RESUMO
Background: Kandelia obovata is an important mangrove species extensively distributed in Eastern Asia that is susceptible to low-temperature stress. NAC (NAM, ATAF1/2 and CUC2) domain proteins are transcription factors (TFs) that play various roles in plant growth and development and in the plant response to environmental stresses. Nevertheless, genome-wide analyses of K. obovata NAC genes (KoNACs) and their responses to chilling stress have rarely been studied. Methods: The KoNAC gene family was identified and characterized using bioinformatic analysis, the subcellular location of some NAC proteins was confirmed using confocal microscopy analysis, and the KoNACs that responded to chilling stress were screened using RNA-seq and qRT-PCR analysis. Results: A total of 79 KoNACs were identified, and they were unequally distributed across all 18 chromosomes of K. obovata. The KoNAC proteins could be divided into 16 subgroups according to the phylogenetic tree based on NAC family members of Arabidopsis thaliana. The KoNACs exhibited greater synteny with A. thaliana sequences than with Oryza sativa sequences, indicating that KoNACs underwent extensive evolution after the divergence of dicotyledons and monocotyledons. Segmental duplication was the main driving force of the expansions of KoNAC genes. Confocal microscopy analysis verified that the four randomly selected KoNACs localized to the nucleus, indicating the accuracy of the bioinformatic predictions. Tissue expression pattern analysis demonstrated that some KoNAC genes showed tissue-specific expression, suggesting that these KoNACs might be important for plant development and growth. Additionally, the expression levels of 19 KoNACs were significantly (15 positively and 4 negatively) induced by cold treatment, demonstrating that these KoNACs might play important roles during cold stress responses and might be candidate genes for the genetic engineering of K. obovata with enhanced chilling stress tolerance. Coexpression network analysis revealed that 381 coexpressed pairs (between 13 KoNACs and 284 other genes) were significantly correlated. Conclusions: Seventy-nine KoNACs were identified in K. obovata, nineteen of which displayed chilling-induced expression patterns. These genes may serve as candidates for functional analyses of KoNACs engaged in chilling stress. Our results lay the foundation for evolutionary analyses of KoNACs and their molecular mechanisms in response to environmental stress.
RESUMO
Background: Kandelia obovata, a dominant mangrove species, is widely distributed in tropical and subtropical areas. Low temperature is the major abiotic stress that seriously limits the survival and growth of mangroves. WRKY transcription factors (TFs) play vital roles in responses to biotic and abiotic stresses. However, genome-wide analysis of WRKY genes in K. obovata and their responses to chilling stress have not been reported. Methods: Bioinformatic analysis was used to identify and characterize the K. obovata WRKY (KoWRKY) gene family, RNA-seq and qRT-PCR analyses were employed to screen KoWRKYs that respond to chilling stress. Results: Sixty-four KoWRKYs were identified and they were unevenly distributed across all 18 K. obovata chromosomes. Many orthologous WRKY gene pairs were identified between Arabidopsis thaliana and K. obovata, showing high synteny between the two genomes. Segmental duplication events were found to be the major force driving the expansion for the KoWRKY gene family. Most of the KoWRKY genes contained several kinds of hormone- and stress-responsive cis-elements in their promoter. KoWRKY proteins belonged to three groups (I, II, III) according to their conserved WRKY domains and zinc-finger structure. Expression patterns derived from the RNA-seq and qRT-PCR analyses revealed that 9 KoWRKYs were significantly upregulated during chilling acclimation in the leaves. KEGG pathway enrichment analysis showed that the target genes of KoWRKYs were significantly involved in 11 pathways, and coexpression network analysis showed that 315 coexpressed pairs (KoWRKYs and mRNAs) were positively correlated. Conclusion: Sixty-four KoWRKYs from the K. obovata genome were identified, 9 of which exhibited chilling stress-induced expression patterns. These genes represent candidates for future functional analysis of KoWRKYs involved in chilling stress related signaling pathways in K. obovata. Our results provide a basis for further analysis of KoWRKY genes to determine their functions and molecular mechanisms in K. obovata in response to chilling stress.