Biology subjects

Pucker, B.

Publications and source records attributed to Pucker, B..

At least 19 recordsLinked to original sources

bioRxiv2026

Comparative Genomics Reveals Genetic Factors Associated with Intraspecific Pigmentation Differences in Cassava

Cassava produces starch-filled tubers which are a crucial food source for many people, especially in Africa. This plant species is famous for drought tolerance and resilience, but changing conditions due to climate change require genetic improvements to ensure sustainable cassava cultivation. In this context, a comprehensive understanding of the genetic basis responsible for a variety of traits is beneficial. Here, we present insights into the anthocyanin metabolism, responsible for coloration and protection of cassava structures during stress. Intraspecific variation in the transcriptional control of the anthocyanin biosynthesis was characterized through comparative genomics and phylogenetic analyses. A member of the R2R3-MYB transcription factor family, orthologous to crucial activators of anthocyanin biosynthesis in other plant species, was identified as a genetic hotspot for pigment accumulation capacity between cultivars. Orthologs of this gene appeared absent from three cassava cultivars without pronounced anthocyanin pigmentation, while being present in cultivars known for anthocyanin formation. This discovery corroborates previous reports about the importance of transcriptional regulators in the intraspecific diversity of anthocyanin pigmentation.

plant biology↗

bioRxiv2026

Muscari commutatum genome sequences reveal genetic basis of flower pigmentation

Flower colour is an important component of floral signaling, yet the genetic basis of naturally occurring colour variation remains poorly understood in many non-model plant groups. Here, we compare genome sequences of a normally pigmented and a largely unpigmented individual of Muscari commutatum, a species typically characterized by very dark violet flowers. Both assemblies were approximately 3.5 Gbp in size, with contig N50 values exceeding 130 Mbp and BUSCO completeness of about 99%. Most structural genes of the flavonoid biosynthesis and the corresponding transcriptional regulators were identified in both plants. Strikingly, no flavanone 3-hydroxylase (F3H) gene was detected in the genome sequence of the unpigmented plant, and additional manual searches provided no evidence for a bona fide F3H copy. Comparison of the two genome sequences revealed a lack of local synteny around the F3H locus of the anthocyanin-pigmented plant, suggesting that F3H might have been lost in the unpigmented plant in association with a genomic rearrangement.

plant biology↗

bioRxiv2026

XpBrew and PanXpresso - automatic RNA-seq processing workflow and comprehensive collection of gene expression data

Rapid developments in sequencing technologies have reduced the costs of transcriptomic experiments and resulted in a plethora of publicly available RNA-seq datasets. This is a valuable resource that can be harnessed to obtain novel biological insights through data upcycling. In this wake, we introduce XpBrew, an end-to-end Python workflow that was applied to generate PanXpresso, a comprehensive collection of gene expression datasets covering the taxonomic breadth of plants, animals, fungi, bacteria and archaea. XpBrew (https://github.com/PuckerLab/XpBrew) and PanXpresso (https://doi.org/10.60507/FK2/OBIGQH) are freely available.

bioinformatics↗

bioRxiv2026

Anthocyanin biosynthesis gene activation in nitrogen deprived Utricularia gibba L. under light or darkness

Utricularia gibba L. is an aquatic carnivorous plant with a diverse set of capabilities. Reddening of traps frequently occurs in old in vitro cultures. While anthocyanins are often responsible for red coloration in plants, not every plant turns red. Stress factors like high light or excess sucrose have previously been shown to induce the formation of anthocyanins. Here, we hypothesized the red trap formation to be dependent on nutrient deprivation and tested nitrogen deprivation. The results suggest, that only in combination with light, nitrogen deficiency leads to the activation of the complete anthocyanin biosynthesis pathway and visible red coloration. However, in darkness, anthocyanin biosynthesis appears generally less active compared to light conditions and expression of most anthocyanin biosynthesis genes is not significantly upregulated under nitrogen deficiency.

plant biology↗

bioRxiv2026

Valeriana officinalis genome sequence reveals candidate genes for valerenic acid biosynthesis and flavonoid metabolism

