfering the recognition of classical splicing sites or by disrupting exon splicing enhancers and generating exon splicing silencers. This is the most comprehensive study to date on pre-mRNA splicing of exonic variants in ADPKD-associated disease-causing genes in consideration of the increasing number of identified variants in PKD1 and PKD2 gene in recent years. These results emphasize the significance of assessing the effect of exon single nucleotide variants in ADPKD at the mRNA level.

EBV-encoded miRNAs BHRF1-1 and BART2-5p aggravate post- transplant lymphoproliferative disorder via LZTS2-PI3K-AKT axis.

Post-transplant lymphoproliferative disorder (PTLD) is one of the most serious complications after transplantation. Epstein-Barr virus (EBV) is a key pathogenic driver of PTLD. About 80% of PTLD patients are EBV positive. However, the accuracy of preventing and diagnosing EBV-PTLD by monitoring EBV DNA load is limited. Therefore, new diagnostic molecular markers are urgently needed. EBV-encoded miRNAs can regulate a variety of EBV-associated tumors and are expected to be potential diagnostic markers and therapeutic targets. We found BHRF1-1 and BART2-5p were significantly elevated in EBV-PTLD patients, functionally promoting proliferation and inhibiting apoptosis in EBV-PTLD. Mechanistically, we first found that LZTS2 acts as a tumor suppressor gene in EBV-PTLD, and BHRF1-1 and BART2-5p can simultaneously inhibit LZTS2 and activate PI3K-AKT pathway. This study shows that BHRF1-1 and BART2-5p can simultaneously inhibit the expression of tumor suppressor LZTS2, and activate the PI3K-AKT pathway, leading to the occurrence and development of EBV-PTLD. Therefore, BHRF1-1 and BART2-5p are expected to be potential diagnostic markers and therapeutic targets for EBV-PTLD patients.

Research Note: Association of VIPR-1 gene polymorphism with growth traits in meat type Japanese quail (Coturnix Japonica).

This study aimed to analyze the polymorphism of the vasoactive intestinal peptide receptor-1 (VIPR-1) gene and its association with growth traits in quail using the PCR-RFLP and sequencing techniques. Genomic DNA was extracted from blood samples of 36 female Savimalt (SV) quails and 49 female French Giant (FG) quails. Growth traits were measured and used for VIPR-1 gene analysis, as body weight (BW), tibia length (TL), chest width (CW), chest depth (CD), sternum length (SL), body length (BL), and tibia circumference (TC). The results showed that 2 SNPs (BsrD I and HpyCH4 IV) were detected in exon 4 to 5 and exon 6 to 7 of the VIPR-1 gene, respectively. The results of association showed that the BsrD I site was not significantly associated with growth traits at 3 or 5 wk of age in the SV strain (P < 0.05), while the BsrD I site was significantly associated with BL at 3 or 5 wk of age in FG (P < 0.05). The HpyCH4 IV site was significantly associated with TL, CW, CD, SL, and BL at 3 wk of age in the SV strain (P < 0.05), while the HpyCH4 IV site was significantly correlated with BW, CW, SL, and BL at 5 wk of age in SV (P < 0.05). The HpyCH4 IV site was significantly associated with TL and TC at 3 wk of age in FG (P < 0.05), while the HpyCH4 IV site was significantly associated with TC at 5 wk of age in FG (P < 0.05). Four haplotype combinations based on 2 SNPs showed significantly association with BW, CW, CD, SL, BL, and TC at 3 or 5 wk of age in SV (P < 0.05). There was not significant association between 3 haplotype combinations with growth trait at 3 or 5 wk of age in FG (P > 0.05). In conclusion, the VIPR-1 gene could be used as a molecular genetic marker to improve growth traits in quail.

The activation of mTOR signalling modulates DNA methylation by enhancing DNMT1 translation in hepatocellular carcinoma.

