Biology subjects

Scanlon, M. J.

Publications and source records attributed to Scanlon, M. J..

14 recordsLinked to original sources

bioRxiv2026

Cloning and characterization of a novel maize leaf area modifier and its effects across elite germplasm

Plant architecture is a crucial component of maize productivity. Tailoring architectural component traits like leaf area and angle can increase productivity by promoting deeper light penetration into the canopy and better resource utilization. Novel genetic variants can increase the rate of gain for optimized plant architecture. Here, we map a moderate-effect mutation denoted reduced leaf area1 (rdla1) to the RAGGED5 (RGD5) locus and characterize it as a transposon insertion allele. Mutant leaf area reductions were most extreme in mid-upper canopy positions. Photosynthetic gas exchange rates were not significantly impacted in rdla1 relative to wild-type, indicating that mutant leaf structure, but not function, is altered. Functional annotations of RDLA1 were supported by metabolite profiles suggesting a role in cuticular wax biosynthesis. Introgression of the rdla1 allele into 27 commercially relevant genetic backgrounds identified differences in effect size across genotypes, revealing modifier effects that could serve as targets for modulating plant architecture.

plant biology↗

bioRxiv2024

Integrative multi-omic analysis identifies genes associated with cuticular wax biogenesis in adult maize leaves

Studying the genetic basis of leaf wax composition and its correlation with leaf cuticular conductance (gc) is crucial for improving crop water-use efficiency. The leaf cuticle, which comprises a cutin matrix and various waxes, functions as an extracellular hydrophobic layer, protecting against water loss upon stomatal closure. To address the limited understanding of genes associated with the natural variation of leaf cuticular waxes and their connection to gc, we conducted statistical genetic analyses using leaf transcriptomic, metabolomic, and physiological data sets collected from a maize (Zea mays L.) panel of [~]300 inbred lines. Through a random forest analysis with 60 cuticular wax traits, it was shown that high molecular weight wax esters play an important role in predicting gc. Integrating results from genome-wide and transcriptome-wide studies (GWAS and TWAS) via a Fishers combined test revealed 231 candidate genes detected by all three association tests. Among these, 11 genes exhibit known or predicted roles in cuticle-related processes. Throughout the genome, multiple hotspots consisting of GWAS signals for several traits from one or more wax classes were discovered, identifying four additional plausible candidate genes and providing insights into the genetic basis of correlated wax traits. Establishing a partially shared genetic architecture, we identified 35 genes for both gc and at least one wax trait, with four considered plausible candidates. Our study uncovered the genetic control of maize leaf waxes, establishing a link between wax composition and gc, with implications for potentially breeding more water-use efficient maize. SIGNIFICANCE STATEMENTWe exploited natural variation in the abundance of maize leaf cuticular waxes to identify genetic determinants of wax composition and its relationship to cuticle function as a barrier against water loss. We identified a set of strongly supported candidate genes with plausible functions in cuticular wax biosynthesis or deposition and added to the evidence for wax esters as the most important wax for water barrier function, offering new tools for modification of cuticle-dependent traits.

genetics↗

bioRxiv2024

A multiplexed transcriptomic analysis of a plant embryonic hourglass

Zoologists have adduced morphological convergence among embryonic stages of closely related taxa, which has been called the phylotypic stage of embryogenesis. Transcriptomic analyses reveal a phylotypic hourglass pattern of gene expression during plant as well as animal embryogenesis, characterized by the accumulation of evolutionarily older and conserved transcripts during mid-embryogenesis, whereas younger less-conserved transcripts predominate at earlier and later embryonic stages. However, widespread comparisons of embryonic gene expression across animal phyla describe an inverse hourglass pattern, where gene expression is correlated during early and late stages but not during mid-development. Here, multiplexed spatial-transcriptomic analyses is used to investigate embryo ontogeny and homology in maize, which has novel, grass-specific morphology. An embryonic-organ genetic network is identified, replete for ancient/conserved genes manifesting a phylotypic hourglass during mid-embryogenesis. Transcriptomic comparisons of grass embryo ontogeny with that of a moss Physcomitrium patens identify a conserved, inverse hourglass pattern across plant phyla, as in animals. The data indicate that the plant phylotypic stage and that of animals is characterized by ancient gene network expression during early histo- and morphogenesis and polarized embryonic development. The data reveal an ancient, convergent mechanism for the evolution of morphological novelty.

