Evolutionary Divergence Research Articles

Linking phenotypic variation to patterns of genetic isolation along a speciation continuum.

Investigating taxa at varying stages of divergence can shed light on the evolutionary forces that lead to reproductive isolation and eventual speciation. The forces promoting isolation vary in space and time, which makes it difficult to reconstruct the trajectory that resulted in the divergence observed among species today. The red macroalgal genus Plocamium is known worldwide for its cryptic genetic and chemical diversity. Previous work on the genus Plocamium in Antarctica observed two haplotypes along the western Antarctic Peninsula that have been treated as the same species. Using 10 microsatellite loci, we observed that these two haplotypes correspond to two highly divergent, co-occurring genetic entities in Antarctic Plocamium, which are located within close vicinity of each other at the same sites. The morphology of the reproductive structures, a feature commonly used to identify cryptic species in Plocamium, as well as the timing of reproduction, differed significantly between the two genetic entities. Altogether, this suggests that two Antarctic Plocamium species exist on the western Antarctic Peninsula. We observed evidence for high levels of selfing in both genetic entities, which likely exacerbated the lack of gene flow between them. In addition, we identified concomitant chemodiversity that generates compelling evidence of early evolutionary divergence within one of these entities. This chemodiversity has ecological consequences for its main grazer, which alludes to one putative evolutionary driver of divergence. Antarctic Plocamium spp. provide a promising model system for investigating the eco-evolutionary forces that initiate and maintain species boundaries.

An enhanced dung beetle optimizer with multiple strategies for robot path planning

In order to make up for the shortcomings of the original dung beetle optimization algorithm, such as low population diversity, insufficient of global exploration ability, being easy to fall into local optimization and unsatisfactory convergence accuracy, etc. An improved dung beetle optimization algorithm using hybrid multi- strategy is proposed. Firstly, the cubic chaotic mapping approach is used to initialize the population to improve the diversity, expand the search range of the solution space, and enhance the global optimization ability. Secondly, the cooperative search algorithm is utilized to strength communication between individual dung beetles and dung beetle groups in foraging stage to expand the search range of the solution space and enhance the global optimization ability. Thirdly, T-distribution mutation and differential evolutionary variation strategies are introduced to provide perturbation to enhance the diversity of the population and avoid falling into local optimization. Fourthly, the proposed algorithm(named as SSTDBO) is compared with other optimization algorithms, including GODBO, QHDBO, DBO, KOA, NOA, WOA and HHO, by 29 benchmark test functions in CEC2017. The results show that the proposed algorithm has stronger robustness and optimization ability, and algorithm’s performance has substantially enhanced. Finally, the proposed algorithm is applied to solve the real-world robot path planning engineering cases, to demonstrate its effectiveness in dealing with real optimization engineering cases, which further verified how noteworthy the enhanced strategy’s efficacy and the enhanced algorithm’s superiority are in addressing real-world engineering cases.

Different intestinal ecological niches drive the divergent evolution of probiotics in the gut

Different intestinal ecological niches drive the divergent evolution of probiotics in the gut

Understanding HLA-DQ in renal transplantation: a mini-review

Human leukocyte antigen (HLA) mismatching, particularly with HLA-DQ, significantly impacts the development of donor-specific antibodies (DSA) and transplant outcomes. HLA-DQ antibodies are highly immunogenic and detrimental, necessitating advanced high-resolution HLA typing to improve mismatch assessment and clinical risk evaluation. Traditional serological or low-resolution typing often misclassifies mismatches, leading to inaccuracies in assessing immunogenicity and predicting outcomes. Emerging molecular mismatch algorithms refine immunogenicity assessments by analyzing amino acid differences and structural interactions. These tools show promise for personalizing transplant protocols but have limitations, such as variability in predicting individual patient outcomes. Immunogenicity of mismatches also depends on evolutionary divergence and specific amino acid differences, with studies revealing that certain evolutionary lineages and polymorphisms influence T-cell alloreactivity and DSA development. Complexities in HLA-DQ protein expression, including combinatorial diversity of heterodimers and inter-isotypic heterodimers, further complicate risk evaluation. Expression levels, influenced by tissue specificity and inflammatory stimuli, and alternative splicing of HLA-DQ transcripts add additional layers of variability. Future clinical applications, enabled by high-resolution HLA typing, may include refined graft selection, improved DSA monitoring, and individualized therapy. However, understanding the precise mechanisms of HLA-DQ immunogenicity remains a priority for advancing transplantation science and enhancing patient outcomes.

