Structural variation discovery in wheat using PacBio high-fidelity sequencing.

Structural variations (SVs) pervade plant genomes and contribute substantially to the phenotypic diversity. However, most SVs were ineffectively assayed due to their complex nature and the limitations of early genomic technologies. By applying the PacBio high-fidelity (HiFi) sequencing for wheat genomes, we performed a comprehensive evaluation of mainstream long-read aligners and SV callers in SV detection. The results indicated that the accuracy of deletion discovery is markedly influenced by callers, accounting for 87.73% of the variance, whereas both aligners (38.25%) and callers (49.32%) contributed substantially to the accuracy variance for insertions. Among the aligners, Winnowmap2 and NGMLR excelled in detecting deletions and insertions, respectively. For SV callers, SVIM achieved the best performance. We demonstrated that combining the aligners and callers mentioned above is optimal for SV detection. Furthermore, we evaluated the effect of sequencing depth on the accuracy of SV detection, revealing that low-coverage HiFi sequencing is sufficiently robust for high-quality SV discovery. This study thoroughly evaluated SV discovery approaches and established optimal workflows for investigating structural variations using low-coverage HiFi sequencing in the wheat genome, which will advance SV discovery and decipher the biological functions of SVs in wheat and many other plants.

Genetic basis of genome size variation of wheat.

Research on various species has revealed a connection between genome size variation and the physiological and ecological characteristics of the species, suggesting that it could be a crucial factor influencing a species' adaptability to different environments. Wheat, being one of the world's three primary grains, holds significance in this regard. Investigating the genome size of wheat and analyzing the genetic factors contributing to its variation could offer valuable insights for enhancing wheat agronomic traits. This project has developed a conservative site ratio calculation approach to determine the size of the wheat genome. Additionally, it employs flow cytometry and k-mer distribution analysis to validate this method. Furthermore, the researchers use re-sequencing data to investigate the impact of environmental selection pressure and transposon dynamics on the variation in the size of the wheat genome. The findings from this study demonstrate a strong relationship between the size of the wheat genome and several environmental factors. These results serve as a valuable reference for understanding the development of variation in the size of the hetero-hexaploid wheat genome. Moreover, they contribute to advancing fundamental research on the genetic mechanisms underlying wheat characteristics. Additionally, the study paves the way for exploring new research directions in wheat breeding, which holds promise for future advancements in this field.

miR-182-5p attenuates Schistosoma japonicum-induced hepatic fibrosis by targeting tristetraprolin.

Egg granuloma formation in the liver is the main pathological lesion caused by Schistosoma japonicum infection, which generally results in liver fibrosis and may lead to death in advanced patients. MicroRNAs (miRNAs) regulate the process of liver fibrosis, but the putative function of miRNAs in liver fibrosis induced by S. japonicum infection is largely unclear. Here, we detect a new miRNA, miR-182-5p, which shows significantly decreased expression in mouse livers after stimulation by soluble egg antigen (SEA) of S. japonicum or S. japonicum infection. Knockdown or overexpression of miR-182-5p in vitro causes the increased or decreased expression of tristetraprolin (TTP), an important immunosuppressive protein in the process of liver fibrosis. Furthermore, knockdown of miR-182-5p in vivo upregulates TTP expression and significantly alleviates S. japonicum-induced hepatic fibrosis. Our data demonstrate that downregulation of miR-182-5p increases the expression of TTP in mouse livers following schistosome infection, which leads to destabilization of inflammatory factor mRNAs and attenuates liver fibrosis. Our results uncover fine-tuning of liver inflammatory reactions related to liver fibrosis caused by S. japonicum infection and provide new insights into the regulation of schistosomiasis-induced hepatic fibrosis.

Genetic and transcriptomic dissection of an artificially induced paired spikelets mutant of wheat (Triticum aestivum L.).