Valeriana officinalis is the scientific name for valerian, a plant known for producing valerenic acid, a compound with anxiolytic properties. Anxiety disorders represent a significant global health crisis, impacting everyday lives. As the global demand for natural, non-synthetic anxiety treatments rises, V. officinalis has emerged as a promising, yet underutilized, medicinal resource. Understanding its genome is the first step toward unraveling the biosynthetic genes underlying valerenic acid production, facilitating further research into its production. Here, we report the first genome sequence of valerian, with an assembly size of 3.3 Gbp and an N50 of 110.8 Mbp, and its corresponding annotation with 96.6% completeness, providing a foundational resource for studying the genetic basis of specialized metabolism in valerian. The value of this genome sequence for discoveries in specialized metabolism is demonstrated by the identification of the flavonoid biosynthesis gene repertoire and the selection of strong candidate genes for valerenic acid biosynthesis. This genome sequence holds the potential to support future functional studies aimed at elucidating the regulation of medically relevant metabolite pathways in V. officinalis.

genomics↗

bioRxiv2026

Genome sequence of the medicinal plant Urtica dioica reveals the genetic basis of the flavonoid metabolism

BackgroundUrtica dioica, also known as stinging nettle, is a widespread plant that can indicate high nitrogen availability in the soil. It is probably best known for the pain caused by touching it. U. dioica is also recognized as a medicinal plant with reports claiming applicability against numerous diseases. ResultsA highly continuous genome sequence was constructed based on nanopore long read sequencing data. The total assembly size is 1.1 Gbp with an N50 of 40.7 Mbp. RNA-seq data and hints from other species were integrated to produce a high quality annotation of the protein encoding genes. This genomic resource enabled the identification of genes involved in the flavonoid biosynthesis. A particular focus was on anthocyanin biosynthesis genes as these are crucial for high light and nitrogen deprivation stress response, which is revealed by redding of the leaves. ConclusionThis genomic resource provides the basis for future studies unraveling the biosynthesis pathways underlying various medically important compounds produced by stinging nettles.

plant biology↗

bioRxiv2026

Rubus armeniacus genome sequence reveals the secrets of blackberry anthocyanin biosynthesis

Here, we present a comprehensive multiomics analysis of anthocyanin biosynthesis in Rubus armeniacus, known for its dark fruits. A phased genome sequence of the tetraploid blackberry was generated, achieving an N50 of 34 Mb with an assembly size of 1.2 Gbp based on Oxford Nanopore Technology sequencing (ONT). The BUSCO score for the total assembly shows a high completeness of 99.1%. The assembly was separated into 4 pseudohaplophases, with the pseudohaplophase A representing the R. armeniacus genome in 7 chromosome scale contigs, with an N50 of 46 Mbp and 98.8% conserved BUSCO genes. A total of 118,183 protein coding genes were annotated within the genome assembly and all relevant genes encoding enzymes and transcriptional regulators of the anthocyanin biosynthesis pathway were identified within each pseudohaplophase. To further understand the underlying cause of dark pigmentation, the gene expression was analysed during different stages of berry development revealing a strong induction of anthocyanin biosynthesis genes including the anthocyanin activating subgroup 6 MYB transcriptions during the berry ripening process. Further, a quantification of cyanidin-3-O-glucoside in methanolic berry extract, utilizing a UHPLC-HRAM-MS analysis, revealed an approximately 500-fold increase of cyanidin-3-O-glucoside from green to black fruit, indicating that dark pigmentation in R. armeniacus results from high anthocyanin accumulation. Significance statementThis study provides a multiomics analysis of the dark pigmentation of Rubus armeniacus, including a high quality phased assembly and an in-depth analysis of the anthocyanin biosynthesis pathway. A transcriptional and metabolomic analysis revealed that dark berry pigmentation is caused by a high accumulation of cyanidin-3-O-glucoside during fruit ripening.

genomics↗

bioRxiv2026

A MaBBX21-MaGCN5 Histone Acetyltransferase Module Regulates Flavonoid Biosynthesis to Improve Heat and UV B Stress Tolerance in Banana (Musa acuminata)