plant biology↗

bioRxiv2022

The ratio of auxin to cytokinin controls leaf development and meristem initiation in Physcomitrium patens

Crosstalk between auxin and cytokinin contributes to widespread developmental processes, including root and shoot meristem maintenance, phyllotaxy, and vascular patterning. Although auxin and cytokinin are potent regulators of plant development, our understanding of crosstalk between these hormones is limited to few model systems. The moss Physcomitrium patens (formerly Physcomitrella patens) is a powerful system for studying plant hormone function. Auxin and cytokinin play similar roles in regulating moss gametophore (upright shoot) architecture, as they do in flowering plant shoots. However, auxin-cytokinin crosstalk is poorly understood in moss. Here we find that the ratio of auxin to cytokinin is an important determinant of development in P. patens, especially during leaf development and branch stem cell initiation. Addition of high levels of auxin to P. patens gametophores blocks leaf outgrowth. However, simultaneous addition of high levels of both auxin and cytokinin partially restores leaf outgrowth, suggesting that the ratio of these hormones is the overriding factor. Likewise, during branch initiation and outgrowth, chemical inhibition of auxin synthesis phenocopies cytokinin application. Finally, cytokinin insensitive mutants resemble plants with altered auxin signaling and are hypersensitive to auxin. In summary, our results suggest that the ratio between auxin and cytokinin signaling is the basis for developmental decisions in the moss gametophore.

plant biology↗

bioRxiv2021

Integration of GWAS and TWAS to elucidate the genetic architecture of natural variation for leaf cuticular conductance in maize

The cuticle, a hydrophobic layer of cutin and waxes synthesized by plant epidermal cells, is the major barrier to water loss when stomata are closed. Dissecting the genetic architecture of natural variation for maize leaf cuticular conductance (gc) is important for identifying genes relevant to improving crop productivity in drought-prone environments. To this end, we performed an integrated genome- and transcriptome-wide association study (GWAS/TWAS) to identify candidate genes putatively regulating variation in leaf gc. Of the 22 plausible candidate genes identified, five were predicted to be involved in cuticle precursor biosynthesis and export, two in cell wall modification, nine in intracellular membrane trafficking, and seven in the regulation of cuticle development. A gene encoding an INCREASED SALT TOLERANCE1-LIKE1 (ISTL1) protein putatively involved in intracellular protein and membrane trafficking was identified in GWAS and TWAS as the strongest candidate causal gene. A set of maize nested near-isogenic lines that harbor the ISTL1 genomic region from eight donor parents were evaluated for gc, confirming the association between gc and ISTL1 in a haplotype-based association analysis. The findings of this study provide novel insights into the role of regulatory variants in the development of the maize leaf cuticle, and will ultimately assist breeders to develop drought-tolerant maize for target environments. Sentence summaryWe performed an integrated GWAS/TWAS and identified 22 candidate genes putatively regulating variation in maize leaf gc. The association between gc and the strongest candidate causal gene, ISTL1, was validated with maize nested near-isogenic lines.

genetics↗

bioRxiv2021

Boundary domain genes were recruited to suppress bract growth and promote branching in maize

Grass inflorescence development is diverse and complex and involves sophisticated but poorly understood interactions of genes regulating branch determinacy and leaf growth. Here, we use a combination of transcript profiling, genetic and phylogenetic analyses to investigate tasselsheath1 (tsh1) and tsh4, two maize genes that simultaneously suppress inflorescence leaf growth and inhibit branching. We identify a regulatory network of inflorescence leaf suppression that involves the phase change gene tsh4 upstream of tsh1 and the ligule identity gene liguleless2 (lg2). We also find that a series of duplications in the tsh1 gene lineage facilitated its shift from boundary domain in non-grasses to suppressed inflorescence leaves of grasses. Collectively, these results suggest that the boundary domain genes tsh1 and lg2 were recruited to inflorescence leaves where they suppress growth and regulate a non-autonomous signaling center that promotes inflorescence branching, an important component of yield in cereal grasses.