KEY MESSAGE: Morphological, genetic and transcriptomic characterizations of an EMS-induced wheat paired spikelets (PS) mutant were performed. A novel qualitative locus WPS1 on chromosome 1D was identified. Grain yield of wheat is significantly associated with inflorescence or spike architecture. However, few genes related to wheat spike development have been identified and their underlying mechanisms are largely unknown. In this study, we characterized an ethyl methanesulfonate (EMS)-induced wheat mutant, wheat paired spikelets 1 (wps1). Unlike a single spikelet that usually develops at each node of rachis, a secondary spikelet appeared below the primary spikelet at most of the rachis nodes of wps1. The microscope observation showed that the secondary spikelet initiated later than the primary spikelet. Genetic analysis suggested that the PS of wps1 is controlled by a single dominant nuclear gene, designated WHEAT PAIRED SPIKELETS 1 (WPS1). Further RNA-seq based bulked segregant analysis and molecular marker mapping localized WPS1 in an interval of 208.18-220.92 Mb on the chromosome arm 1DL, which is different to known genes related to spike development in wheat. By using wheat omics data, TraesCS1D02G155200 encoding a HD-ZIP III transcription factor was considered as a strong candidate gene for WPS1. Transcriptomic analysis indicated that PS formation in wps1 is associated with auxin-related pathways and may be regulated by networks involving TB1, Ppd1, FT1, VRN1, etc. This study laid the solid foundation for further validation of the causal gene of WPS1 and explored its regulatory mechanism in PS formation and inflorescence development, which may benefit to kernel yield improvement of wheat based on optimization or design of spike architecture in the future.

Comprehensive analyses of prognostic biomarkers and immune infiltrates among histone lysine demethylases (KDMs) in hepatocellular carcinoma.

BACKGROUND: Histone lysine demethylases (KDMs) are closely related to the occurrence and development of different tumors through epigenetic mechanisms. However, the prognosis and immune infiltration of KDMs in hepatocellular carcinoma (HCC) remain undefined. METHODS: In the current study, we analyzed the expression of KDMs on HCC patients using the Oncomine, GEPIA, UALCAN, Kaplan-Meier Plotter, cBioPortal, GeneMANIA, STRING, Metascape, GSEA, and TIMER databases. Finally, we investigated KDM expression in HCC by qRT-PCR, Western blotting, and IHC. RESULTS: We found that KDM3A/3B/5A/5B and KDM6A were upregulated in HCC patients, while KDM6B and KDM8 were downregulated. The high expressions of KDM1A/2B/3B/5B/5C were markedly related to tumor stages and grades of HCC patients. The abnormal expression of KDM1A/1B/3A/4A/5A/5C/6A/6B/7A and KDM8 were associated with HCC patients' prognosis. Also, we found that HCC tissues presented higher expression levels of KDM1A/2A/5A/5B and lower expression levels of KDM6B. The function of KDMs was primarily related to the histone demethylase activity and cell cycle, p53 signaling pathway, pathways in cancer, transcriptional mis-regulation in cancer, viral carcinogenesis, and FoxO signaling pathway. Furthermore, we indicated that the pathways most involved were the mitotic spindle and DNA repair. Additionally, we found that the expression of KDM1A/1B/3A/4A/5B/5C and KDM6A were significantly correlated with HCC immune infiltration. CONCLUSIONS: Overall, our current results indicated that KDM1A/1B/3A/4A/5B/5C and KDM6A could be novel prognostic biomarkers and provide insights into potential immunotherapy targets to HCC patients.

Puerarin promotes the osteogenic differentiation of rat dental follicle cells by promoting the activation of the nitric oxide pathway.

Puerarin regulates the osteoblast differentiation of umbilical cord mesenchymal stem cells. This study, hereby, explored the effects of puerarin on the osteogenic differentiation of dental follicle cells (DFCs) for the first time. Rat DFCs (rDFCs) were isolated and identified. After the rDFCs were treated by Puerarin and cultured in osteogenic induction medium, the viability, osteogenic differentiation, and the activities of alkaline phosphatase (ALP) and nitric oxide (NO) were detected. Besides, the secretion of cyclic guanosine monophosphate (cGMP) and expressions of collagen I, osteocalcin (OC), osteopontin (OPN), runt-related transcription factor 2 (RUNX2), soluble guanylate cyclase (SGC), and protein kinase G 1 (PKG-1) were further determined or quantified. Puerarin enhanced the viability and osteogenic differentiation, and increased the activities of ALP, NO, and cGMP and the expressions of Collagen I, OC, OPN, RUNX2, SGC, and PKG-1 in rDFCs. After the co-treatment with puerarin and L-NMMA (NO synthase inhibitor), the promotive effects of Puerarin on cell viability, osteogenic differentiation, and the expressions of collagen I, OC, OPN, RUNX2, SGC, and PKG-1 in rDFCs were reversed by L-NMMA. Puerarin boosted the osteogenic differentiation of rDFCs by activating the NO pathway.

