Integrated Single-cell and Bulk RNA Sequencing Analysis Cross Talk between Ferroptosis-related Genes and Prognosis in Oral Cavity Squamous Cell Carcinoma.

BACKGROUND: Ferroptosis is a new type of programmed apoptosis and plays an important role in tumour inhibition and immunotherapy. OBJECTIVE: In this study, we aimed to explore the potential role of ferroptosis-related genes (FRGs) and the potential therapeutic targets in oral cavity squamous cell carcinoma (OCSCC). METHODS: The transcription data of OCSCC samples were obtained from the Cancer Genome Atlas (TCGA) database as a training dataset. The prognostic FRGs were extracted by univariate Cox regression analysis. Then, we constructed a prognostic model using the least absolute shrinkage and selection operator (LASSO) and Cox analysis to determine the independent prognosis FRGs. Based on this model, risk scores were calculated for the OCSCC samples. The model's capability was further evaluated by the receiver operating characteristic curve (ROC). Then, we used the GSE41613 dataset as an external validation cohort to confirm the model's predictive capability. Next, the immune infiltration and somatic mutation analysis were applied. Lastly, single-cell transcriptomic analysis was used to identify the key cells. RESULTS: A total of 12 prognostic FRGs were identified. Eventually, 6 FRGs were screened as independent predictors and a prognostic model was constructed in the training dataset, which significantly stratified OCSCC samples into high-risk and low-risk groups based on overall survival. The external validation of the model using the GSE41613 dataset demonstrated a satisfactory predictive capability for the prognosis of OCSCC. Further analysis revealed that patients in the highrisk group had distinct immune infiltration and somatic mutation patterns from low-risk patients. Mast cell infiltrations were identified as prognostic immune cells and played a role in OCSCC partly through ferroptosis. CONCLUSION: We successfully constructed a novel 6 FRGs model and identified a prognostic immune cell, which can serve to predict clinical prognoses for OCSCC. Ferroptosis may be a new direction for immunotherapy of OCSCC.

Genomic insights into the genetic signatures of selection and seed trait loci in cultivated peanut.

INTRODUCTION: Cultivated peanut (Arachis hypogaea L.) is an important oil crop for human nutrition and is cultivated in >100 countries. However, the present knowledge of its genomic diversity, evolution, and loci related to the seed traits is limited. OBJECTIVES: Our study intended to (1) uncover the population structure and the demographic history of peanuts, (2) identify signatures of selection that occurred during peanut improvement breeding, and (3) detect and verify the functions of candidate genes associated with seed traits. METHODS: We explored the population relationship and the evolution of peanuts using a largescale single nucleotide polymorphism dataset generated from the genome-wide resequencing of 203 cultivated peanuts. Genetic diversity and genomic scan analyses were applied to identify selective loci for genomic-selection breeding. Genome-wide association studies, transgenic experiments, and RNA-seq were employed to identify the candidate genes associated with seed traits. RESULTS: Our study revealed that the 203 resequenced accessions were divided into four genetic groups, consistent with their botanical classification. Moreover, the var. peruviana and var. fastigiata subpopulations have diverged to a greater extent than the others, and var. peruviana may be the earliest variant in the evolution from tetraploid ancestors. A recent dramatic expansion in the effective population size of the cultivated peanuts ca. 300-500 years ago was also noted. Selective sweeps underlying quantitative trait loci and genes of seed size, plant architecture, and disease resistance coincide with the major goals of improved peanut breeding compared with the landrace and cultivar populations. Genome-wide association testing with functional analysis led to the identification of two genes involved in seed weight and seed length regulation. CONCLUSION: Our study provides valuable information for understanding the genomic diversity and the evolution of peanuts and serves as a genomic basis for improving peanut cultivars.

Bicolor angelfish (Centropyge bicolor) provides the first chromosome-level genome of the Pomacanthidae family.

