Author: slquinlan

Tree Physiol. 2025 Jul 11:tpaf081. doi: 10.1093/treephys/tpaf081. Online ahead of print.

ABSTRACT

The success of CRISPR genome editing studies depends critically on the precision of guide RNA (gRNA) design. Sequence polymorphisms in outcrossing tree species pose design hazards that can render CRISPR genome editing ineffective. Despite recent advances in tree genome sequencing with haplotype resolution, sequence polymorphism information remains largely inaccessible to various functional genomics research efforts. The Populus VariantDB v3.2 addresses these challenges by providing a user-friendly search engine to query sequence polymorphisms of heterozygous genomes. The database accepts short sequences, such as gRNAs and primers, as input for searching against multiple poplar genomes, including hybrids, with customizable parameters. We provide examples to showcase the utilities of VariantDB in improving the precision of gRNA or primer design. The platform-agnostic nature of the probe search design makes Populus VariantDB v3.2 a versatile tool for the rapidly evolving CRISPR field and other sequence-sensitive functional genomics applications. The database schema is expandable and can accommodate additional tree genomes to broaden its user base.

PMID:40643191 | DOI:10.1093/treephys/tpaf081

Tree Physiol. 2025 Jul 11:tpaf080. doi: 10.1093/treephys/tpaf080. Online ahead of print.

ABSTRACT

Sulfate-proton co-transporters (SULTRs) mediate sulfate uptake, transport, storage, and assimilation in plants. The SULTR family has historically been classified into four groups (SULTR1-SULTR4), with well-characterized roles for SULTR groups 1, 2, and 4. However, the functions of the large and diverse SULTR3 group remain poorly understood. Here, we present an updated phylogenetic analysis of SULTRs across angiosperms, including multiple early-divergent lineages. Our results suggest that the enigmatic SULTR3 group comprises four distinct subfamilies that predate the emergence of angiosperms, providing a basis for reclassifying the SULTR family into seven subfamilies. This expanded classification is supported by subfamily-specific gene structures and amino acid substitutions in the substrate-binding pocket. Structural modeling identified three serine residues uniquely lining the substrate-binding pocket of SULTR3.4, enabling three hydrogen bonds with the phosphate ion. The data support the proposed neofunctionalization of this subfamily for phosphate allocation within vascular tissues. Transcriptome analysis of Populus tremula × alba revealed divergent tissue expression preferences among SULTR subfamilies and between genome duplicates. We observed partitioned expression in vascular tissues among the four SULTR3 subfamilies, with PtaSULTR3.4a and PtaSULTR3.2a preferentially expressed in primary and secondary xylem, respectively. Gene coexpression analysis revealed coordinated expression of PtaSULTR3.4a with genes involved in phosphate starvation responses and nutrient transport, consistent with a potential role in phosphate homeostasis. In contrast, PtaSULTR3.2a was strongly coexpressed with lignification and one-carbon metabolism genes and their upstream transcription regulators. PtaSULTR3.2a belongs to a eudicot-specific branch of the SULTR3.1 subfamily found only in perennial species, suggesting a specialized role in lignifying tissues. Together, our findings provide a refined phylogenetic framework for the SULTR family and suggest that the expanded SULTR3 subfamilies have undergone neofunctionalization during the evolution of vascular and perennial plants.

PMID:40643194 | DOI:10.1093/treephys/tpaf080

J Gen Virol. 2025 Jul;106(7). doi: 10.1099/jgv.0.002119.