plant biology↗

bioRxiv2021

Cytokinin-CLAVATA crosstalk is an ancient mechanism regulating shoot meristem homeostasis in land plants

Plant shoots grow from stem cells within Shoot Apical Meristems (SAMs), which produce lateral organs while maintaining the stem cell pool. In the model flowering plant Arabidopsis, the CLAVATA (CLV) pathway functions antagonistically with cytokinin signaling to control the size of the multicellular SAM via negative regulation of the stem cell organizer WUSCHEL (WUS). Although comprising just a single cell, the SAM of the model moss Physcomitrium patens (formerly Physcomitrella) performs equivalent functions during stem cell maintenance and organogenesis, despite the absence of WUS-mediated stem cell organization. Our previous work showed that the stem cell-delimiting function of the CLV pathway receptors CLAVATA1 (CLV1) and RECEPTOR-LIKE PROTEIN KINASE2 (RPK2) is conserved in the moss P. patens. Here, we use P. patens to assess whether CLV-cytokinin crosstalk is also an evolutionarily conserved feature of stem cell regulation. Genetic analyses reveal that CLV1 and RPK2 regulate SAM proliferation via separate pathways in moss. Surprisingly, cytokinin receptor mutants also form ectopic stem cells in the absence of cytokinin signaling. Through modeling, we identified regulatory network archtectures that recapitulated the stem cell phenotypes of clv1 and rpk2 mutants, cytokinin application, cytokinin receptor mutations, and higher-order combinations of these perturbations. These models predict that CLV1 and RPK2 act through separate pathways wherein CLV1 represses cytokinin-mediated stem cell initiation and RPK2 inhibits this process via a separate, cytokinin-independent pathway. Our analysis suggests that crosstalk between CLV1 and cytokinin signaling is an evolutionarily conserved feature of SAM homeostasis that preceded the role of WUS in stem cell organization.

plant biology↗

bioRxiv2021

Fatty acid binding proteins shape the cellular response to activation of the glucocorticoid receptor

Glucocorticoids are steroid hormones that are essential for life in mammals. Therapeutically, they are some of the most cost-effective drugs for the treatment of inflammatory diseases ranging from skin rashes to COVID-19, but their use is limited by adverse effects. Glucocorticoids exert their effects via the glucocorticoid receptor, a type I nuclear hormone receptor which modulates gene expression. The transcriptional activity of some related, but nuclear restricted, type II nuclear hormone receptors can be enhanced by a family of intracellular transport proteins, the fatty acid binding proteins (FABPs). We find that the transcriptional activity of the GR can be altered by a sub-set of FABP family members dependent on the GR-ligand. The ability of some FABPs to selectively promote or limit the transcriptional activity of the GR in a ligand-dependent manner could facilitate the discovery of drugs that narrow GR activity to only the desired subset of therapeutically relevant genes.

pharmacology and toxicology↗

bioRxiv2021

Identification and characterization of two drug-like fragments that bind to the same cryptic binding pocket of Burkholderia pseudomallei DsbA