Genetic and molecular characterization of determinant of six-rowed spike of barley carrying vrs1.a4.

KEY MESSAGE: Decisive role of reduced vrs1 transcript abundance in six-rowed spike of barley carrying vrs1.a4 was genetically proved and its potential causes were preliminarily analyzed. Six-rowed spike 1 (vrs1) is the major determinant of the six-rowed spike phenotype of barley (Hordeum vulgare L.). Alleles of Vrs1 have been extensively investigated. Allele vrs1.a4 in six-rowed barley is unique in that it has the same coding sequence as Vrs1.b4 in two-rowed barley. The determinant of row-type in vrs1.a4 carriers has not been experimentally identified. Here, we identified Vrs1.b4 in two-rowed accessions and vrs1.a4 in six-rowed accessions from the Qinghai-Tibet Plateau at high frequency. Genetic analyses revealed a single nuclear gene accounting for row-type alteration in these accessions. Physical mapping identified a 0.08-cM (~ 554-kb) target interval on chromosome 2H, wherein Vrs1 was the most likely candidate gene. Further analysis of Vrs1 expression in offspring of the mapping populations or different Vrs1.b4 and vrs1.a4 lines confirmed that downregulated expression of vrs1.a4 causes six-rowed spike. Regulatory sequence analysis found a single 'TA' dinucleotide deletion in vrs1.a4 carriers within a 'TA' tandem-repeat-enriched region ~ 1 kb upstream of the coding region. DNA methylation levels did not correspond to the expression difference and therefore did not affect Vrs1 expression. More evidence is needed to verify the causal link between the 'TA' deletion and the downregulated Vrs1 expression and hence the six-rowed spike phenotype.

Identification and Validation of a Novel Locus Controlling Spikelet Number in Bread Wheat (Triticum aestivum L.).

Spikelet number is an important target trait for wheat yield improvement. Thus, the identification and verification of novel quantitative trait locus (QTL)/genes controlling spikelet number are essential for dissecting the underlying molecular mechanisms and hence for improving grain yield. In the present study, we constructed a high-density genetic map for the Kechengmai1/Chuanmai42 doubled haploid (DH) population using 13,068 single-nucleotide polymorphism (SNP) markers from the Wheat 55K SNP array. A comparison between the genetic and physical maps indicated high consistence of the marker orders. Based on this genetic map, a total of 27 QTLs associated with total spikelet number per spike (TSN) and fertile spikelet number per spike (FSN) were detected on chromosomes 1B, 1D, 2B, 2D, 3D, 4A, 4D, 5A, 5B, 5D, 6A, 6B, and 7D in five environments. Among them, five QTLs on chromosome 2D, 3D, 5A, and 7D were detected in multiple environments and combined QTL analysis, explaining the phenotypic variance ranging from 3.64% to 23.28%. Particularly, QTsn/Fsn.cib-3D for TSN and FSN [phenotypic variation explained (PVE) = 5.97-23.28%, limit of detection (LOD) = 3.73-18.51] is probably a novel locus and located in a 4.5-cM interval on chromosome arm 3DL flanking by the markers AX-110914105 and AX-109429351. This QTL was further validated in other two populations with different genetic backgrounds using the closely linked Kompetitive Allele-Specific PCR (KASP) marker KASP_AX-110914105. The results indicated that QTsn/Fsn.cib-3D significantly increased the TSN (5.56-7.96%) and FSN (5.13-9.35%), which were significantly correlated with grain number per spike (GNS). We also preliminary analyzed the candidate genes within this locus by sequence similarity, spatial expression patterns, and collinearity analysis. These results provide solid foundation for future fine mapping and cloning of QTsn/Fsn.cib-3D. The developed and validated KASP markers could be utilized in molecular breeding aiming to increase the grain yield in wheat.