BACKGROUND: Both dysregulation of mechanistic target of rapamycin (mTOR) signalling and DNA methylation patterns have been shown to be closely associated with tumor progression and serve as promising targets for hepatocellular carcinoma (HCC) therapy. Although their respective roles in HCC have been extensively revealed, the existence of molecular interactions between them remains largely unknown. METHODS: The association of DNA methylation and mTOR signalling in HCC tissues and cell lines was assessed. A Kaplan‒Meier analysis was applied to estimate the overall survival (OS) and recurrence-free survival (RFS) of HCC patients. The modulation of DNMT1 by mTOR in HCC cell lines was determined. The effect of the drug combination in cell lines and mouse models was examined. RESULTS: The results showed that the DNA methylation level was positively associated with the activation of mTOR signalling in HCC tissues and cell lines. Moreover, HCC patients with higher DNA methylation levels and enhanced activation of mTOR signalling exhibited the worst prognosis. Then, we screened methylation-related enzymes and found that the activation of mTOR signalling increased DNMT1 expression and activity. In addition, mTOR enhanced the translational efficiency of DNMT1 in a 4E-BP1-dependent manner, which is based on the pyrimidine rich translational element (PRTE)-containing 5'UTR of DNMT1. Moreover, we demonstrated that the combined inhibition of mTOR and DNMT synergistically inhibited HCC growth in vitro and in vivo. CONCLUSIONS: In addition to some already identified pro-cancer downstream molecules, the activation of mTOR signalling was found to promote DNA methylation by increasing the translation of DNMT1. Furthermore, combined targeting of mTOR and DNMT1 has been demonstrated to have a more effective tumor suppressive function in HCC.

Highly sensitive droplet digital PCR for detection of RET fusion in papillary thyroid cancer.

BACKGROUND: Thyroid cancer is the most frequent malignancy of the endocrine system, of which papillary thyroid cancer (PTC) is the predominant form with a rapid increasing incidence worldwide. Rearranged during transfection (RET) fusions are common genetic drivers of PTC and the potent RET inhibitor selpercatinib has been recently approved for treating advanced or metastatic RET fusion-positive thyroid cancer. In this study we aimed to develop a droplet digital PCR (ddPCR) system to accurately detect RET fusion in PTC samples. METHODS: The frequency and distribution of RET fusions in PTC were analyzed using genomic data of 402 PTC patients in The Cancer Genome Atlas (TCGA) database. To establish the ddPCR system for detecting CCDC6::RET fusion, a plasmid containing CCDC6::RET infusion fragment was constructed as standard template, the annealing temperature and concentrations of primers and probe were optimized. The analytical performance of ddPCR and quantitative reverse transcription PCR (qRT-PCR) were assessed in standard templates and tissue samples from 112 PTC patients. Sanger sequencing was performed in all the RET fusion-positive samples identified by ddPCR. RESULTS: RET fusions were observed in 25 (6.2%) of the 402 TCGA samples, and 15 (60%) of the RET fusion-positive patients had the CCDC6::RET fusion. Compared with qRT-PCR, the ddPCR method showed a lower limit of detection (128.0 and 430.7 copies/reaction for ddPCR and qRT-PCR, respectively). When applying the two methods to 112 tissue samples of PTC, eleven (9.8%) CCDC6::RET fusion-positive samples were detected by qRT-PCR, while ddPCR identified 4 additional positive samples (15/112, 13.4%). All the CCDC6::RET fusion-positive cases identified by ddPCR were confirmed by Sanger sequencing except for one case with 0.14 copies/uL of the fusion. CONCLUSION: The accurate and sensitive ddPCR method reported here is powerful to detection CCDC6::RET fusion in PTC samples, application of this method would benefit more RET fusion-positive patients in the clinic.

Research Note: Association of IGF-1R gene polymorphism with egg quality and carcass traits of quail (Coturnix japonica).

