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Dynamic and non-additive gene regulation shapes maize responses to simultaneous salt and cold stress.

Salt and cold stresses often occur together in nature and severely impact crop productivity, yet their transcriptional regulation remains poorly understood. Here, we conducted a time-series transcriptomic analysis of maize under salt, cold, and their combination at 0, 6, 12, and 24 h. Differential expression analysis revealed dynamic, condition-specific gene responses grouped into eight distinct temporal patterns. Promoter motif analysis of genes within each pattern identified 5-39 significantly enriched motifs, with over 40% lacking known counterparts, suggesting the involvement of previously uncharacterized cis-regulatory elements in stress-responsive transcriptional regulation. By comparing combined stress responses to the sum of single-stress effects, we found that about 74% of DEGs showed non-additive patterns, suggesting that combined stress triggers a distinct transcriptional program. Evolutionary analysis showed that additive DEGs tend to be more recently evolved, subject to weaker purifying selection, and enriched in transposed duplications, contrasting with the stronger constraint observed in non-additive DEGs. WGCNA identified 24 co-expression modules, among which 65 hub DEGs were detected in modules significantly correlated with specific stress conditions. Furthermore, we reconstructed 228, 20, and 200 sequential transcription factor cascades spanning 6 h, 12 h, and 24 h under cold, salt, and combined stress, respectively, with no cascade shared across all three conditions. Together, these results reveal that maize responses to combined salt and cold stress are largely non-additive and temporally dynamic, with distinct evolutionary patterns underlying different response types, offering insights and candidate regulators for enhancing crop stress resilience.

Zea mays

Engineering cold stress resilience in capsicum annuum through functional genomics and precision breeding.

This review synthesizes the molecular mechanisms of cold tolerance in pepper, integrating multi-omics data,genome editing, and precision breeding strategies to accelerate the development of cold-resilient cultivars. Cold stress is a significant environmental factor that affects the growth, productivity, and fruit quality of Capsicum annuum by impairing membrane integrity photosynthesis and cellular redox homeostasis. Although pepper has several endogenous cold-responsive regulators such as CaNAC035 and CabHLH035, along with antioxidant defense systems, its cold tolerance remains limited due to low transcriptional activation of key regulators, functional redundancy among cold-responsive genes, and the polygenicity of cold tolerance. These complexities, combined with low genetic diversity and linkage drag, have hindered the improvement of cold-resistant cultivars through conventional breeding. This review brings together the recent progress in understanding the molecular mechanisms of cold stress perception, signal transduction, transcriptional regulation, metabolic reprogramming, and phytohormone interactions in pepper. Precision Breeding 2.0 is a new innovation that combines the integration of multi-omics-based target identification with next-generation genome-editing techniques, allowing precise and multiplex engineering of complex and interconnected regulatory networks instead of single genes. We cover new approaches such as engineering the DREB/CBF pathway, allele-specific editing and targeted disruption of negative regulators to enhance the pathway(s) involved in cold response. Moreover, we propose a roadmap for integration of transcriptomics, proteomics, metabolomics, high-throughput phenomics, and speed breeding to accelerate the identification, validation, and deployment of superior alleles to boost cold tolerance. This review provides a foundation for developing climate-resilient pepper cultivars by connecting functional genomics with precision genome engineering approaches to maintain productivity under variable environmental conditions.

Capsicum

Genome-wide identification and evolutionary analysis of the ERF-VII gene family in the tea plant (Camellia sinensis) and functional characterization of CsRAP2.2 in response to cold stress.

The ERF-VII gene family, a critical branch of the AP2/ERF superfamily, is central to plant stress adaptation. However, its evolutionary history and function in tea plant (Camellia sinensis) remain unclear. Here, we performed integrated evolutionary, genomic, and functional analyses of ERF-VII genes across 14 plant lineages and 20 tea plant cultivars. The phylogenetic analysis revealed that ERF-VII proteins originated after vascular plant divergence, coinciding with the emergence of the N-terminal MCGGA/I motif linked to the oxygen-dependent N-degron pathway. Gymnosperms retained few conserved members, whereas angiosperms exhibited lineage-specific expansion-extensive in monocots via whole-genome duplication, moderate in eudicots with functional diversification. Pan-genome analysis across 20 tea plant cultivars further revealed varietal differences in ERF-VII gene distribution. Transcriptome profiling via the Tea Plant Information Archive identified CsRAP2.2 as a cold-inducible ERF-VII member with sustained expression under low-temperature stress. Functional assays demonstrated that silencing CsRAP2.2 reduced cold tolerance, while overexpression in tea leaves and heterologous expression in Arabidopsis thaliana enhanced cold tolerance by maintaining photosystem II efficiency, reducing membrane lipid peroxidation, and improving antioxidant capacity. Weighted gene co-expression network analysis positioned CsRAP2.2 as a regulatory hub integrating cold, hormone, and oxygen-sensing pathways. These results clarify the evolutionary trajectory of ERF-VII genes and establish CsRAP2.2 as a core cold-tolerance regulator in tea plant. These findings may inform future breeding of cold-resilient tea cultivars.