ABSTRACT

Orthotospovirus tomatomaculae [tomato spotted wilt virus (TSWV)] is a major pathogen in horticultural and row crops worldwide including the USA. In this study, tomato spotted wilt disease incidence was monitored in Arachis hypogaea (peanut; year 1990 to 2024) and Nicotiana tabacum (tobacco; year 2000 to 2024) in commercial farmers’ fields in the Southeastern USA. Furthermore, nucleocapsid (N), nonstructural movement (NSm) and nonstructural silencing suppressor (NSs) protein gene sequences of TSWV global populations from North America, South America, Europe, Asia-Pacific, Africa and Australia were compared with local US population and analysed to understand the genetic variability in the virus genome. In our study, full-length sequences of 94 N, 111 NSm and 78 NSs genes were amplified from TSWV-infected A. hypogaea (peanut), Capsicum annuum (pepper), N. tabacum (tobacco) and Solanum lycopersicum (tomato). nt-based phylogenetic analysis of N, NSm and NSs genes correlated with the geographical location of the TSWV isolates, with notably higher substitution rates in the population of recent years. In addition, the least genetic variability was observed in the N gene of the local population upon comparison with other global TSWV population. The neutrality test of TSWV suggested a non-neutral evolution of the virus genome. Low variation among the selected genes might be attributed to strong purifying selection pressure in the populations. Furthermore, estimation of selection pressure (dN/dS) on small (S) segment-encoded N protein and nonstructural protein showed higher purifying selection than the movement protein encoded by the medium (M) segment of the TSWV isolates. Single-likelihood ancestor counting suggested an overall negative selection pressure on several codons of the selected genes, which indicated that natural selection and population bottleneck events might have influenced the evolution of TSWV. Our study also deciphered high gene flow and low genetic differentiation amid the different TSWV population sets. Additionally, BEAST analysis of TSWV N gene sequences from GA predicted the most common recent ancestor existed ~25 years ago. This data was further correlated with disease incidence data from peanut and tobacco crops obtained in the last three decades. These findings suggest the intermixing of TSWV isolates between peanut, pepper, tobacco and tomato crops, while the virus genome has undergone strong purifying selection.

PMID:40622855 | DOI:10.1099/jgv.0.002119

Plant J. 2025 Jul;123(1):e70304. doi: 10.1111/tpj.70304.

ABSTRACT

Targeted demethylation by DNA glycosylases (DNGs) results in differential methylation between parental alleles in the endosperm, which drives imprinted expression. Here, we performed RNA sequencing on endosperm derived from DNG mutant mdr1 and wild-type (WT) endosperm. Consistent with the role of DNA methylation in gene silencing, we find 108 genes and 96 TEs differentially expressed (DE) transcripts that lost expression in the hypermethylated mdr1 mutant. Compared with other endosperm transcripts, the mdr1 targets are enriched for TEs (particularly Helitrons), and DE genes are depleted for both core genes and GO term assignments, suggesting that the majority of DE transcripts are TEs and pseudo-genes. By comparing DE genes to imprinting calls from prior studies, we find that the majority of DE genes have maternally biased expression, and approximately half of all maternally expressed genes (MEGs) are DE in this study. In contrast, no paternally expressed genes (PEGs) are DE. DNG-dependent imprinted genes are distinguished by maternal demethylation and expression primarily in the endosperm, so we also performed Enzymatic Methyl-seq on hybrids to identify maternal demethylation and utilized a W22 gene expression atlas to identify genes expressed primarily in the endosperm. Overall, approximately ⅔ of all MEGs show evidence of regulation by DNGs. Taken together, this study solidifies the role of MDR1 in the regulation of maternally expressed, imprinted genes and TEs and identifies subsets of genes with DNG-independent imprinting regulation.

PMID:40587880 | DOI:10.1111/tpj.70304

BioTech (Basel). 2025 Jun 12;14(2):48. doi: 10.3390/biotech14020048.

ABSTRACT

Interspecific and intersectional crosses have introduced valuable genetic traits for blueberry (Vaccinium sect. Cyanococcus) cultivar improvement. Introgression from Vaccinium species at the diploid, tetraploid, and hexaploid levels has been found in cultivated blueberries. Continued efforts to integrate wild blueberry genetic resources into blueberry breeding are essential to broaden the genetic diversity of cultivated blueberries. However, performing heteroploid crosses among Vaccinium species is challenging. Polyploid induction through tissue culture has been useful in bridging ploidy barriers. Mixoploid or chimeric shoots often are produced, along with solid polyploid mutants. These chimeras are mostly discarded because of their genome instability and the difficulty in identifying periclinal mutants carrying germline mutations. Since induced polyploidy in blueberries often results in a low frequency of solid mutant lines, it is important to recover solid polyploids through chimera dissociation. In this study, two vegetative propagation methods, i.e., axillary and adventitious shoot induction, were evaluated for their efficiency in chimera dissociation. Significantly higher rates of chimera dissociation were found in adventitious shoot induction compared to axillary shoot induction. Approximately 89% and 82% of the adventitious shoots induced from mixoploid lines 145.11 and 169.40 were solid polyploids, respectively, whereas only 25% and 53% of solid polyploids were recovered through axillary shoot induction in these lines. Effective chimera dissociation provides useful and stable genetic materials to enhance blueberry breeding.