DiSulfide Bond forming proteins (DSB) play a crucial role in the pathogenicity of many Gram-negative bacteria. Disulfide bond protein A (DsbA) catalyzes the formation of disulfide bonds necessary for the activity and stability of multiple substrate proteins, including many virulence factors. Hence, DsbA is an attractive target for the development of new drugs to combat bacterial infections. Here, we identified two fragments - 1 (bromophenoxy propanamide) and 2 (4-methoxy-N-phenylbenzenesulfonamide), that bind to the DsbA from the pathogenic bacterium Burkholderia pseudomallei, the causative agent of melioidosis. Crystal structures of the oxidized B. pseudomallei DsbA (termed BpsDsbA) co-crystallized with 1 or 2 suggests that both fragments bind to a hydrophobic pocket that is formed by a change in the side chain orientation of tyrosine 110. This conformational change opens a "cryptic" pocket that is not evident in the apo-protein structure. This binding location was supported by 2D-NMR studies which identified a chemical shift perturbation of the tyrosine 110 backbone amide resonance of more than 0.05 ppm upon addition of 2 mM of fragment 1 and over 0.04 ppm upon addition of 1 mM of fragment 2. Although binding was detected by both X-ray crystallography and NMR, the binding affinity (KD) for both fragments was low (above 2 mM), suggesting weak interactions with BpsDsbA. This conclusion is also supported by the modelled crystal structures which ascribe partial occupancy to the ligands in the cryptic binding pocket. Small fragments such as 1 and 2 are not expected to have high binding affinity due to their size and the relatively small surface area that can be involved in intermolecular interactions. However, their simplicity makes them ideal for functionalization and optimization. Identification of the binding sites of 1 and 2 to BpsDsbA could provide a starting point for the development of more potent novel antimicrobial compounds that target DsbA and bacterial virulence. SynopsisDescribes the binding properties of two drug-like fragments to a conformationally dynamic site in the disulfide-bond forming protein A from Burkholderia pseudomallei.

molecular biology↗

bioRxiv2020

Plant stem cell organization and differentiation at single-cell resolution

Plants maintain populations of pluripotent stem cells in shoot apical meristems (SAMs), which continuously produce new aboveground organs. We used single-cell RNA sequencing to achieve an unbiased characterization of the transcriptional landscape of the maize shoot stem-cell niche and its differentiating cellular descendants. Stem cells housed in the SAM tip are engaged in genome integrity maintenance and exhibit a low rate of cell division, consistent with their contributions to germline and somatic cell fates. Surprisingly, we find no evidence for a canonical stem cell organizing center subtending these cells. In addition, we use trajectory inference to trace the gene expression changes that accompany cell differentiation. These data provide a valuable scaffold on which to better dissect the genetic control of plant shoot morphogenesis.

plant biology↗

bioRxiv2019

Genome-wide association study for maize leaf cuticular conductance identifies candidate genes involved in the regulation of cuticle development

The cuticle, a hydrophobic layer of cutin and waxes synthesized by plant epidermal cells, is the major barrier to water loss when stomata are closed at night and under water-limited conditions. Elucidating the genetic architecture of natural variation for leaf cuticular conductance (gc) is important for identifying genes relevant to improving crop productivity in drought-prone environments. To this end, we conducted a genome-wide association study of gc of adult leaves in a maize inbred association panel that was evaluated in four environments (Maricopa, AZ, and San Diego, CA in 2016 and 2017). Five genomic regions significantly associated with gc were resolved to seven plausible candidate genes (ISTL1, two SEC14 homologs, cyclase-associated protein, a CER7 homolog, GDSL lipase, and {beta}-D-XYLOSIDASE 4). These candidates are potentially involved in cuticle biosynthesis, trafficking and deposition of cuticle lipids, cutin polymerization, and cell wall modification. Laser microdissection RNA sequencing revealed that all these candidate genes, with the exception of the CER7 homolog, were expressed in the zone of the expanding adult maize leaf where cuticle maturation occurs. With direct application to genetic improvement, moderately high average predictive abilities were observed for whole-genome prediction of gc in locations (0.46 and 0.45) and across all environments (0.52). The findings of this study provide novel insights into the genetic control of gc and have the potential to help breeders more effectively develop drought-tolerant maize for target environments.\n\nArticle summaryThe cuticle serves as the major barrier to water loss when stomata are closed at night and under water-limited conditions and potentially relevant to drought tolerance in crops. We performed a genome-wide association study to elucidate the genetic architecture of natural variation for maize leaf cuticular conductance. We identified epidermally expressed candidate genes that are potentially involved in cuticle biosynthesis, trafficking and deposition, cutin polymerization, and cell wall modification. Finally, we observed moderately high predictive abilities for whole-genome prediction of leaf cuticular conductance. Collectively, these findings may help breeders more effectively develop drought-tolerant maize.