Camellia sinensis

Uncovering molecular regulatory networks of low-temperature stress response in Trachinotus ovatus via integrated transcriptome and metabolome analyses.

Golden pompano (Trachinotus ovatus) is one of the most economically important marine fish species in China. It is susceptible to low-temperature stress, which significantly challenges its production and supply. Nevertheless, study on the regulatory mechanisms underlying low-temperature stress responses in golden pompano remains limited. Here, we firstly performed a time-series transcriptome analysis to reconstruct dynamic response patterns under low-temperature stress in golden pompano. Transcriptome profiling identified common differentially expressed genes (DEGs), including fos, hlf, and hmgb1, as well as condition-specific DEGs across distinct low-temperature stress groups. Based on cluster analysis, all DEGs were classified into five distinct expression patterns, reflecting diversified regulation of expression in golden pompano during low-temperature stress. Furthermore, condition-specific regulatory modules were explored via weighted gene co-expression network analysis (WGCNA), highlighting that the two module hub genes, serbf2 and lipc, might respond to low-temperature stress by regulating the lipid catabolic process. Subsequently, untargeted metabolomic analysis revealed that glycerophospholipid metabolism was a significantly enriched common pathway, highlighting its crucial role in mediating the response to low-temperature stress. Finally, by integrating transcriptomic and metabolomic analyses, a gene-metabolite interaction network associated with glycerophospholipid metabolism under low-temperature stress was established. These findings underscore the significance of multiple candidate genes and glycerophospholipid metabolism in golden pompano's response to low-temperature stress, thereby laying a solid molecular foundation for the development of low-temperature-tolerant fish strains.

Animals

Distinct molecular responses to acute cold exposure revealed by comparative transcriptomic and metabolomic profiling in the bay scallop Argopecten irradians.

Acute cold stress can elicit distinct molecular responses even when bay scallop populations show similar phenotypic outcomes. We compared a seventh-generation fast-growing bay scallop line (BS) with a commercial control population (CC) during a 72-h acute cold exposure at -1 ± 0.3 °C. RNA-seq was used as the discovery layer, representative BS cold-responsive genes were evaluated by qRT-PCR, and paired LC-MS profiles provided a comparative metabolic layer. At baseline, 138 genes differed between BS and CC; after cold exposure, 134 of these baseline differences disappeared and 61 of 65 cold-state differences newly emerged. BS showed a larger transcriptomic response magnitude than CC, with 1129 cold-responsive genes compared with 28 genes in CC, and this ordering remained robust across multiple sensitivity analyses. Survival after 72 h was identical in BS and CC (83/90, 92.2% in each population). Biochemical responses were time-dependent and marker-specific: CAT, LZM, T-SOD and T-AOC showed population-by-time interactions, whereas GSH-Px and MDA did not, and the 72-h differences were not consistently favourable to BS. Metabolomic cold effects were strongly concordant between populations, and no feature showed a significant population-by-cold interaction. Features putatively assigned to arachidonic acid metabolism were enriched, but this provider-annotated pathway signal remains exploratory because authentic-standard confirmation was not performed. These findings indicate population-specific differences in molecular responsiveness but do not establish superior cold tolerance in BS.

Animals

The chloroplast 16S rRNA dimethyltransferase BrPFC1 is required for Brassica rapa development under chilling stress.

Chloroplast ribosomal RNA (Ch-rRNA) methylation is critical for plant development and response to low temperatures. Several Ch-rRNA methyltransferases and their catalytic modes, as well as biological relevance, have been reported in model plant species. However, Ch-rRNA methyltransferases and their functional significance remain poorly characterized in crops, including leafy vegetables such as Chinese cabbage. In this study, we screened an EMS-mutagenized Chinese cabbage population and identified a yellow inner leaf (yif) mutant. This mutant develops yellowing inner leaves with reduced chlorophyll accumulation and ultrastructure-impaired chloroplasts under low-temperature conditions. Genetic analysis revealed a premature termination mutation in BrPFC1, encoding the chloroplast-localized 16S rRNA dimethyltransferase. The BrPFC1 mutation (yif) disrupts the dimethylation of 16S rRNA. The cold-sensitive phenotype of the yif mutant can be explained by temperature-dependent defects in the maturation and assembly of chloroplast ribosomes at 4°C. Through integrated analysis of chloroplast and nuclear transcriptomes coupled with translational profiling at 25°C and 4°C, we established that low temperature preferentially upregulates transcripts encoding nuclear-derived ribosomal proteins, while defective 16S rRNA specifically compromises the translational efficiency of chloroplast-encoded photosynthetic complex and ribosomal protein at 4°C. These findings establish rRNA modification by BrPFC1 as a critical regulatory layer for optimizing chloroplast translational efficiency at 4°C, providing mechanistic insights into post-translational adaptation strategies in Chinese cabbage.