Insulin-like growth factor 1 receptor (IGF-1R) gene is the main effector of insulin-like growth factor (IGF), which plays an important role in growth, development and reproduction of the animal organism. This study aimed to investigate the association of IGF-1R gene single nucleotide polymorphisms (SNPs) with egg quality and carcass traits of quail by direct sequencing. In this study, genomic DNA was extracted from quail blood samples of 46 Chinese yellow (CY) quail, 49 Beijing white (BW) quail and 48 Korean (KO) quail strains. Egg quality and carcass traits were measured and used for IGF-1R gene analysis in 3 quail strains. The results showed that 2 SNPs (A57G and A72T) of the IGF-1R gene were detected in 3 quail strains. The A57G was significantly associated with yolk width (YWI) in BW strain (P < 0.05). Whereas A72T was significantly associated with egg shell thickness (EST) in BW strain (P < 0.05), and significantly associated with egg weight (EW), egg long (EL), and egg short (ES) in KO strain (P < 0.05). Haplotypes based on 2 SNPs showed significant effect on EST in 3 quail strains (P < 0.05), it also has a significant effect on EW in KO strain (P < 0.05). Meanwhile, A72T was significantly associated with liver weight (LW) and dressing percentage (DP) in 3 strains (P < 0.05). Haplotypes showed significant effect on LW (P < 0.05). Therefore, the IGF-1R gene may be a molecular genetic marker to improve egg quality and carcass traits in quails.

SEPT2 crotonylation promotes metastasis and recurrence in hepatocellular carcinoma and is associated with poor survival.

BACKGROUND: Hepatocellular carcinoma (HCC) metastasis and recurrence lead to therapy failure, which are closely associated with the proteome. However, the role of post-translational modification (PTM) in HCC, especially for the recently discovered lysine crotonylation (Kcr), is elusive. RESULTS: We investigated the correlation between crotonylation and HCC in 100 tumor tissues and performed stable isotope labeling by amino acids and liquid chromatography tandem mass spectrometry in HCC cells, and we found that crotonylation was positively correlated with HCC metastasis, and higher crotonylation in HCC cells facilitated cell invasiveness. Through bioinformatic analysis, we found that the crotonylated protein SEPT2 was significantly hypercrotonylated in highly invasive cells, while the decrotonylated mutation of SEPT2-K74 impaired SEPT2 GTPase activity and inhibited HCC metastasis in vitro and in vivo. Mechanistically, SIRT2 decrotonylated SEPT2, and P85α was found to be the downstream effector of SEPT2. Moreover, we identified that SEPT2-K74cr was correlated with poor prognosis and recurrence in HCC patients, thus indicating its clinical potential as an independent prognostic factor. CONCLUSIONS: We revealed the role of nonhistone protein crotonylation in regulating HCC metastasis and invasion. Crotonylation facilitated cell invasion through the crotonylated SEPT2-K74-P85α-AKT pathway. High SEPT2-K74 crotonylation predicted poor prognosis and a high recurrence rate in HCC patients. Our study revealed a novel role of crotonylation in promoting HCC metastasis.

Research Note: Polymorphisms of gonadotrophin-releasing hormone gene and their association with growth traits in quail (Coturnix Coturnix).

This study aimed to identify polymorphisms of gonadotropin-releasing hormone (GnRH) gene and their association with growth traits in quail by PCR and direct sequencing. Genomic DNA was extracted from quail blood samples of 36 from Savimalt (SV) and 49 from French Giant (FG). Growth traits were measured and used for candidate gene analysis, as body weight (BW), shank length (SL), chest width (CW), chest depth (CD), breastbone length (BBL), body length (BL), and shank circumference (SC). The results showed that a total of 20 SNPs were detected in GnRH gene, whereas 8 SNPs were significantly associated with growth traits (P < 0.05). The T215C, G279A, C458T, A520G, and C547G were significantly associated with SL at 3 wk of age in the FG strain, whereas A583T was significantly related to BBL and BL, and C591T was significantly related to SL, BBL, and BL, whereas A592G was significantly correlated with SL, CW, CD, BBL, and BL (P < 0.05). The 8 SNPs were significantly related to CW, CD, and BBL at 3 wk of age in the SV strain, whereas A583T, C591T, and A592G were significantly associated with BW (P < 0.05). The G279A showed significant correlations with SL at 5 wk of age in FG, whereas A583T showed significant associations with SC in FG, and C591T was significantly associated with BW and SC in FG, whereas A592T was significantly related to BW, SL, and CD in FG (P < 0.05). The T215C, G279A, C458T, A520G, and C547G were significantly correlated with BW, CW, BBL, and BL at 5 wk of age in SV, whereas A583T, C591T, and A592G were significantly related to BW, SL, CW, BBL, and BL (P < 0.05). Haplotypes based on 8 SNPs showed significant correlation with BW, SL, CW, CD, BBL, BL, and SC in FG (P < 0.05). In conclusion, the GnRH gene could be used as a molecular genetic marker to provide theoretical foundation to improve growth traits in quail.

