Circular RNAs (circRNAs) serve as covalently closed single-stranded RNAs and have been proposed to influence plant development and stress resistance. Grapevine is one of the most economically valuable fruit crops cultivated worldwide and is threatened by various abiotic stresses. Herein, we reported that a circRNA (Vv-circPTCD1) processed from the second exon of the pentatricopeptide repeat family gene PTCD1 was preferentially expressed in leaves and responded to salt and drought but not heat stress in grapevine. Additionally, the second exon sequence of PTCD1 was highly conserved, but the biogenesis of Vv-circPTCD1 is species-dependent in plants. It was further found that the overexpressed Vv-circPTCD1 can slightly decrease the abundance of the cognate host gene, and the neighboring genes are barely affected in the grapevine callus. Furthermore, we also successfully overexpressed the Vv-circPTCD1 and found that the Vv-circPTCD1 deteriorated the growth during heat, salt, and drought stresses in Arabidopsis. However, the biological effects on grapevine callus were not always consistent with those of Arabidopsis. Interestingly, we found that the transgenic plants of linear counterpart sequence also conferred the same phenotypes as those of circRNA during the three stress conditions, no matter what species it is. Those results imply that although the sequences are conserved, the biogenesis and functions of Vv-circPTCD1 are species-dependent. Our results indicate that the plant circRNA function investigation should be conducted in homologous species, which supports a valuable reference for further plant circRNA studies.

The genus Vitis, known for its economically important fruit—grape—is divided into three geographical groups, American, East Asian, and Eurasian, along with a hybrid group. However, previous studies on grape phylogeny using chloroplast genomes have been hindered by limited sample sizes and inconsistent methodologies, resulting in inaccuracies. In this study, we employed the GetOrganelle software with consistent parameters to assemble the chloroplast genomes of 21 grape cultivars, ensuring comprehensive representation across four distinct groups. A comparative analysis of the 21 grape cultivars revealed structural variation, showing chloroplast genome sizes ranging from 160,813 bp to 161,275 bp. In 21 Vitis cultivars, genome annotation revealed 134 to 136 genes, comprising 89 to 91 protein-coding genes (PCGs), 37 tRNAs, and 8 rRNAs. Our observations have pinpointed specific occurrences of contraction and expansion phenomena at the interfaces between inverted repeat (IR) regions and single-copy (SC) regions, particularly in the vicinity of the rpl2, ycf1, ndhF, and trnN genes. Meanwhile, a total of 193 to 198 SSRs were identified in chloroplast genomes. The diversification pattern of chloroplast genomes exhibited strong concordance with the phylogenetic relationships of the Euvitis subgenera. Phylogenetic analysis based on conserved chloroplast genome strongly clustered the grape varieties according to their geographical origins. In conclusion, these findings enhance our understanding of chloroplast genome variation in Vitis populations and have important implications for cultivar selection, breeding, and conservation efforts.

Summary: Circular RNAs (circRNAs) are single‐stranded, covalently closed RNA molecules that arise from exon back‐splicing. The identification and function investigation of circRNAs have been comprehensively explored in plants, however, the regulatory mechanisms are largely unknown. This study employed genetic transformation, circRNA pull‐down assay, ribonucleoprotein immunoprecipitation (RIP), and the circRNA trimolecular fluorescence complementation (cTriFC) system to investigate the function and regulatory mechanisms of a circRNA Vv‐circCOR27 in grapevine. Vv‐circCOR27 expression was suppressed under heat stress and was markedly lower in thermotolerant cultivars. In addition, overexpression of Vv‐circCOR27 , using an expression cassette with endogenous introns inserted into circRNA‐producing exons to minimize the background level of cognate linear RNA, was shown to exacerbate thermotolerance in grapevine embryogenic calli and seedlings. Biochemical assays demonstrate that Vv‐circCOR27 binds directly to heat shock protein 90.2b (VvHSP90.2b). Further evidence indicated that Vv‐circCOR27 inhibits the interaction between VvHsfA7a and VvHSP90.2b in vivo , which resulted in the downregulation of small heat shock protein (sHSP) genes under heat stress in Vv‐circCOR27 overexpression calli. Collectively, this finding suggests a model in which thermotolerance is fine‐tuned by Vv‐circCOR27 via competitively binding to VvHSP90.2b, thereby attenuating its interaction with VvHsfA7a and subsequent transactivation. This work advances understanding of circRNA regulatory mechanisms in plants.