Distinct domain regions of NIN and NLP1 mediate symbiotic and nitrate signaling in Medicago truncatula.

Nodule Inception (NIN) and NIN-like protein 1 (NLP1), both belonging to the RWP-RK type transcription factors, play critical roles in plant development. Specifically, NIN is pivotal in facilitating root nodule symbiosis in nitrogen-starved conditions, while NLP1 coordinates nodulation in response to nitrate level. In this study, we conducted domain swapping experiments between NIN and NLP1 in Medicago truncatula to elucidate the functional significance of their respective domains. The findings reveal that the C-terminal regions, including the RWP-RK and PB1 domains of NIN, can substitute for those of NLP1, whereas reciprocal substitution do not yield equivalent outcomes. Moreover, our data emphasize the critical role of PB1-mediated interactions for NLP1's activity, a feature not essential for NIN. Additionally, the N-terminal segment, conserved in NLPs but containing deletions or mutations in NIN, is essential for the proper functioning of both NIN and NLP1. Collectively, our research suggests the evolutionary divergence of NIN from ancestral NLPs, indicating specific adaptations that have enabled NIN as a central regulator in root nodulation processes.

A comparative analysis of sequence composition in different lots of a phage display peptide library during amplification

BackgroundTo develop efficient selection strategies and improve the discovery of promising ligands, it is highly desirable to analyze the sequence composition of naïve phage display libraries and monitor the evolution of their peptide content during successive rounds of amplification. In the current study, we performed a comparative analysis of the compositional features in different lots of the same naïve phage display library and monitored alterations in their peptide compositions during three rounds of amplification.MethodsWe conducted three rounds of duplicate serial amplification of two different lots of the Ph.D.™-12 phage display library. DNA from the samples was subjected to Next-Generation Sequencing (NGS) using an Illumina platform. The NGS datasets underwent a variety of bioinformatic analyses using Python and MATLAB scripts.ResultsWe observed substantial heterogeneity in the sequence composition of the two lots indicated by differences in the enhanced percentage of wildtype clones, reduced diversity (number of unique sequences), and increased enrichment factors (EFs) during amplification as well as by observing no common sequence between lots and decreased number of common sequences between the naïve library and the consecutive rounds of amplification for each lot. We also found potential propagation-related target-unrelated peptides (TUPs) with the highest EFs in the two lots, which were displayed by the fastest-propagating phage clones. Furthermore, motif analysis of the most enriched subpopulation of amplified libraries led to the identification of some motifs hypothesized to contribute to the increased amplification rates of the respective phage clones.ConclusionOur results highlight tremendous heterogeneity in the peptide composition of different lots of the same type of naïve phage display library, and the divergent evolution of their compositional features during amplification rounds at the amino acid, peptide, and motif levels. Our findings can be instrumental for phage display researchers by bringing fundamental insights into the vast extent of non-uniformity between phage display libraries and by providing a clear picture of how these discrepancies can lead to different evolutionary fates for the peptide composition of phage pools, which can have profound impacts on the outcome of phage display selections through biopanning.

Organ-specific microenvironments drive divergent T cell evolution in acute graft-versus-host disease.