Occurrence, spatial heterogeneity, and risk assessment of perfluoroalkyl acids (PFAAs) in the major rivers of the Tibetan Plateau.

The Tibetan Plateau (TP) is home to the headwaters of major rivers in Asia, yet their water quality security on a large spatial scale is scarcely studied, especially in regard to emerging organic pollutants. In this study, a systematic field campaign was carried out along Yarlung Tsangpo River, Nu River, Lancang River and Jinsha River, and 13 perfluoroalkyl acids (PFAAs) were analyzed. The total concentrations of PFAAs in the river waters of the TP were in the range of 0.58-7.46 ng/L, containing a high proportion of perfluorobutanoic acid (PFBA) and perfluorobutane sulfonate (PFBS) with average values of 56.7 %. Elevated PFAA loadings were found for the midstream of Yarlung Tsangpo River in central Tibet. Geodetector results indicated that precipitation, solar radiation and vegetation type were the top three influential factors contributing to the observed spatial heterogeneity. When interactions with human activities were taken into account, the explanatory power was significantly enhanced and rose above 0.70, highlighting the increased risks for TP rivers from the combined effects of natural environments and anthropogenic activities. Risk assessments suggest a low risk is posed to the alpine aquatic ecosystems and human health. The discharge fluxes of PFAAs via riverine export were estimated at 94-425 kg/year, which is one to two orders of magnitude lower than their mass loadings in major rivers worldwide. Our study underlined the need for further attention to the increased risk of water resource quality on the central TP in the context of long-range transport, increased cryosphere melting and local emission.

Microplastics in a Remote Lake Basin of the Tibetan Plateau: Impacts of Atmospheric Transport and Glacial Melting.

Plastic pollution is fast becoming one of the most pressing global issues that we currently face. Remote areas, such as the polar regions and the Tibetan Plateau, are now also exposed to microplastic contamination. However, with the impact of global warming, the transport of microplastics within the glacier-lake basins in such regions remains unclear. In this work, the Nam Co Basin in the Tibetan Plateau was selected to study the characteristics of microplastics in the rain fallout, lake water, glacial runoff, and non-glacial runoff. Fiber and films were the most common microplastic morphologies in all water samples; a higher proportion (37%) of light-weighing polypropylene and small-size (50-300 µm, ∼30%) microplastics were found in the glacial runoff. Air mass trajectory analysis showed that microplastics could be transported through the atmosphere over a distance of up to 800 km. For microplastic loading in lakes, the atmospheric fallout was estimated to be 3.3 tons during the monsoon season, whereas the contributions of glacial runoff (∼41 kg) and non-glacial runoff (∼522 kg) were relatively low. For the microplastic loading in glaciers, the atmospheric deposition was ∼500 kg/yr, and the output caused by glacial melting only accounted 8% of the total atmospheric input. All these results suggested that the dominant pathway through which microplastics enter remote mountainous lake basins is atmospheric deposition, and once deposited on glaciers, microplastics will be stored for a long time. This work provides quantitative evidence elucidating the fate of microplastics in alpine lake environments.

Perfluoroalkyl substances in precipitation from the Tibetan Plateau during monsoon season: Concentrations, source regions and mass fluxes.