PMID:40558397 | DOI:10.3390/biotech14020048

bioRxiv [Preprint]. 2025 May 28:2025.05.23.655667. doi: 10.1101/2025.05.23.655667.

ABSTRACT

Repairing a damaged body part is critical for the survival of any organism. In plants, tissue damage induces rapid responses that activate defense, regeneration and wound healing. While early wound signaling mediated by phytohormones, electrical signals and reactive oxygen species is well-characterized, the mechanisms governing the final stages of wound healing remain poorly understood. Here, we show that wounding in Arabidopsis leaves induces localized cooling, likely due to evaporative water loss, accompanied by the activation of cold-responsive genes. The subsequent disappearance of localized cooling and deactivation of cold-responsive genes serve as a quantitative marker of wound healing. Based on these observations, we developed a workflow by leveraging computer vision and deep learning to monitor the dynamics of wound healing. We found that CBFs transcription factors relay injury-induced cooling signal to wound healing. Thus, our work advances our understanding of tissue repair and provides a tool to quantify wound healing in plants.

PMID:40502075 | PMC:PMC12154623 | DOI:10.1101/2025.05.23.655667

Phytopathology. 2025 Jun 22. doi: 10.1094/PHYTO-03-25-0093-R. Online ahead of print.

ABSTRACT

Phony peach disease (PPD), caused by Xylella fastidiosa subsp. multiplex (Xfm), poses a significant threat to commercial peach orchards in Georgia. Early and accurate detection is essential for effective disease management, yet visual assessment remains the primary approach for diagnosing PPD symptoms due to the high cost and logistical challenges of qPCR-based detection of Xfm. We evaluated the accuracy of visual PPD assessment and examined the factors influencing rater performance, symptom reliability, and optimal survey deployment strategies with CART/Random Forest analyses and simulations. Internode length was the most reliable symptom for PPD identification in two peach cultivars, consistently outperforming other physical traits such as canopy flatness and shape. Primer pair C06Xf-bamA had the greatest relative sensitivity, making it the preferred choice for qPCR confirmation. Principal component analysis suggested that rater experience significantly improved agreement with qPCR results and repeated assessments of the same orchards further enhanced consistency for raters. Simulations results suggested that deploying two experienced raters may provide the highest detection diagnostic accuracy for survey purposes, particularly when qPCR-based pathogen detection is unavailable. Last, PPD-affected trees, through PCR verification and visual identification, exhibited higher mortality rates than Xfm-negative trees, reinforcing the need for early detection and removal to limit disease spread. These findings underscore the importance of strategic rater deployment, targeted symptom selection, and integrating molecular diagnostics when feasible.

PMID:40544457 | DOI:10.1094/PHYTO-03-25-0093-R

bioRxiv [Preprint]. 2025 May 27:2025.05.22.655462. doi: 10.1101/2025.05.22.655462.

ABSTRACT

Meiotic drive elements are regions of the genome that are transmitted to progeny at frequencies that exceed Mendelian expectations, often to the detriment of the organism. In maize there are three prevalent chromosomal drive elements known as Abnormal chromosome 10 (Ab10), K10L2, and the B chromosome. There has been much speculation about how these drivers might interact with each other and the environment in traditional maize landraces and their teosinte ancestors. Here we used genotype-by-sequencing data to score more than 10,000 maize and teosinte lines for the presence or absence of each driver. Less than ~0.5% of modern inbred lines carry chromosomal drivers. Among individuals from 5331 open-pollinated landraces, 6.32% carried Ab10, 5.16% carried K10L2, and 12.28% carried at least one B chromosome. Using a GWAS approach we identified unlinked loci that associate with the presence or absence of the selfish genetic elements. Many genetic modifiers are positively associated with the drivers, suggesting that there may have been selection for alleles that ameliorate their negative fitness consequences. We then assessed the contributions of population structure, associated loci, and the environment on the distribution of each chromosomal driver. There was no significant relationship between any chromosomal driver and altitude, contrary to conclusions based on smaller studies. Our data suggest that the distribution of the major chromosomal drivers is primarily influenced by neutral processes and the deleterious fitness consequences of the drivers themselves. While each driver has a unique relationship to genetic background and the environment, they are largely unconstrained by either.