Tissue-specific T cell immune responses play a critical role in maintaining organ health but can also drive immune pathology during both autoimmunity and alloimmunity. The mechanisms controlling intratissue T cell programming remain unclear. Here, we leveraged a nonhuman primate model of acute graft-versus-host disease (aGVHD) after allogeneic hematopoietic stem cell transplantation to probe the biological underpinnings of tissue-specific alloimmune disease using a comprehensive systems immunology approach including multiparameter flow cytometry, population-based transcriptional profiling, and multiplexed single-cell RNA sequencing and TCR sequencing. Transcriptional profiling revealed substantial biological differences between T cells infiltrating the lung and liver during aGVHD. These included enrichment for transcriptional pathways controlling extracellular matrix remodeling and chemotaxis in the lung and enrichment for transcriptional pathways linked to nucleic acid metabolism and proliferation in the liver. Single-cell RNA sequencing and TCR sequencing substantiated divergent organ-specific transcriptional programing of tissue-infiltrating T cells, which was linked to clonal expansion, with expanded clones progressively enriched for C-X3-C motif chemokine receptor 1 (CX3CR1)-expressing CD8 effector T cells in the lung and eomesodermin (EOMES)-expressing CD8 effector-memory T cells in the liver. This divergent evolution of T cells was maintained even for T cells sharing the same TCRs, indicating its independence from antigen specificity. Together, these results provide insights into the role that tissue microenvironment-derived signals play in local T cell transcriptional programming during alloimmune-mediated clonal expansion and suggest potential opportunities to develop tissue-specific therapeutics to curtail pathogenic immunity after transplant.

Evolutionary divergence between homologous X-Y chromosome genes shapes sex-biased biology.

Sex chromosomes are a fundamental aspect of sex-biased biology, but the extent to which homologous X-Y gene pairs ('the gametologs') contribute to sex-biased phenotypes remains hotly debated. Although these genes tend to exhibit large sex differences in expression throughout the body (XX females can express both X members, and XY males can express one X and one Y member), there is conflicting evidence regarding the degree of functional divergence between the X and Y members. Here we develop and apply co-expression fingerprint analysis to characterize functional divergence between the X and Y members of 17 gametolog gene pairs across >40 human tissues. Gametolog pairs exhibit functional divergence between the sexes that is driven by divergence between the X versus Y members (assayed in males), and this within-pair divergence is greatest among pairs with evolutionarily distant X and Y members. These patterns reflect that X versus Y gametologs show coordinated patterns of asymmetric coupling with large sets of autosomal genes, which are enriched for functional pathways and gene sets implicated in sex-biased biology and disease. Our findings suggest that the X versus Y gametologs have diverged in function and prioritize specific gametolog pairs for future targeted experimental studies.

Novel H16N3 avian influenza viruses isolated from migratory gulls in China in 2023

As a rare subtype of avian influenza virus, H16 viruses are predominant in gulls but rarely found in domestic birds. The low prevalence of H16 viruses has limited our understanding of their epidemiology and evolutionary dynamics. In this study, we isolated three novel H16N3 viruses from migratory gulls in East Asian–Australasian Flyway in eastern China in 2023, which are significantly different from previously identified isolates. To fully understand the epidemiology and genetics characteristics of the global H16 viruses, we compared the host divergence of several rare subtypes and determined that the H13 and H16 subtypes were predominantly pooled into different species of gulls by sharing their internal genes, whereas the waterfowl of Anatidae served as the primary natural reservoirs of the H8, H11, H12, H14, and H15 subtypes. Detailed phylogenetic analysis revealed the evolutionary divergence of globally circulating H16 viruses and their frequent gene reassortment. Furthermore, the gull origin H13 and H16 viruses collectively served as gene donors for the newly emerged highly pathogenic clade 2.3.4.4b H5N1 viruses because the H13/H16-like PA, NP, and NS genes have been introduced into circulating H5N1 viruses since May 2022 in Europe. To date, the H5N1 reassortants containing the H13/H16-like gene segments have been detected in wild and domestic birds and resulted in mammal and human infections. These results improve our knowledge of the ecology and genetics of H16 viruses and emphasize the need for surveillance to monitor the emergence of novel avian influenza viruses in migratory birds.

Co-Conservation of Synaptic Gene Expression and Circuitry in Collicular Neurons.