Flavonoids are important specialized metabolites that contribute to plant pigmentation, stress adaptation, and nutritional value. In banana, a major global staple crop, their accumulation is highly tissue-specific, with very low levels in the edible pulp, and the mechanisms underlying this spatial distribution remain unclear. Here, integrative transcriptomic and metabolomic analyses across vegetative and reproductive tissues reveal that light-responsive regulatory networks control tissue-specific flavonoid biosynthesis. We identified a B-box transcription factor MaBBX21 as a key positive regulator of flavonoid biosynthesis. Its overexpression enhances flavonoid accumulation, whereas knockdown leads to a reduction in flavonoid levels in banana. Mechanistically, MaBBX21 interacts with MaHY5 and directly activates anthocyanin biosynthesis genes (MaDFR2 and MaANS). It also regulates metabolic flux by binding to the MaWRKY23 promoter, promoting flavonol biosynthesis through activation of MaFLS1 while partially repressing the anthocyanin branch. In addition, MaBBX21 introduces an epigenetic layer by activating the histone acetyltransferase MaGCN5, increasing H3K9 acetylation at target promoters, including MaDFR2, MaANS, and MaBBX21 itself, thereby forming a positive feedback loop. Functionally, MaBBX21 overexpression enhances flavonoid accumulation, ROS scavenging, and tolerance to heat and UV-B stress, whereas knockdown lines show reduced metabolite levels and increased stress sensitivity. Collectively, these results define a MaBBX21-MaWRKY23/MaGCN5 regulatory axis that integrates transcriptional regulation, and chromatin modification to control flavonoid biosynthesis, providing a foundation for improving nutritional quality and stress resilience in banana.

plant biology↗

bioRxiv2026

Lack of evidence for anthocyanins contributing to pigmentation of Chenopodium quinoa

While most plant lineages are pigmented by anthocyanins, several families in the Caryophyllales represent a major exception by showing a replacement of anthocyanin pigmentation by betalain pigmentation. The mutual exclusion of anthocyanins and betalains at the family level has been well established for over 50 years and has been mechanistically explained. Chenopodiaceae are a betalain-pigmented lineage lacking a key anthocyanin biosynthesis gene and lacking the key activating transcription factor of the anthocyanin biosynthesis. A publication by Zhang et al., 2024 claims that anthocyanins would be responsible for the red pigmentation in leaves of Chenopodium quinoa. Here, we assessed this study and reanalyzed the RNA-seq datasets generated in this study to demonstrate that there is no evidence for anthocyanin biosynthesis, but activity of the betalain and carotenoid biosynthesis could explain the observed pigmentation of quinoa leaves.

plant biology↗

bioRxiv2026

NanoPlasmiQC: Full plasmid sequencing with ONT long-reads and automatic data analysis

Long-read sequencing has shown a rapid technological development during the last years. It has been established as the standard method for the sequencing of plant genomes and has also gained importance for full plasmid sequencing. As Sanger sequencing has a limited read length of about 1 kb, long read sequencing offers a great advantage, as the full plasmid can be sequenced in one read. Here, we present a cost-effective workflow to sequence full plasmids and compare the results against an expectation. The per plasmid cost of this workflow is determined by the number of plasmids investigated simultaneously, but can be lower than the price of a single Sanger sequencing reaction. We developed a workflow for automatic data processing, which allows us to complete sequencing and data analysis within a day.

genomics↗

bioRxiv2026

A hidden T-DNA-linked inversion-duplication causes a pronounced light-dependent phenotype in Arabidopsis

T-DNA insertion mutants are widely used to disrupt genes and infer their functions, yet the insertions can also trigger unintended genomic changes that confound phenotypic interpretation. Here, we used T-DNA insertion mutants affecting the major mitochondrial malate dehydrogenase (MDH1) and the heterodimeric NAD-dependent malic enzymes (ME1 and ME2) to examine their functional coordination across photoperiods and irradiance regimes. Under short days, especially at low light intensity, mdh1xme2 mutants were markedly smaller than wild type and, unexpectedly, than the mdh1xme1xme2 triple mutant, and they showed a more pronounced reduction in photosynthetic capacity. ME1 was undetectable in mdh1xme2, implying that the double and triple mutants effectively lack heterodimeric ME and should therefore behave similarly, contrary to what we observed. Whole-genome analysis resolved this discrepancy by revealing that the MDH1 T-DNA insertion in mdh1xme2 is accompanied by a major rearrangement, a 137-kbp duplication downstream of the insertion site, which was absent in the mdh1xme1xme2 triple mutant. This duplication increased gene dosage and elevated transcript abundance across the duplicated interval, while proteomics detected 5 of the 38 encoded proteins, including PEPC1. mdh1xme2 accumulated oxaloacetate-derived amino acids and displayed an altered carbon/nitrogen balance, making PEPC1 a plausible contributor to the exacerbated mdh1xme2 phenotype. Together, our data indicate that a T-DNA-linked structural variant can amplify expression of dozens of genes and intensify phenotypes at specific conditions, thereby affecting the interpretation of genotype-phenotype relationships. Because Agrobacterium-mediated DNA transfer also underpins many genome-editing workflows, our findings argue that structural validation around insertion/editing loci should be considered essential when interpreting T-DNA-derived plant lines.