PMID:40501570 | PMC:PMC12154789 | DOI:10.1101/2025.05.22.655462

Mol Plant Pathol. 2025 Jun;26(6):e70107. doi: 10.1111/mpp.70107.

ABSTRACT

Acidovorax citrulli, the causal pathogen of bacterial fruit blotch of cucurbits, relies on a functional type III secretion system (T3SS) for pathogenicity. Two-component systems (TCSs) are primary signal transduction mechanisms for bacteria to detect and adapt to various environmental conditions. However, the role of TCS on regulating T3SS and other virulence factors in response to environmental stimuli is still poorly understood in A. citrulli. Here, we report the identification of a conserved TCS, OmpR/EnvZ, involved in hypersensitive response (HR) induction in Nicotiana benthamiana by screening a transposon-insertion library in the group II strain xjL12 of A. citrulli. Transcription analysis confirmed that OmpR_Ac/EnvZ_Ac was upregulated in response to elevated osmotic pressure, low and high pH conditions, and host environment. Deletions of envZ_Ac, ompR_Ac, or both envZ_Ac and ompR_Ac in A. citrulli attenuated virulence to melon seedlings and mature leaf tissues, and delayed HR in N. benthamiana. OmpR_Ac was activated by EnvZ_Ac and directly bound to the promoter region of hrpG, a major regulator of T3SS. This binding activated hrpG transcription and promoted T3SS assembly in T3SS-inducing medium, XVM2. Additionally, the OmpR_Ac/EnvZ_Ac mutants of A. citrulli displayed reduced swimming motility due to impaired flagella formation, but also had enhanced biofilm formation and exopolysaccharide production. OmpR_Ac/EnvZ_Ac regulation of these virulence factors in A. citrulli depended on its own conserved phosphorylation sites. This work illuminates a signalling pathway for regulating the T3SS and provides insights into the OmpR/EnvZ-mediated virulence regulatory network in A. citrulli.

PMID:40524436 | DOI:10.1111/mpp.70107

mSphere. 2025 Jun 17:e0002325. doi: 10.1128/msphere.00023-25. Online ahead of print.

ABSTRACT

Allicin tolerance (alt) clusters in phytopathogenic bacteria, which provide resistance to thiosulfinates like allicin, are challenging to find using conventional approaches due to their varied architecture and the paradox of being vertically maintained within genera despite likely being horizontally transferred. This results in significant sequential diversity that further complicates their identification. Natural language processing (NLP), like techniques such as those used in DeepBGC, offers a promising solution by treating gene clusters like a language, allowing for identifying and collecting gene clusters based on patterns and relationships within the sequences. We curated and validated alt-like clusters in Pantoea ananatis 97-1R, Burkholderia gladioli pv. gladioli FDAARGOS 389, and Pseudomonas syringae pv. tomato DC3000. Leveraging sequences from the RefSeq bacterial database, we conducted comparative analyses of gene synteny, gene/protein sequences, protein structures, and predicted protein interactions. This approach enabled the discovery of several novel alt-like clusters previously undetectable by other methods, which were further validated experimentally. Our work highlights the effectiveness of NLP-like techniques for identifying underrepresented gene clusters and expands our understanding of the diversity and utility of alt-like clusters in diverse bacterial genera. This work demonstrates the potential of these techniques to simplify the identification process and enhance the applicability of biological data in real-world scenarios.IMPORTANCEThiosulfinates, like allicin, are potent antifeedants and antimicrobials produced by Allium species and pose a challenge for phytopathogenic bacteria. Phytopathogenic bacteria have been shown to utilize an allicin tolerance (alt) gene cluster to circumvent this host response, leading to economically significant yield losses. Due to the complexity of mining these clusters, we applied techniques akin to natural language processing to analyze Pfam domains and gene proximity. This approach led to the identification of novel alt-like gene clusters, showcasing the potential of artificial intelligence to reveal elusive and underrepresented genetic clusters and enhance our understanding of their diversity and role across various bacterial genera.

PMID:40525872 | DOI:10.1128/msphere.00023-25