Identification of long noncoding RNAs involved in resistance to downy mildew in Chinese cabbage

Brassica downy mildew,a severe disease caused by Hyaloperonospora brassicae,can cause enormous economic losses in Chinese cabbage(Brassica rapa L.ssp.pekinensis)production.Although some research has been reported recently concerning the underlying resistance to this disease,no studies have identified or characterized long noncoding RNAs involved in this defense response.In this study,using high-throughput RNA sequencing,we analyzed the disease-responding mRNAs and long noncoding RNAs in two resistant lines(T12–19 and 12–85)and one susceptible line(91–112).Clustering and Gene Ontology analysis of differentially expressed genes(DEGs)showed that more DEGs were involved in the defense response in the two resistant lines than in the susceptible line.Different expression patterns and proposed functions of differentially expressed long noncoding RNAs among T12–19,12–85,and 91–112 indicated that each has a distinct disease response mechanism.There were significantly more cis-and trans-functional long noncoding RNAs in the resistant lines than in the susceptible line,and the genes regulated by these RNAs mostly participated in the disease defense response.Furthermore,we identified a candidate resistance-related long noncoding RNA,MSTRG.19915,which is a long noncoding natural antisense transcript of a MAPK gene,BrMAPK15.Via an agroinfiltration-mediated transient overexpression system and virus-induced gene silencing technology,BrMAPK15 was indicated to have a greater ability to defend against pathogens.MSTRG.19915-silenced seedlings showed enhanced resistance to downy mildew,probably because of the upregulated expression of BrMAPK15.This research identified and characterized long noncoding RNAs involved in resistance to downy mildew,laying a foundation for future in-depth studies of disease resistance mechanisms in Chinese cabbage.

Genome-wide analysis of changes in miRNA and target gene expression reveals key roles in heterosis for chinese cabbage biomass

Heterosis is a complex phenomenon in which hybrids show better phenotypic characteristics than their parents do.Chinese cabbage(Brassica rapa L.spp.pekinensis)is a popular leafy crop species,hybrids of which are widely used in commercial production;however,the molecular basis of heterosis for biomass of Chinese cabbage is poorly understood.We characterized heterosis in a Chinese cabbage hybrid cultivar and its parental lines from the seedling stage to the heading stage;marked heterosis of leaf weight and biomass yield were observed.Small RNA sequencing revealed 63 and 50 differentially expressed microRNAs(DEMs)at the seedling and early-heading stages,respectively.The expression levels ofthe majority of miRNA clusters in the hybrid were lower than the mid-parent values(MPVs).Using degradome sequencing,we identi fied 1,819 miRNA target genes.Gene ontology(GO)analyses demonstrated that the target genes ofthe MPV-DEMs and low parental expression level dominance(ELD)miRNAs were signi ficantly enriched in leaf morphogenesis,leaf development,and leaf shaping.Transcriptome analysis revealed that the expression levels of photosynthesis and chlorophyll synthesis-related MPV-DEGs(differentially expressed genes)were signi ficantly different in the F_(1) hybrid compared to the parental lines,resulting in increased photosynthesis capacity and chlorophyll content in the former.Furthermore,expression of genes known to regulate leaf development was also observed at the seedling stage.Arabidopsis plants overexpressing BrGRF4.2 and bra-miR396 presented increased and decreased leaf sizes,respectively.These results provide new insight into the regulation of target genes and miRNA expression patterns in leaf size and heterosis for biomass of B.rapa.

Editing of eIF(iso)4E.c confers resistance against Turnip mosaic virus in Brassica rapa

Turnip mosaic virus(TuMV)constitutes one of the primary diseases affecting Brassica rapa,severely impacting its production and resulting in crop failures in various regions worldwide.Recent research has demonstrated the significance of plant translation initiation factors,specifically the eIF4E and eIF4G family genes,as essential recessive disease resistance genes.In our study,we conducted evolutionary and gene expression studies,leading us to identify e IF(iso)4E.c as a potential TuMV-resistant gene.Leveraging CRISPR/Cas9 technology,we obtained mutant B.rapa plants with edited eIF(iso)4E.c gene.We confirmed eIF(iso)4E.c confers resistance against TuMV through phenotypic observations and virus content evaluations.Furthermore,we employed ribosome profiling assays on eif(iso)4e.c mutant seedlings to unravel the translation landscape in response to TuMV.Interestingly,we observed a moderate correlation between the fold changes in gene expression at the transcriptional and translational levels(R^(2)=0.729).Comparative analysis of ribosome profiling and RNA-seq data revealed that plant-pathogen interaction,and MAPK signaling pathway-plant pathways were involved in eIF(iso)4E.c-mediated TuMV resistance.Further analysis revealed that sequence features,coding sequence length,and normalized minimal free energy,influenced the translation efficiency of genes.Our study highlights that the loss of e IF(iso)4E.c can result in a highly intricate translation mechanism,acting synergistically with transcription to confer resistance against TuMV.

A Genomic Variation Map Provides Insights into the Genetic Basis of Spring Chinese Cabbage (Brassica rapa ssp.pekinensis)Selection

Chinese cabbage is the most consumed leafy crop in East Asian countries.However,premature bolting induced by continuous low temperatures severely decreases the yield and quality of the Chinese cabbage, and therefore restricts its planting season and geographic distribution.In the past 40years,spring Chinese cabbage with strong winterness has been selected to meet the market demand.Here,we report a genome variation map of Chinese cabbage generated from the resequencing data of 194 geographically diverse accessions of three ecotypes.In-depth analyses of the selection sweeps and genome-wide patterns revealed that spring Chinese cabbage was selected from a specific population of autumn Chinese cabbage around the area of Shandong peninsula in northern China.We identified 23 genomic loci that underwent intensive selection,and further demonstrated by gene expression and haplotype analyses that the incorporation of elite alleles of VERNALISATION INSENTIVE 3.1(BrVIN3.1)and FLOWER LOCUS C 1(BrFLC1)is a determinant genetic source of variation during selection.Moreover,we showed that the quantitative response of BrVIN3.1 to cold due to the sequence variations in the cis elements of the BrVlN3.1 promoter significantly contributes to bolting-time variation in Chinese cabbage.Collectively, our study provides valuable insights into the genetic basis of spring Chinese cabbage selection and will facilitate the breeding of bolting-resistant Varieties by molecular-marker-assisted selection,transgenic or gene editingapproaches.