plant biology↗

bioRxiv2026

Genome sequence of Tacca chantrieri reveals the genetic basis of floral pigmentation

Tacca chantrieri, black bat flower, has showy flowers often appearing almost black. Here, we present the genome sequence and corresponding annotation to identify the genetic basis of the pigmentation. Candidate genes associated with the anthocyanin biosynthesis were identified based on this genome sequence and investigated with respect to their properties. The best dihydroflavonol 4-reductase (DFR) candidate, which harbours all amino acid residues believed to be required for DFR activity, shows a threonine in the substrate preference determining position where most characterized DFRs display asparagine or aspartate. This amino acid residue appears to be frequent in the Dioscoreaceae family as a comprehensive investigation revealed.

plant biology↗

bioRxiv2026

Mitochondrial malate metabolism acts as a control hub for photosynthesis and carbon-nitrogen balance in Arabidopsis

Malate is a central metabolite in plant energy metabolism and biosynthesis and serves as a major carrier of carbon and reducing equivalents between chloroplasts, the cytosol, and mitochondria. However, how individual malate-converting systems contribute to physiology in specific subcellular compartments remains incompletely understood. Here, we investigated the impact of combined loss of mitochondrial malate dehydrogenase (MDH) and NAD-dependent malic enzyme (NAD-ME) activity in Arabidopsis thaliana by integrating reverse genetics, physiological analyses, transcriptomics, quantitative proteomics, and metabolite profiling. Specifically, we generated triple mutants (mdh1xme1xme2) lacking the predominant mitochondrial isoform MDH1 together with both NAD-ME subunits, thereby reducing overall mitochondrial malate conversion capacity. By growing plants under contrasting photoperiod and irradiance regimes to vary photosynthetic demand on malate-linked fluxes, we uncovered a conditional phenotype that was most pronounced under short-day/low-light conditions. Under these conditions, mdh1xme1xme2 exhibited impaired growth and photosynthetic performance, accompanied by cytosolic redox imbalance and altered chloroplast ultrastructure. Transcriptomic profiling revealed that low light unmasks a dawn-phase bottleneck in establishing photosynthetic and redox homeostasis. Consistent with this, the low-light plastid proteome revealed a reallocation away from chloroplast translation and photosynthetic capacity toward proteome maintenance, photoprotection/repair, and iron/ROS management, consistent with a protective acclimation state that nevertheless constrains carbon gain under energy limitation. Low light also triggered C/N imbalance and ammonium accumulation in the mutants. In contrast, increasing irradiance or extending the photoperiod largely alleviated these defects. Together, our results identified mitochondrial malate conversion capacity as a key control point coupling respiratory energy supply and redox homeostasis to photosynthetic metabolism when photosynthetic energy input is limiting.

plant biology↗

bioRxiv2026

Begonia manicata genome sequence reveals genetic basis underlying ornamental pigmentation

Plant genome sequences provide access to the gene repertoire of a species. This facilitates basic research, biotechnological processes, or horticultural applications. Here, we present the genome sequence of Begonia manicata and unravel the genes underlying the pigmentation of red structures emerging from its leaves and stems. Structural genes of the anthocyanin biosynthesis and corresponding regulatory genes were discovered to be upregulated in these red structures suggesting that the pigmentation is caused by the accumulation of anthocyanins. Our work provides a resource for future studies on pigmentation of Begoniaceae.

plant biology↗

bioRxiv2026

Pathogen-induced red pigmentation uncovers a conserved floral defense in Asteraceae

Flower colour is predominantly selected for visual appeal and pollinator-driven evolution, however its role in plant-pathogen interaction remains unclear. Here, we investigated the relationship between anthocyanin accumulation and resistance to the necrotrophic pathogen Botrytis cinerea in Chrysanthemum morifolium. Using a unique set of spontaneous colour mutants sharing a common genetic background, we demonstrate that higher anthocyanin levels are associated with enhanced resistance to B. cinerea. Furthermore, we also observed pathogen induced anthocyanin accumulation at pathogen penetration sites in the white cultivars. Transcriptomics and metabolomics analyses revealed shift in metabolic pathways from flavonols towards flavone and anthocyanin biosynthesis during infections. A broader survey of taxonomically distant flowering species identified pathogen-induced red pigmentation is largely restricted to the members of the Asteraceae family. Notably, we observed divergence between wild and domesticated species, where the wild species retained the inducible pigmentation response and exhibited resistant to B. cinerea, whereas the cultivated species showed loss of this response, correlating with its increased susceptibility. Overall, our findings indicate that breeding for ornamental traits may have inadvertently selected against floral immunity.