The Bicolor Angelfish, Centropyge bicolor, is a tropical coral reef fish. It is named for its striking two-color body. However, a lack of high-quality genomic data means little is known about the genome of this species. Here, we present a chromosome-level C. bicolor genome constructed using Hi-C data. The assembled genome is 650 Mbp in size, with a scaffold N50 value of 4.4 Mbp, and a contig N50 value of 114 Kbp. Protein-coding genes numbering 21,774 were annotated. Our analysis will help others to choose the most appropriate de novo genome sequencing strategy based on resources and target applications. To the best of our knowledge, this is the first chromosome-level genome for the Pomacanthidae family, which might contribute to further studies exploring coral reef fish evolution, diversity and conservation.

The Chromosome Level Genome and Genome-wide Association Study for the Agronomic Traits of Panax Notoginseng.

The Chinese ginseng Panax notoginseng is a domesticated herb with significant medicinal and economic value. Here we report a chromosome-level P. notoginseng genome assembly with a high (∼79%) repetitive sequence content. The juxtaposition with the widely distributed, closely related Korean ginseng (Panax ginseng) genome revealed contraction of plant defense genes (in particular R-genes) in the P. notoginseng genome. We also investigated the reasons for the larger genome size of Panax species, revealing contributions from two Panax-specific whole-genome duplication events and transposable element expansion. Transcriptome data and comparative genome analysis revealed the candidate genes involved in the ginsenoside synthesis pathway. We also performed a genome-wide association study on 240 cultivated P. notoginseng individuals and identified the associated genes with dry root weight (63 genes) and stem thickness (168 genes). The P. notoginseng genome represents a critical step toward harnessing the full potential of an economically important and enigmatic plant.

An integrated Asian human SNV and indel benchmark established using multiple sequencing methods.

Sequencing technologies have been rapidly developed recently, leading to the breakthrough of sequencing-based clinical diagnosis, but accurate and complete genome variation benchmark would be required for further assessment of precision medicine applications. Despite the human cell line of NA12878 has been successfully developed to be a variation benchmark, population-specific variation benchmark is still lacking. Here, we established an Asian human variation benchmark by constructing and sequencing a stabilized cell line of a Chinese Han volunteer. By using seven different sequencing strategies, we obtained ~3.88 Tb clean data from different laboratories, hoping to reach the point of high sequencing depth and accurate variation detection. Through the combination of variations identified from different sequencing strategies and different analysis pipelines, we identified 3.35 million SNVs and 348.65 thousand indels, which were well supported by our sequencing data and passed our strict quality control, thus should be high confidence variation benchmark. Besides, we also detected 5,913 high-quality SNVs which had 969 sites were novel and  located in the high homologous regions supported by long-range information in both the co-barcoding single tube Long Fragment Read (stLFR) data and PacBio HiFi CCS data. Furthermore, by using the long reads data (stLFR and HiFi CCS), we were able to phase more than 99% heterozygous SNVs, which helps to improve the benchmark to be haplotype level. Our study provided comprehensive sequencing data as well as the integrated variation benchmark of an Asian derived cell line, which would be valuable for future sequencing-based clinical development.

GAA compound heterozygous mutations associated with autophagic impairment cause cerebral infarction in Pompe disease.

Clinical manifestations of the late-onset adult Pompe disease (glycogen storage disease type II) are heterogeneous. To identify genetic defects of a special patient population with cerebrovascular involvement as the main symptom, we performed whole-genome sequencing (WGS) analysis on a consanguineous Chinese family of total eight members including two Pompe siblings both had cerebral infarction. Two novel compound heterozygous variants were found in GAA gene: c.2238G>C in exon 16 and c.1388_1406del19 in exon 9 in the two patients. We verified the function of the two mutations in leading to defects in GAA protein expression and enzyme activity that are associated with autophagic impairment. We further performed a gut microbiome metagenomics analysis, found that the child's gut microbiome metagenome is very similar to his mother. Our finding enriches the gene mutation spectrum of Pompe disease, and identified the association of the two new mutations with autophagy impairment. Our data also indicates that gut microbiome could be shared within Pompe patient and cohabiting family members, and the abnormal microbiome may affect the blood biochemical index. Our study also highlights the importance of deep DNA sequencing in potential clinical applications.