The superior colliculus (SC), a midbrain sensorimotor hub, is anatomically and functionally similar across vertebrates, but how its cell types have evolved is unclear. Using single-nucleus transcriptomics, we compared the SC's molecular and cellular organization in mice, tree shrews, and humans. Despite over 96 million years of evolutionary divergence, we identified ∼30 consensus neuronal subtypes, including Cbln2 + neurons that form the SC-pulvinar circuit in mice and tree shrews. Synapse-related genes were among the most conserved, unlike neocortex, suggesting co-conservation of synaptic genes and circuitry. In contrast, cilia-related genes diverged significantly across species, highlighting the potential importance of the neuronal primary cilium in SC evolution. Additionally, we identified a novel inhibitory SC neuron in tree shrews and humans but not mice. Our findings reveal that the SC has evolved by conserving neuron subtypes, synaptic genes, and circuitry, while diversifying ciliary gene expression and an inhibitory neuron subtype.

In situ diversification and adaptive introgression in Taiwanese Scutellaria.

Island habitats provide unique opportunities to study speciation. Recent work indicates that both ex situ origination and in situ speciation contribute to island species diversity. However, clear evidence of local adaptation of endemic plant species on islands requires in-depth studies, which are scarce. This study underscores the importance of local adaptation in maintaining species boundaries by examining how adaptive introgression, hybridization, and local adaptation contribute to genetic variation in island species. Multilocus genome scanning of 51 nuclear genes was used to investigate the evolutionary relationships of the Scutellaria species complex on Taiwan Island and assess the role of in situ diversification in generating high endemism and genetic diversity. Interspecies introgressions were detected by phylogenetic networks and ABBA-BABA-based analysis, suggesting ongoing or recent speciation processes. Coalescent-based simulation identified hybrid speciation in Scutellaria taiwanensis and Scutellaria hsiehii, with evidence of hybridization between more than two parental species. Genotype-environment association studies revealed that the influence of climate, particularly precipitation- and temperature-related factors, contributed to adaptive genetic divergence between species. Additionally, adaptive introgression related to environmental pressures that may have facilitated the colonization of new island habitats were identified. This research illustrates how hybridization, introgression, and adaptation shaped the evolutionary histories and divergence of this island-endemic plant species complex and sheds light on the multifaceted mechanisms of speciation on semi-isolated islands.

Decoding MexB efflux pump genes: structural, molecular, and phylogenetic analysis of multidrug-resistant and extensively drug-resistant Pseudomonas aeruginosa.

Emerging drug resistance in Pseudomonas aeruginosa is of great concern in clinical settings. P. aeruginosa activates its efflux-pump system in order to evade the effect of antibiotics. The current investigation aims to detect MexB genes in P. aeruginosa, their structural and molecular analysis and their impact on antimicrobial susceptibility profiling. A total of 42 clinical specimens were aseptically collected from hospitalized patients who had underlying infections related to medical implants. Matrix-assisted laser desorption ionization-time of flight (MALDI-ToF) were used for the identification of isolates. The methods used in this study were antibiotic susceptibility profiling, minimum inhibitory concentration (MIC), polymerase chain reaction (PCR), sanger sequencing, phylogenetic analysis, MolProbity score, Ramachandran plot analysis and multiple sequence alignment. The highest resistance was shown by P. aeruginosa against cefoperazone (67%), gentamycin and amikacin (66%) each, followed by cefotaxime (64%). The prevalence of multi-drug resistant (MDR) and extensively drug resistant (XDR) was 57% and 12%, respectively. The presence of an active efflux-pump system was indicated by the MexB genes found in most of the resistant isolates (p<0.05). Following addition of efflux pump inhibitor carbonyl cyanide m-chlorophenyl hydrazone (CCCP), a significant decrease (p<0.05) in MIC was observed in resistance, that revealed the presence of active efflux pump system. Phylogenetic analysis revealed evolutionary relationships with the P. aeruginosa strains isolated in Switzerland, Denmark and Germany. Protein domain architecture revealed that MexB gene proteins were involved in particular efflux pump function. Protein sequences aligned by multiple sequence alignment revealed conserved regions and sequence variants, which suggested antibiotic translocation and evolutionary divergence. These highly conserved regions could be used for diagnostic purposes of efflux pump MexB genes. To avoid their spread in hospital settings, responsible authorities ought to begin rigorous initiatives in order to reduce the prevalence of multi-drug resistant, extensively drug resistant, and efflux pump carrying isolates in clinical settings.