Chloroplasts

Analysis of genetic differences underlying chilling stress tolerance using whole genome Re-Sequencing in walnut (Juglans regia L.).

Walnut (Juglans regia L.) is prized worldwide for both its nutritional value and economic importance, yet it remains vulnerable to cold stress, with significant differences in tolerance among varieties. This study combined physiological analyses with whole-genome resequencing (WGS) to evaluate the cold stress responses of two varieties, &#x2018;Qingxiang&#x2019; and &#x2018;Liaoning No.8&#x2019;. Under chilling stress (0&#xa0;&#xb0;C), we measured electrolyte leakage and antioxidant enzyme activity, applying both exogenous methyl jasmonate (MeJA) and the jasmonate inhibitor DIECA. Genomic variations were analyzed using WGS. Results showed that &#x2018;Liaoning No.8&#x2019; exhibited superior cold tolerance. Application of MeJA reduced electrolyte leakage by 37% and MDA accumulation by 52% on average, whereas DIECA exacerbated stress-related damage. WGS achieved 16.24&#x2013;16.26&#xd7; coverage and identified 2.73&#x2013;2.78&#xa0;million SNPs, 378&#x2013;382k InDels, 25&#x2013;26k SVs, and 7.2&#x2013;7.9k CNVs. Twenty genes containing sequence variants showed transcriptional responses under cold stress that were significantly correlated with mutation density (r&#x2009;=&#x2009;0.62, P&#x2009;<&#x2009;0.01). One gene, XM_018985465.2, which lacked SNPs in &#x2018;Liaoning No.8&#x2019;, was expressed 4.2 times more in this variety, suggesting cis-regulatory influence. These findings highlight the role of jasmonic acid signaling in enhancing cold tolerance in walnut and offer genomic insights into its underlying adaptive mechanisms.

Juglans

Integrative analysis of transcriptome and chromatin accessibility reveals promoter-proximal regulation and identifies candidate ABC transporters associated with cold stress responses in maize.

BACKGROUND: Low-temperature stress is a formidable environmental constraint that severely limits the growth and productivity of maize (Zea mays L.), particularly during the highly vulnerable early seedling stage. While cold tolerance is a critical agronomic objective, the integrated transcriptional and epigenetic regulatory mechanisms that govern this trait remain largely elusive. Characterizing these coordinated molecular networks is fundamental to the genetic enhancement of cold resilience in maize. METHODS: Using two maize inbred lines contrasting in chilling response (ZHB12 tolerant, B73 sensitive), we performed integrative time&#x2011;course RNA&#x2011;seq and ATAC&#x2011;seq to thoroughly and systematically characterize the precise dynamic interplay between gene expression and chromatin accessibility under cold stress conditions at the seedling stage. RESULTS: Physiological assessments confirmed that ZHB12 possesses superior cold tolerance, manifested by significantly attenuated electrolyte leakage and reduced foliar damage compared to B73. Transcriptomic profiling revealed a massive, time-dependent divergence in gene expression between the two genotypes, with a major regulatory transition identified at 24&#xa0;h of cold exposure. Functional enrichment analysis demonstrated that ZHB12 preferentially activates a robust defense repertoire, including Photosystem II electron transport, diterpenoid biosynthesis, and ATP biosynthetic pathways. Notably, multiple ATP-binding cassette (ABC) transporter genes were coordinately upregulated under chilling, suggesting their potential involvement in cellular homeostasis. ATAC-seq analysis indicated that cold stress is associated with chromatin remodeling in ZHB12, with increased accessibility observed in proximal promoter regions. Integrative analysis identified a core set of dual-responsive genes, in which increased promoter accessibility coincided with transcriptional upregulation. These genes were predominantly enriched in transporter activity and transcriptional regulation, suggesting potential epigenetic link to the superior stress response of ZHB12. CONCLUSION: Our findings reveal extensive transcriptional and chromatin accessibility changes in ZHB12 under cold stress. The observed associations between promoter accessibility and gene activation, particularly in genes involved in transport processes, highlight candidate regulators potentially contributing to cold tolerance. This study provides a molecular framework and identifies high-value candidate genes that may inform future efforts in breeding cold-tolerant maize, pending functional validation.

Zea mays

Construction of cDNA library of Dalbergia odorifera induced by low temperature stress and screening of low temperature tolerant genes.