Circular RNAs (circRNAs) are covalently closed RNA molecules that regulate various biological processes in plants. However, the functions of most identified circRNAs remain unclear. Here, we report a nucleoplasmic-localized circRNA, Vv-circRCD1, derived from exons 2 and 3 of the grape VvRCD1 gene. Overexpression of Vv-circRCD1 significantly shortened primary root length and increased root hair number and length, notably, and improved the salt tolerance in Arabidopsis. Transient overexpression also significantly enhanced salt tolerance of grapevines. In silico analyses confirmed direct sequence complementarity between Vv-circRCD1 and the Vvi-miR399 family, and Vv-circRCD1 and Vvi-miR399 target genes (involved in salt stress responses) showed consistent expression patterns under salt stress, indicating a Vv-circRCD1–Vvi-miR399–target gene regulatory module may mediate salt tolerance. These results not only identified Vv-circRCD1 as a novel regulator of grapevine salt tolerance, but also highlighted its potential in improving crop stress resistance, providing a practical reference for crop breeding.

The grapevine, a globally significant fruit and an essential fruit tree species in China, is vulnerable to the adverse effects of high temperatures. Understanding the roles of microRNA and transcription factors in plant development and stress resistance is crucial for mitigating the impact of high temperature on grape growth and yield. This study investigates the response of miRNA to high-temperature stress in grape leaves. The expression level of Vvi-miR3633a was found to be inhibited under heat treatment in both Thompson seedless and Shen yue varieties, while its potential target genes (Vv-Atg36 and Vv-GA3ox2) were induced. Through transgenic overexpression experiments, it was demonstrated that Vvi-miR3633a plays a role in thermal response by affecting the expression of target genes. Furthermore, under heat stress conditions, overexpression of Vvi-miR3633a in grape callus decreased heat resistance compared to the control group (CK). The study also revealed that the target genes of Vvi-miR3633a regulate the expression of oxidase synthesis genes VvSOD and VvCAT, leading to reduced oxidase synthesis which may compromise the oxidation system. Additionally, the expression level of heat shock proteins in the transgenic lines was changed compared to the control (CK). Overall, this research provides valuable insights into understanding the molecular mechanisms involved in different crossing/breeding programs to produce heat-resistant grape varieties. Such varieties can be appropriate to propagate in warm climate areas with high temperature conditions.

Grapes (Vitis spp.) are a globally significant fruit crop with a long history of cultivation and substantial cultivar diversity. Their high genetic differentiation and complex evolutionary history make them a valuable system for studying plant evolution. The chloroplast genome, known for its structural conservation and uniparental inheritance, offers a reliable molecular marker for phylogenetic reconstruction. In this study, we sequenced and assembled the complete chloroplast genomes of nine representative grape cultivars, analyzed their phylogenetic relationships, and compared structural variations. All chloroplast genomes displayed a typical quadripartite structure, with high conservation in genomic architecture, gene order and content, codon usage, and simple sequence repeats (SSRs). However, additional sequence comparisons revealed seven regions with high variation, including the genes rbcL and ndhF, and the intergenic regions rps16-trnQ, ndhC-trnV, accD-psaI, ndhF-rpl32, and trnL-ccsA. At the same time, seven natural variation sites were identified in the amino acid sequences of rbcL and ndhF. Additionally, the study’s maximum likelihood (ML) phylogenetic trees and photosynthetic index measurements suggest that developmental characteristics of grape photosynthesis may be related to the evolutionary origins of different populations. This phylogenetic classification not only elucidates the evolutionary origins of these germplasm resources but also provides a foundation for molecular-assisted breeding by identifying distinct genetic groups.