Atmospheric wet deposition is an important process for the occurrence of perfluoroalkyl substances (PFASs) in polar/remote mountain regions; however, there are limited data on PFASs in precipitation from the Tibetan Plateau (TP). Precipitation (rain from May to October 2017) was therefore collected across the TP to investigate the concentrations, composition profiles, sources, and fluxes of perfluoroalkyl acids (PFAAs). The average ∑PFAA concentrations ranged from 212.3 pg L-1 to 547.7 pg L-1, and perfluoroalkyl carboxylic acids (PFCAs) accounted for 87% of the measured PFAAs (mean value). Significant positive associations (p < 0.05) were found for most PFCAs in the southeast TP, indicating that they may come from similar sources. The monthly PFAA deposition flux ranged from 12.6 to 68.9 ng m-2 month-1, decreasing from east to west. As climate of the eastern TP is controlled mainly by the Indian monsoon, indicating that the Indian monsoon plays an important role in delivering PFAAs to the TP. PCA (principal component analysis) combined with back-trajectory analysis was used to estimate the atmospheric transport pathways, and the PSCF (potential source contribution function) model was applied to define the potential source regions of individual PFAAs. The results suggested that northeast India, Bangladesh, and southern Nepal are the potential sources of C4-C7 PFCAs; C8-C10 PFCAs are more influenced by emissions from southern Nepal and Bhutan; while the source regions of long-chain PFCAs (C11-C12) can be attributed to northern India and Pakistan. Specifically, PFOS (perfluorooctane sulfonic acid) has a local contribution from the central TP.

The ETS Inhibitor YK-4-279 Suppresses Thyroid Cancer Progression Independent of TERT Promoter Mutations.

Hotspot mutations in the core promoter region of the telomerase reverse transcriptase (TERT) gene have been well established to associate with aggressive clinical characteristics, radioiodine refractory, tumor recurrence, and mortality in thyroid cancer. Several E-twenty-six (ETS) transcription factors were reported to selectively bound to the mutant TERT promoter and activated TERT expression. In this study we aimed to investigate whether TERT promoter mutations confer sensitivity to ETS inhibitor YK-4-279 in thyroid cancer cells and whether this inhibitor could be served as a potential therapeutic agent for thyroid cancer. In vitro assays showed that YK-4-279 treatment sharply suppressed cell viability, colony formation, migration, and invasion, as well as induced cell cycle arrest and apoptosis in a panel of thyroid cancer cells. The cell viability after YK-4-279 treatment was similar between cell lines harboring mutant and wild-type TERT promoters. Furthermore, YK-4-279 treatment reduced both luciferase activity and mRNA expression of TERT independent of TERT promoter mutation status. Data from RNA-seq further revealed that YK-4-279 significantly affected biological processes including DNA replication and cell cycle. Reduced DNA helicase activity and decreased expression of several helicase genes were observed after YK-4-279 treatment. Moreover, YK-4-279 significantly inhibited tumor growth and induced apoptosis in a xenograft mice model. Thus, ETS inhibitor YK-4-279 suppressed TERT expression and conferred anti-tumor activity in a TERT promoter mutation-independent manner, and it could be a potential agent for the treatment of advanced thyroid cancers.

RNF135 Promoter Methylation Is Associated With Immune Infiltration and Prognosis in Hepatocellular Carcinoma.

RING finger protein 135 has an important role in the occurrence of many cancers; however its regulation and function of RNF135 in hepatocellular carcinoma remains unknown. The promoter methylation status and mRNA expression of RNF135 was evaluated by methylation-specific PCR, semi-quantitative RT-PCR, and real-time quantitative PCR in HCC tissues and cell lines, and further analyzed from The Cancer Genome Atlas database. Wound healing assay, transwell migration, cell viability, and colony formation assay were performed to investigate the function of RNF135. GSEA analysis, TIMER database, and ESTIMATE algorithm were used to decipher the associated pathway and immune infiltration. The survival analysis was applied to assess the prognostic value of RNF135. RNF135 expression was downregulated in HCC tissues and 5 of 8 HCC cell lines, and was negatively correlated with its promoter hypermethylation. Demethylating regent decitabine restored RNF135 expression on the cellular level. Knockdown of RNF135 expression enhanced the migration of HCC cells, while RNF135 overexpression and decitabine treatment repressed cell migration. Bioinformatics analysis and immunohistochemistry revealed a positive relationship between RNF135 expression and six immune cell infiltrates (B cells, CD4+ T cells, CD8+ T cells, neutrophils, macrophages, and dendritic cells). Survival analysis disclosed that RNF135 hypermethylation is independently associated with poor clinical outcomes in HCC. Decreased RNF135 expression driven by promoter hypermethylation frequently occurred in HCC and associated with prognosis of HCC. RNF135 functions as a tumor suppressor and is involved in tumor immune microenvironment in HCC.