To systematically analyze the gene function of Dalbergia odorifera, the seedlings of D. odorifera were treated with low-temperature stress for 6&#x2009;h. Total RNA was extracted from a mixture of seedling roots, stems, and leaves, and a low-temperature-induced D. odorifera yeast cDNA expression library was constructed. The library volume was 1.032&#x2009;&#xd7;&#x2009;108 CFU, and the PCR (Polymerase Chain Reaction) identification of the library bacterial fluid showed that the amplification was around 1000&#x2009;bp, with a single randomly distributed band, indicating that the library had been recombinantly inserted into the pYES2 vector. The GO (Gene Ontology) analysis showed that the library genes were mainly involved in metabolic and stress signaling pathways. The KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway enrichment analysis showed that the genes were primarily related to energy and metabolic pathways. Twenty-one genes were screened or obtained at -20&#xb0;C for low-temperature tolerance. In addition, the organ expression profiles of the candidate genes were analyzed based on RNA-seq data, and the expression profiles of the candidate genes under low-temperature stress were also examined. The construction of the yeast library provides genetic resources for the analysis of the mechanism of low-temperature tolerance of D. odorifera, which is important for comprehending and utilizing the genetic resources of D. odorifera.

Gene Library

Genome-wide identification and cold-stress-responsive expression analysis of the NOX gene family in Cucumis melo.

NADPH oxidases (NOXs) are crucial enzymes for reactive oxygen species (ROS) generation in plants and play vital roles in growth, development, and stress responses. To elucidate the sequence characteristics of the NOX gene family and its low-temperature response patterns in melon (Cucumis melo L.), this study conducted genome-wide identification and expression profiling of NOX family members using bioinformatics analysis, RNA-seq transcriptome sequencing, and real-time quantitative PCR (RT-qPCR). The results revealed that eight NOX members were identified in the melon genome, distributed across six chromosomes. All members harbored conserved domains including Ferric_reductase, FAD_binding_8, NAD_binding_6, and NADPH_Ox, and the encoded proteins were generally basic and hydrophilic. Phylogenetic analysis classified the NOX proteins into five subgroups. Synteny analysis indicated the presence of only one pair of intraspecific duplicated genes in melon, which was under purifying selection. The promoter regions contained multiple hormone- and stress-responsive cis-acting elements, with CmNOX2 and CmNOX4 harboring low-temperature responsive elements. Following treatment at 4&#x2103; for 24 h and 48 h, leaf relative electrolyte leakage (REL) increased from 28.33% to 42.67% and 52.67%, respectively; transcriptome analysis identified 5,633 and 6,882 differentially expressed genes (DEGs), respectively. Cold-responsive genes exhibited significant differential expression, with SLAC1 and CPK19 showing sustained upregulation. RT-qPCR results demonstrated that the expression of CmNOX2, CmNOX5, CmNOX6, and CmNOX7 was significantly downregulated after low-temperature treatment, whereas CmNOX4 expression was significantly upregulated at 48 h. Integrating promoter elements and expression characteristics, CmNOX4 may represent an important candidate gene involved in melon low-temperature response. This study systematically characterized the structure, evolution, and expression patterns of the melon NOX gene family, identified candidate genes responsive to low temperature, and provides a reference for further investigation into the mechanisms underlying melon cold adaptation.

Cucumis melo

Maternal and developmental temperature modulate adult response to temperature in Drosophila melanogaster.

Beyond inherited genes and environmentally induced changes in gene expression, phenotypes can also be shaped by parental effects-an effect from a parental phenotype that causes modifications in offspring traits, which cannot be solely explained by the parental or offspring genomes. Such effects may prepare offspring for future environmental conditions and contribute to phenotypic plasticity, including responses to temperature. While temperature-induced plasticity has been extensively studied, the relative contributions of parental versus direct environmental cues remain poorly understood. The fruit fly Drosophila melanogaster is a powerful model for studying physiological and behavioral adaptation to temperature. Flies inhabit environments spanning broad thermal ranges and show evidence of parental effects, such as increased heat tolerance in offspring from warm-reared parents. Here, we exposed mothers to two experimental temperatures and split their broods between the same two temperatures to estimate the relative importance of maternal and developmental effects on adult physiological and developmental responses to temperature. We find that the reaction norms of locomotor activity under gradually increasing temperatures, responses to heat-shock and cold-shock, and fecundity are mostly governed by direct plastic responses to developmental environment. We detected comparatively weak maternal effects in the response to heat-shock, fecundity, and grand-offspring survival where matched environments counteracted the effects of direct offspring experience. We conclude that thermal experience during development is the primary determinant of phenotypic plasticity in D. melanogaster, while maternal experience contributes a small but non-negligible component.

Animals