An Active-Site Bro̷nsted Acid-Base Catalyst Destabilizes Mandelate Racemase and Related Subgroup Enzymes: Implications for Catalysis.

Enzymes of the enolase superfamily (ENS) are mechanistically diverse, yet share a common partial reaction, i.e., the metal-assisted, Bro̷nsted base-catalyzed abstraction of the α-proton from a carboxylate substrate to form an enol(ate) intermediate. Although the catalytic machinery responsible for the initial deprotonation reaction has been conserved, divergent evolution has led to numerous ENS members that catalyze different overall reactions. Using differential scanning calorimetry, we examined the contribution of the Bro̷nsted acid-base catalysts to the thermostability (Tm) of four members of the mandelate racemase (MR)-subgroup of the ENS: MR, d-tartrate dehydratase, l-talarate/galactarate dehydratase, and l-fuconate dehydratase. Each enzyme contains an active-site Lys (part of a KxK motif) and His, which act as Bro̷nsted acid-base catalysts. The KxK → KxM substitutions increased the thermostability in all four enzymes with the effect being most prominent for MR (ΔTm = +8.6 °C). The KxK → MxK substitutions decreased the thermostability in all four enzymes, and the His → Asn substitution had a significant stabilizing effect only on MR. Thus, the active sites of MR-subgroup enzymes are destabilized by the Lys Bro̷nsted acid-base catalyst, suggesting that the destabilization energy may be used to drive a conformational change of the enzyme to yield a catalytically competent protonation state upon substrate binding.

Cross-species single-cell transcriptomics reveals neuronal similarities and heterogeneity in amniote pallium.

The amniote pallium, a vital component of the forebrain, exhibits considerable evolutionary divergence across species and mediates diverse functions, including sensory processing, memory formation, and learning. However, the relationships among pallial subregions in different species remain poorly characterized, particularly regarding the identification of homologous neurons and their transcriptional signatures. In this study, we utilized single-nucleus RNA sequencing to examine over 130 000 nuclei from the macaque ( Macaca fascicularis) neocortex, complemented by datasets from humans ( Homo sapiens), mice ( Mus musculus), zebra finches ( Taeniopygia guttata), turtles ( Chrysemys picta bellii), and lizards ( Pogona vitticeps), enabling comprehensive cross-species comparison. Results revealed transcriptomic conservation and species-specific distinctions within the amniote pallium. Notable similarities were observed among cell subtypes, particularly within PVALB + inhibitory neurons, which exhibited species-preferred subtypes. Furthermore, correlations between pallial subregions and several transcription factor candidates were identified, including RARB, DLX2, STAT6, NR3C1, and THRB, with potential regulatory roles in gene expression in mammalian pallial neurons compared to their avian and reptilian counterparts. These results highlight the conserved nature of inhibitory neurons, remarkable regional divergence of excitatory neurons, and species-specific gene expression and regulation in amniote pallial neurons. Collectively, these findings provide valuable insights into the evolutionary dynamics of the amniote pallium.

Vanadium-Dependent Haloperoxidase Gene Evolution in Brown Algae: Evidence for Horizontal Gene Transfer.

