An elucidation of how the selection pressures caused by habitat environments affect plant plastid genomes and lead to the adaptive evolution of plants, is a very intense area of research in evolutionary biology. The genus Paphiopedilum is a predominant group of orchids that includes over 66 species with high horticultural and ornamental value. However, owing to the destructive exploitation and habitat deterioration of wild germplasm resources of Paphiopedilum, it needs more molecular genetic resources and studies on this genus. The chloroplast is cytoplasmically inherited and often used in evolutionary studies. Thus, for this study, we newly sequenced, assembled and annotated five chloroplast genomes of the Paphiopedilum species. The size of these genomes ranged from 155,886 bp (P. henryanum) to 160,503 bp (P. ‘GZSLKY’ Youyou) and they contained 121–122 genes, which consisted of 76 protein coding genes, eight ribosomal RNAs, and 37–38 transfer RNAs. Combined with the other 14 Paphiopedilum species, the characteristics of the repeat sequences, divergent hotspot regions, and the condo usage bias were evaluated and identified, respectively. The gene transfer analysis showed that some fragments of the ndh and ycf gene families were shared by both the chloroplast and nucleus. Although the genomic structure and gene content was conserved, there was a significant boundary shift caused by the inverted repeat (IR) expansion and small single copy (SSC) contraction. The lower GC content and loss of ndh genes could be the result of adaptive evolutionary responses to its unique habitats. The genes under positive selection, including accD, matK, psbM, rpl20, rps12, ycf1, and ycf2 might be regarded as potential candidate genes for further study, which significantly contribute to the adaptive evolution of Paphiopedilum.

Wall-associated kinase (WAK) and WAK-like kinase (WAKL) are receptor-like kinases (RLKs), which play important roles in signal transduction between the cell wall and the cytoplasm in plants. WAK/WAKLs have been studied in many plants, but were rarely studied in the important economic walnut tree. In this study, 27 and 14 WAK/WAKL genes were identified in Juglans regia and its wild related species Juglans mandshurica, respectively. We found tandem duplication might play a critical role in the expansion of WAK/WAKL gene family in J. regia, and most of the WAK/WAKL homologous pairs underwent purified selection during evolution. All WAK/WAKL proteins have the extracellular WAK domain and the cytoplasmic protein kinase domain, and the latter was more conserved than the former. Cis-acting elements analysis showed that WAK/WAKL might be involved in plant growth and development, plant response to abiotic stress and hormones. Gene expression pattern analysis further indicated that most WAK/WAKL genes in J. regia might play a role in the development of leaves and be involved in plant response to biotic stress. Our study provides a new perspective for the evolutionary analysis of gene families in tree species and also provides potential candidate genes for studying WAK/WAKL gene function in walnuts.

Cold stress impacts woody tree growth and perennial production, especially when the temperature rapidly changes in late spring. To address this issue, we conducted the genome-wide identification of two important transcription factors (TFs), CBF (C-repeat binding factors) and ICE (inducers of CBF expression), in three walnut (Juglans) genomes. Although the CBF and ICE gene families have been identified in many crops, very little systematic analysis of these genes has been carried out in J. regia and J. sigillata. In this study, we identified a total of 16 CBF and 12 ICE genes in three Juglans genomes using bioinformatics analysis. Both CBF and ICE had conserved domains, motifs, and gene structures, which suggests that these two TFs were evolutionarily conserved. Most ICE genes are located at both ends of the chromosomes. The promoter cis-regulatory elements of CBF and ICE genes are largely involved in light and phytohormone responses. Based on 36 RNA sequencing of leaves from four walnut cultivars (‘Zijing’, ‘Lvling’, ‘Hongren’, and ‘Liao1’) under three temperature conditions (8 °C, 22 °C, and 5 °C) conditions in late spring, we found that the ICE genes were expressed more highly than CBFs. Both CBF and ICE proteins interacted with cold-related proteins, and many putative miRNAs had interactions with these two TFs. These results determined that CBF1 and ICE1 play important roles in the tolerance of walnut leaves to rapid temperature changes. Our results provide a useful resource on the function of the CBF and ICE genes related to cold tolerance in walnuts.

The YABBY gene family is a plant transcription factor that exists in all seed plants. YABBY family members have been studied extensively in various plants and were to play significant roles in plant growth and development. Juglans, especially walnuts, are important economic tree species that are widely distributed worldwide. However, the identification and related research of YABBY in Juglans have not been reported to date. In this study, we identified 19 YABBY genes from two Juglans species, namely, J. regia and J. mandshurica. Ten JrYABBY genes and nine JmYABBY genes were divided into five subfamilies (YAB1/3, YAB2, INO, CRC, and YAB5). Sequence analysis revealed that all encoded YABBY protein sequences had a highly conserved YABBY and C2C2 zinc-finger domains. An analysis of the assumed cis-acting elements revealed that JrYABBY and JmYABBY genes were deeply involved in phytohormone and light responses. Further, gene expression pattern analysis suggested that most walnut YABBY genes were likely involved in peel and flower development and responses to biotic stress. This study not only suppled novel insights into the evolutionary basis of YABBY gene families in Juglans, but also provided clues for the further functional verification and investigation of YABBY genes in other tree species.

Plant NAPDH oxidase (NOX) gene family members are critical producers of ROS and play crucial roles in plant signaling, development, and stress responses. Opisthopappus taihangensis is a pivotal wild genetic resource in Asteraceae. To date, little knowledge exists about the functions of NOXs in O. taihangensis. In this study, seven typical NOXs and seven ferric reduction oxidases (FROs) were identified in O. taihangensis. Dispersed duplication might play a critical role in the expansion of the NOX/FRO gene family in O. taihangensis, and most of the NOX/FRO homologous pairs have undergone purifying selection. Although the results of the collinearity analysis show that these genes were relatively conserved, the gain and loss of members in this gene family occurred frequently during the evolution of Asteraceae. Phylogenetic analysis indicated that the FROs are relatively ancient, and the earliest diverged branch was revealed by the maximum likelihood (ML) tree. The Asteraceae NOXs were divided into six subgroups according to their relationship with Arabidopsis and rice members. The transcriptome profile unveiled tissue-specific expression patterns and complex response modes under drought and salt stresses. OtNOX6 and OtNOX7 could be recognized as important candidates to respond to drought and salt stress. Our results provide clues and references for further in-depth exploration of NOX/FRO function in O. taihangensis and other horticulture plants.

Disentangling how climate oscillations and geographical events significantly influence plants’ genetic architecture and demographic history is a central topic in phytogeography. The deciduous ancient tree species Ulmus macrocarpa is primarily distributed throughout Northern China and has timber and horticultural value. In the current study, we studied the phylogenic architecture and demographical history of U. macrocarpa using chloroplast DNA with ecological niche modeling. The results indicated that the populations’ genetic differentiation coefficient (N_ST) value was significantly greater than the haplotype frequency (G_ST) (p < 0.05), suggesting that U. macrocarpa had a clear phylogeographical structure. Phylogenetic inference showed that the putative chloroplast haplotypes could be divided into three groups, in which the group Ⅰ was considered to be ancestral. Despite significant genetic differentiation among these groups, gene flow was detected. The common ancestor of all haplotypes was inferred to originate in the middle–late Miocene, followed by the haplotype overwhelming diversification that occurred in the Quaternary. Combined with demography pattern and ecological niche modeling, we speculated that the surrounding areas of Shanxi and Inner Mongolia were potential refugia for U. macrocarpa during the glacial period in Northern China. Our results illuminated the demography pattern of U. macrocarpa and provided clues and references for further population genetics investigations of precious tree species distributed in Northern China.