Background: The C2H2 zinc finger protein family plays important roles in plants. However, precisely how C2H2s function in Opisthopappus (Opisthopappus taihangensis and Opisthopappus longilobus) remains unclear. Results: In this study, a total of 69 OpC2H2 zinc finger protein genes were identified and clustered into five Groups. Seven tandem and ten fragment repeats were found in OpC2H2s, which underwent robust purifying selection. Of the identified motifs, motif 1 was present in all OpC2H2s and conserved at important binding sites. Most OpC2H2s possessed few introns and exons that could rapidly activate and react when faced with stress. The OpC2H2 promoter sequences mainly contained diverse regulatory elements, such as ARE, ABRE, and LTR. Under salt stress, two up-regulated OpC2H2s (OpC2H2-1 and OpC2H2-14) genes and one down-regulated OpC2H2 gene (OpC2H2-7) might serve as key transcription factors through the ABA and JA signaling pathways to regulate the growth and development of Opisthopappus species. Conclusion: The above results not only help to understand the function of C2H2 gene family but also drive progress in genetic improvement for the salt tolerance of Opisthopappus species.

Salt stress profoundly affects plant growth, prompting intricate molecular responses, such as alternative splicing (AS), for environmental adaptation. However, the response of AS events to salt stress in Opisthopappus (Opisthopappus taihangensis and Opisthopappus longilobus) remains unclear, which is a Taihang Mountain cliff-dwelling species. Using RNA-seq data, differentially expressed genes (DEGs) were identified under time and concentration gradients of salt stress. Two types of AS, skipped exon (SE) and mutually exclusive exons (MXE), were found. Differentially alternative splicing (DAS) genes in both species were significantly enriched in “protein phosphorylation”, “starch and sucrose metabolism”, and “plant hormone signal transduction” pathways. Meanwhile, distinct GO terms and KEGG pathways of DAS occurred between two species. Only a small subset of DAS genes overlapped with DEGs under salt stress. Although both species likely adopted protein phosphorylation to enhance salt stress tolerance, they exhibited distinct responses. The results indicated that the salt stress mechanisms of both Opisthopappus species exhibited similarities and differences in response to salt stress, which suggested that adaptive divergence might have occurred between them. This study initially provides a comprehensive description of salt responsive AS events in Opisthopappus and conveys some insights into the molecular mechanisms behind species tolerance on the Taihang Mountains.

Opisthopappus is a perennial, endemic herb of the Taihang Mountains in China. Two species of this genus (O. longilobus and O. taihangensis) are important wild genetic resources for Asteraceae; however, their reproductive biology has been lacking until now. This study is the first detailed report on the reproductive biology and breeding systems of two Opisthopappus species. Through field observations, the floral syndromes of O. longilobus and O. taihangensis were found to possess a similar pattern, although O. taihangensis has a relatively larger capitulum, more ray ligules, and disc florets. The flowers of both O. longilobus and O. taihangensis are protandrous, a character that can prevent autogamy at the single-flower level, and insects are required for pollination. Further, brightly ligules, brightly bisexual florets, unique fragrance, and amount of nectar suggest that these species propagate via an entomophilous pollination system. Hymenopteran and Diptera species were observed as the effective pollinators for these two species. The outcrossing index, pollen/ovule ratio and the results of hand pollination indicated that these Opisthopappus species might have a mixed mating system that combines cross-fertilization and partial self-fertilization for O. longilobus and O. taihangensis, outcrossing predominated in the breeding system, while self-pollination played an important role in seed production when insect pollination was unavailable, particularly in a harsh environment, such as the Taihang Mountains cliffs. Meanwhile, O. taihangensis might better adapt to severe surroundings with relatively complex floral syndromes, specifically through the attraction of visiting insects and a high seed set rate. The above results not only provide reference information toward a better understanding of the survival strategies of O. longilobus and O. taihangensis in the Taihang Mountains but also lay a solid foundation for further exploring the molecular mechanisms that underly their adaptation under cliff environments.

When plants are exposed to salt stress, endogenous hormones are essential for their responses through biosynthesis and signal transduction pathways. However, the roles of endogenous hormones in two cliff species (Opisthopappus taihangensis and Opisthopappus longilobus (Opisthopappus genus)) in the Taihang Mountains under salt stress have not been investigated to date. Following different time treatments under 500 mM salt concentrations, 239 differentially expressed gene (DEG)-related endogenous hormones were identified that exhibited four change trends, which in Profile 47 were upregulated in both species. The C-DEG genes of AUX, GA, JA, BR, ETH, and ABA endogenous hormones were significantly enriched in Opisthopappus taihangensis (O. taihangensis) and Opisthopappus longilobus (O. longilobus). During the responsive process, mainly AUX, GA, and JA biosynthesis and signal transduction were triggered in the two species. Subsequently, crosstalk further influenced BR, EHT, ABA, and MAPK signal transduction pathways to improve the salt resistance of the two species. Within the protein–protein interactions (PPI), seven proteins exhibited the highest interactions, which primarily involved two downregulated genes (SAUR and GA3ox) and eight upregulated genes (ACX, MFP2, JAZ, BRI1, BAK1, ETR, EIN2, and SNRK2) of the above pathways. The more upregulated expression of ZEP (in the ABA biosynthesis pathway), DELLA (in the GA signaling pathway), ABF (in the ABA signaling pathway), and ERF1 (in the ETH signaling pathway) in O. taihangensis revealed that it had a relatively higher salt resistance than O. longilobus. This revealed that the responsive patterns to salt stress between the two species had both similarities and differences. The results of this investigation shed light on the potential adaptive mechanisms of O. taihangensis and O. longilobus under cliff environments, while laying a foundation for the study of other cliff species in the Taihang Mountains.

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.

The WRKY gene family comprises important transcription factors widely distributed in plants and plays significant roles in the growth and development, diverse (biotic and abiotic) stress responses, and various biological processes. In the current study, 96 identified HvLWRKY genes were classified into three groups and seven subgroups. Among these, 89 genes possessed the conserved domain WRKYGQK. A total of ten motifs were harbored in HvLWRKY genes with two to four introns. Fragmental duplication was suggested to be the prime force that drove the evolution of HvLWRKY genes. A high degree of collinearity was observed between barley and Triticum spelta. Cis-elements of HvLWRKYs were closely associated with abiotic stress, light response, and hormone response; however, there were differences in the numbers among groups. HvLWRKY genes, even the paralogous gene pairs, from different clades were differentially regulated under cold treatments in two landraces. HvLWRKY33, 43, 44, 57, 65, and 77 were homologous with the relative AtWRKY genes in Arabidopsis thaliana. They are suggested to regulate abiotic and pathogen resistance of two barley landraces via SA and JA pathways. Meanwhile, some genes (for example, HvLWRKY1 and HvLWRKY32) were specifically expressed in either cold-tolerant or cold-sensitive landraces. Under cold stress, different cold-responsive patterns occurred in different barley landraces. These findings provide a foundation for further studies on cold resistance in barley landraces and offer new insights for application of WRKY genes in barley breeding.