plant biology↗

bioRxiv2026

Phylogenetic based dissection of eukaryotic Mo-insertase functionality: From mechanism to complex assembly

Molybdenum cofactor (Moco) biosynthesis is vitally important for all organisms, yet the domain organization of the eukaryotic molybdenum insertase (Mo-insertase) remains enigmatic. We combine extensive phylogenetic reconstructions, sequence analysis and structural modeling in order to uncover evolutionary and functional principles of eukaryotic Mo-insertases. We note, that the vast majority of plant, fungi and animal species evolved fused E- and G-domains, yet the orientation of both domains in the fusion proteins differs among different eukaryotic lineages. Despite the divergent domain arrangements amongst eukaryotic Mo-insertases the E-domain active site is well conserved, with very few tolerated substitutions in >1,000 sequences. Among the Mo-insertases from different eukaryotic species, vertebrate gephyrin is the only Mo-insertase with a dual function as - next to its metabolic function - it scaffolds inhibitory neurotransmitter receptors in the post synapsis. Gephyrin is surprisingly high conserved, including surface patches not directly involved in catalysis and receptor clustering. This profile suggests additional, as yet uncharacterized, functional constraints on gephyrins evolution. Together, our results reveal how eukaryotic Mo-insertases combine evolutionary domain organization plasticity with stringent active site conservation and recognize the evolutionary constraint on gephyri[n]s surface conservation to be extreme, likely due to its mutual metabolic and neuronal function.

bioinformatics↗

bioRxiv2025

DupyliCate - mining, classifying, and characterizing gene duplications

Paralogs, copies of a gene, form an important basis for novelty during evolution. Analysis of such gene duplications is important to understand the emergence of novel traits during evolution. DupyliCate is a Python tool that has been developed for this purpose. With the ability to process multiple datasets concurrently, flexible features, and parameters to set species-specific thresholds, DupyliCate offers a high-throughput method for gene copy identification and analysis. The different available parameters and modes are explored in detail based on the Arabidopsis thaliana datasets. Proof of concept for the tool is presented by characterizing well known duplications in different plants, and its broad applicability is demonstrated by running it on diverse datasets including complex plant genome sequences with high heterozygosity. Further, two case studies involving the evolution of flavonol synthase (FLS) genes in Brassicales, and the evolution of flavonol synthesis regulating myeloblastosis (MYB) transcription factors- MYB12 and MYB111 across a large number of plant species, are presented as exemplar use cases. The tools applicability beyond plants is demonstrated on Escherichia coli, Saccharomyces cerevisiae, and Caenorhabditis elegans datasets. DupyliCate is available at: https://github.com/ShakNat/DupyliCate.

bioinformatics↗

bioRxiv2025

Large-scale Phylogenomics Reveals Systematic Loss of Anthocyanin Biosynthesis Genes at the Family Level in Cucurbitaceae

O_LIPlant pigmentation secrets are among the oldest interests of plant scientists, with pigments such as chlorophyll, carotenoids, flavonoids, and betalains contributing to the diversity of hues in higher plants. Anthocyanins, a class of flavonoids responsible for vibrant shades of pink, red, and blue pigmentation, are almost ubiquitous in angiosperms but are replaced by betalains in some families in the order Caryophyllales. C_LIO_LIWe investigated anthocyanin pigmentation in Cucurbitaceae, one of the largest fruit and vegetable families, characterised by white and yellow flowers and red, orange, and green fruits predominantly pigmented by carotenoids. Using a comprehensive collection of 258 datasets representing 183 unique species across all 15 tribes of Cucurbitaceae, along with a phylogenomics approach, we observed a systematic absence of genes involved in anthocyanin and proanthocyanidin biosynthesis. Absence of the structural genes DFR, ANS, arGST, LAR, and ANR, along with the anthocyanin-related regulatory MYB genes, was consistently confirmed by synteny and phylogenetic analysis. C_LIO_LIThese results suggest that anthocyanin loss in angiosperms is more common than previously assumed. In light of this new discovery, we propose a stepwise loss of anthocyanin pigmentation in Cucurbitaceae, likely accompanied by a partial functional replacement with carotenoid pigmentation. C_LI

plant biology↗