Compared with green plants, brown algae are characterized by their ability to accumulate iodine, contributing to their ecological adaptability in high-iodide coastal environments. Vanadium-dependent haloperoxidase (V-HPO) is the key enzyme for iodine synthesis. Despite its significance, the evolutionary origin of V-HPO genes remains underexplored. This study investigates the genomic and evolutionary dynamics of V-HPOs in brown algae, focusing on Laminariales species, particularly Saccharina japonica. Genomic analyses revealed the extensive expansion of the V-HPO gene family in brown algae, with 88 V-HPOs identified in S. japonica, surpassing the number in red algae. Phylogenetic analysis demonstrated distinct evolutionary divergence between brown and red algal V-HPOs, with the brown algal clade closely related to bacterial V-HPOs. These findings suggest horizontal gene transfer (HGT) played a key role in acquiring V-HPO genes, particularly from Acidobacteriota, a bacterial phylum known for genomic plasticity. Additionally, enriched active transposable elements were identified around V-HPO genomic clusters, highlighting their role in tandem gene duplications and rapid HGT processes. Expression profiling further revealed dynamic regulation of V-HPOs in response to environmental conditions. This study provides new insights into how HGT has driven kelp genomic adaptations and enhances understanding of marine ecological success and evolutionary processes.

Unveiling the Role of GhP5CS1 in Cotton Salt Stress Tolerance: A Comprehensive Genomic and Functional Analysis of P5CS Genes.

Proline, a critical osmoregulatory compound, is integral to various plant stress responses. The P5CS gene, which encodes the rate-limiting enzyme in proline biosynthesis, known as ∆1-pyrroline-5-carboxylate synthetase, is fundamental to these stress response pathways. While the functions of P5CS genes in plants have been extensively documented, their specific roles in cotton remain inadequately characterized. In this study, we identified 40 P5CS genes across four cotton species with diverse sequence lengths and molecular weights. Phylogenetic analysis of 100 P5CS genes from nine species revealed three subgroups, with Gossypium hirsutum closely related to Gossypium barbadense. Collinearity analysis highlighted significant differences in collinear gene pairs, indicating evolutionary divergence among P5CS genes in tetraploid and diploid cotton. Exon-intron structures and conserved motifs correlated with phylogenetic relationships, suggesting functional differentiation. Stress-responsive elements in P5CS promoters suggest involvement in abiotic stress. Expression analysis under salt stress revealed differential expressions of GhP5CS genes, with GhP5CS1 emerging as a potential key regulator. Virus-induced gene silencing confirmed the pivotal role of GhP5CS1 in cotton's salt stress response, as evidenced by increased salt sensitivity in the silenced plants. This study enhances our understanding of the functional diversity and roles of P5CS genes in cotton under stress conditions.

Chromosome-level genome assembly and annotation of largemouth bronze gudgeon (Coreius guichenoti)

Coreius guichenoti, mainly distributed in upstream regions of the Yangtze River China, is currently on the brink of extinction and listed as national secondary protected animal. In this study, we aimed to obtain the chromosome-level genome of C. guichenoti using PacBio and Hi-C techniques. According to the PacBio sequencing, C. guichenoti genome was successfully assembled to 1100.1 Mb size, with a Contig N50 size of 25.0 Mb, and containing 731.0 Mb of repeats. Hi-C sequencing data was utilized for chromosome assembly and 25 chromosome sequences were ultimately yielded, with a total length of 1076.8 Mb. Moreover, a total of 22,506 protein-coding genes were predicted with average intron length of 2293 bp. Evolutionary analysis and divergence time prediction revealed that C. guichenoti was closely related to C. heterodon and they phylogenetically diverged from common ancestor ~20.7 million years ago (Mya), following the separation of Cyprinidae at 28.3 Mya. In the future, the utilization of comparative genomics research is important in elucidating the molecular mechanisms of Ichthyophthirius disease and ensuring the conservation of biological resources.

Scaling up morphological differentiation of pangolin scales: Serial, ontogenetic and evolutionary variation.