genetics↗

bioRxiv2019

Network analyses implicate a role for PHYTOCHROME-mediated light signaling in the regulation of cuticle development in plant leaves

Plant cuticles are composed of wax and cutin, and evolved in the land plants as a hydrophobic boundary that reduces water loss from the plant epidermis. The expanding maize adult leaf displays a dynamic, proximodistal gradient of cuticle development, from the leaf base to the tip. Laser microdissection RNA Sequencing (LM-RNAseq) was performed along this proximodistal gradient, and complementary network analyses identified potential regulators of cuticle biosynthesis and deposition. Correlations between cuticle development and cell wall biosynthesis processes were identified, as well as evidence of roles for auxin and brassinosteroids. In addition, our network analyses suggested a previously undescribed function for PHYTOCHROME-mediated light signaling during cuticular wax deposition. Genetic analyses reveal that the phyB1 phyB2 double mutant of maize exhibits abnormal cuticle composition, supporting predictions of our coexpression analyses. Reverse genetic analyses also show that phy mutants of the moss Physcomitrella patens exhibit abnormal cuticle composition, suggesting a role for light-stimulated development of cuticular waxes during plant evolution.

plant biology↗

bioRxiv2019

Changes in lipid composition and ultrastructure associated with functional maturation of the cuticle during adult maize leaf development

Although extensive prior work has characterized cuticle composition, function, ultrastructure and development in many plant species, much remains to be learned about how these features are interrelated. Moreover, very little is known about the adult maize leaf cuticle in spite of its significance for agronomically important traits in this major crop. We analyzed cuticle composition, ultrastructure, and permeability along the developmental gradient of partially expanded adult maize leaves to probe the relationships between these features. The water barrier property is acquired at the cessation of cell expansion. Wax types and chain lengths accumulate asynchronously along the developmental gradient, while overall wax load does not vary. Cutin begins to accumulate prior to establishment of the water barrier and continues thereafter. Ultrastructurally, pavement cell cuticles consist of an epicuticular layer, a thin cuticle proper that acquires an inner, osmiophilic layer during development, and no cuticular layer. Cuticular waxes of the adult maize leaf are dominated by alkanes and wax esters localized mainly in the epicuticular layer. Establishment of the water barrier coincides with a switch from alkanes to esters as the major wax type, and the emergence of an osmiophilic (likely cutin-rich) layer of the cuticle proper.\n\nHiglight statementChemical, ultrastructural and functional analysis of cuticle development in partially expanded adult maize leaves revealed important roles for wax esters and an osmiophilic, likely cutin-rich, layer in protection from dehydration.

plant biology↗

bioRxiv2018

Bulked-Segregant Analysis Coupled to Whole Genome Sequencing (BSA-Seq) for Rapid Gene Cloning in Maize

Forward genetics remains a powerful method for revealing the genes underpinning organismal form and function, and for revealing how these genes are tied together in gene networks. In maize, forward genetics has been tremendously successful, but the size and complexity of the maize genome made identifying mutant genes an often arduous process with traditional methods. The next generation sequencing revolution has allowed for the gene cloning process to be significantly accelerated in many organisms, even when genomes are large and complex. Here, we describe a bulked-segregant analysis sequencing (BSA-Seq) protocol for cloning mutant genes in maize. Our simple strategy can be used to quickly identify a mapping interval and candidate single nucleotide polymorphisms (SNPs) from whole genome sequencing of pooled F2 individuals. We employed this strategy to identify narrow odd dwarf as an enhancer of teosinte branched1, and to identify a new allele of defective kernel1. Our method provides a quick, simple way to clone genes in maize.\n\nReferences numbers for supporting data:figshare and NCBI SRA links to follow

plant biology↗