Chromosome-level reference genome of the Siamese fighting fish Betta splendens, a model species for the study of aggression.

Background: Siamese fighting fish Betta splendens are notorious for their aggressiveness and accordingly have been widely used to study aggression. However, the lack of a reference genome has, to date, limited the understanding of the genetic basis of aggression in this species. Here, we present the first reference genome assembly of the Siamese fighting fish. Findings: Frist, we sequenced and de novo assembled a 465.24-Mb genome for the B. splendens variety Giant, with a weighted average (N50) scaffold size of 949.03 Kb and an N50 contig size of 19.01 Kb, covering 99.93% of the estimated genome size. To obtain a chromosome-level genome assembly, we constructed one Hi-C library and sequenced 75.24 Gb reads using the BGISEQ-500 platform. We anchored approximately 93% of the scaffold sequences into 21 chromosomes and evaluated the quality of our assembly using the high-contact frequency heat map and Benchmarking Universal Single-Copy Orthologs. We also performed comparative chromosome analyses between Oryzias latipes and B. splendens, revealing a chromosome conservation evolution in B. splendens. We predicted 23,981 genes assisted by RNA-sequencing data generated from brain, liver, muscle, and heart tissues of Giant and annotated 15% repetitive sequences in the genome. Additionally, we resequenced five other B. splendens varieties and detected ∼3.4 M single-nucleotide variations and 27,305 insertions and deletions. Conclusions: We provide the first chromosome-level genome for the Siamese fighting fish. The genome will lay a valuable foundation for future research on aggression in B. splendens.

Frequent alterations in cytoskeleton remodelling genes in primary and metastatic lung adenocarcinomas.

The landscape of genetic alterations in lung adenocarcinoma derived from Asian patients is largely uncharacterized. Here we present an integrated genomic and transcriptomic analysis of 335 primary lung adenocarcinomas and 35 corresponding lymph node metastases from Chinese patients. Altogether 13 significantly mutated genes are identified, including the most commonly mutated gene TP53 and novel mutation targets such as RHPN2, GLI3 and MRC2. TP53 mutations are furthermore significantly enriched in tumours from patients harbouring metastases. Genes regulating cytoskeleton remodelling processes are also frequently altered, especially in metastatic samples, of which the high expression level of IQGAP3 is identified as a marker for poor prognosis. Our study represents the first large-scale sequencing effort on lung adenocarcinoma in Asian patients and provides a comprehensive mutational landscape for both primary and metastatic tumours. This may thus form a basis for personalized medical care and shed light on the molecular pathogenesis of metastatic lung adenocarcinoma.

Detection of inherited mutations for hereditary cancer using target enrichment and next generation sequencing.

Hereditary cancers occur because of inherited gene mutations. Genetic testing has been approved to provide information for risk assessment and rationale for appropriate intervention. Testing methods currently available for clinical use have some limitations, including sensitivity and testing throughput, etc. Next generation sequencing (NGS) has been rapidly evolving to increase testing sensitivity and throughput. It can be potentially used to identify inherited mutation in clinical diagnostic setting. Here we develop an effective method employing target enrichment and NGS platform to detect common as well as rare mutations for all common hereditary cancers in a single assay. Single base substitution across 115 hereditary cancer related genes using YH (the first Asian genome) was characterized to validate our method. Sensitivity, specificity and accuracy of 93.66, 99.98 and 99.97 %, were achieved, respectively. In addition, we correctly identified 53 SNVs and indels of BRCA1 and BRCA2 in two breast cancer specimens, all confirmed by Sanger sequencing. Accuracy in detecting copy number variation (CNV) was corroborated in 4 breast cancer specimens with known CNVs in BRAC1. Application of the method to 85 clinical cases revealed 22 deleterious mutations, 11 of which were novel. In summary, our studies demonstrate that the target enrichment combined with NGS method provides the accuracy, sensitivity, and high throughput for genetic testing for patients with high risk of hereditary or familial cancer.