Pangolins are the most heavily trafficked mammals in the world, largely because of the high demand for their keratinous scales from the traditional Chinese medicine market. While seizures of pangolin material are largely composed of isolated scales, efficient approaches to reach species-level identification are missing. This mostly originates from the lack of comparative studies on the shape of pangolin scales, resulting in knowledge gaps on the imbricated effects of serial, ontogenetic, and evolutionary variations. Using a large sample of micro-CT scan data, we performed the first quantitative 3D analysis of scale shape variation among the eight species of extant pangolins. Our geometric morphometric approach suggests that pangolins grossly share similar trends of serial variation, with scale shape and size being similarly distinct across the different body regions. Relative elongation is by far the main component of scale shape variation at the intraspecific level, although degree of asymmetry and shape of scale bed area also allow distinction among different body areas, especially in adults. At the evolutionary level, Phataginus is the most distinctive genus, with Pseudacteon tricuspis having the most elongated scales overall. Scales of the back (dorsum) appear to be the best discriminator between species, providing one of the few scale shape differences recorded between the genera Smutsia and Manis. Our results provide an unprecedented, upstream understanding of broad differentiation patterns across the scaled body of pangolins. Together with other yet-to-be-explored morphological traits (e.g., scale size, ornamentation, and thickness), scale shape could provide a valuable matrix of information for forensic applications.

Molecular and biochemical evolution of casbene-type diterpene and sesquiterpene biosynthesis in rice.

Casbene and neocembrene are casbene-type macrocyclic diterpenes; their derivatives play significant roles in plant defense and have pharmaceutical applications. We had previously characterized a casbene synthase, TERPENE SYNTHASE 28 (OsTPS28), in rice (Oryza sativa). However, the mechanism of neocembrene biosynthesis in rice remained unclear. In this study, we identified two genes of the TPS-a1 subfamily, OsTPS2 and OsTPS10, encoding a neocembrene synthase and sesquiterpene synthase, respectively, as supported by enzyme activity assays and determination of subcellular localization. Metabolic profiling of rice lines overexpressing either TPS confirmed the catalytic functions of OsTPS2 and OsTPS10, and suggested that OsTPS10 enhances resistance to rice bacterial blight. An evolutionary analysis revealed that OsTPS10 is conserved in monocots and first appeared in wild rice, whereas OsTPS2 and OsTPS28 sequentially evolved through gene duplication, transit peptide recruitment, and mutation of key amino acids such as H362R. In summary, this study not only deepens our understanding of the metabolic pathways and evolutionary history governing the biosynthesis of casbene-type diterpenoids in rice, representing parallel and divergent evolution within the gene family, and offers gene resources for the improvement of rice.

Unraveling TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR Transcription Factors in Safflower: A Blueprint for Stress Resilience and Metabolic Regulation.

Safflower (Carthamus tinctorius L.), a versatile medicinal and economic crop, harbors untapped genetic resources essential for stress resilience and metabolic regulation. The TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR (TCP) transcription factors, exclusive to plants, are pivotal in orchestrating growth, development, and stress responses, yet their roles in safflower remain unexplored. Here, we report the comprehensive identification and characterization of 26 safflower TCP genes (CtTCPs), categorized into Class I (PROLIFERATING CELL FACTOR, PCF) and Class II (CINCINNATA and TEOSINTE BRANCHED1/CYCLOIDEA, CIN and CYC/TB1) subfamilies. Comparative phylogenetics, conserved motif, and gene structure analyses revealed a high degree of evolutionary conservation and functional divergence within the gene family. Promoter analyses uncovered light-, hormone-, and stress-responsive cis-elements, underscoring their regulatory potential. Functional insights from qRT-PCR analyses demonstrated dynamic CtTCP expression under abiotic stresses, including abscisic acid (ABA), Methyl Jasmonate (MeJA), Cold, and ultraviolet radiation b (UV-B) treatments. Notably, ABA stress triggered a significant increase in flavonoid accumulation, correlated with the upregulation of key flavonoid biosynthesis genes and select CtTCPs. These findings illuminate the complex regulatory networks underlying safflower's abiotic stress responses and secondary metabolism, offering a molecular framework to enhance crop resilience and metabolic engineering for sustainable agriculture.