Biology subjects

Stanley, L. E.

Publications and source records attributed to Stanley, L. E..

5 recordsLinked to original sources

bioRxiv2024

Just add water: A simple floral bud injection method for stable Agrobacterium-mediated transformation in two ecotypes of Mimulus guttatus

Stable transformation is the biggest challenge for studying gene function in plants. In most plant species, stable transformation requires the development of arduous tissue culture and regeneration methods that often fail and can cause undesirable genetic changes. Floral transformation methods bypass these challenges by infiltrating Agrobacterium directly into the ovules of developing flowers. However, this method has had limited success outside of a handful of plant species and genotypes within those species. Here, we demonstrate that the floral infiltration method is effective for some, but not all genotypes of the yellow monkeyflower, Mimulus guttatus, a model system for studies of ecology and evolution. To conduct transformation in genotypes where infiltration failed, we developed a novel floral injection method. Beyond expanding the number of genotypes that can be transformed, the floral injection method has other advantages, such as allowing plants to be infiltrated multiple times and a reduction in floral abscission and male sterility. Through a combination of floral infiltration and injection methods, we were able to transform both coastal perennial and inland annual genotypes of M. guttatus, which sets the stage for understanding the molecular genetic underpinnings of local adaptation to the divergent habitats occupied by these distinctive ecotypes. Overall, the development of these efficient transformation methods will open up a wide range of new research avenues in the M. guttatus species complex.

plant biology↗

bioRxiv2023

The role of breakpoint mutations, supergene effects, and ancient nested rearrangements in the evolution of adaptive chromosome inversions in the yellow monkey flower, Mimulus guttatus

Chromosomal inversion polymorphisms are ubiquitous across the diversity of diploid organisms and play a significant role in the evolution of adaptations in those species. Inversions are thought to operate as supergenes by trapping adaptive alleles at multiple linked loci through the suppression of recombination. While there is now considerable support for the supergene mechanism of inversion evolution, the extent to which inversions trap pre-existing adaptive genetic variation versus accumulate new adaptive variants over time remains unclear. In this study, we report new insights into the evolutionary dynamics of a locally adaptive chromosomal inversion polymorphism (inv_chr8A), which contributes to the evolutionary divergence between coastal perennial and inland annual ecotypes of the yellow monkeyflower, Mimulus guttatus. This research was enabled by the sequencing, assembly, and annotation of new annual and perennial genomes of M. guttatus using Oxford Nanopore long-read sequencing technology. In addition to the adaptive inv_chr8A inversion, we identified three other large inversion polymorphisms, including a previously unknown large inversion (inv_chr8B) nested within the inv_chr8A. Through population genomic analyses and comparative genomics, we determined that the nested inv_chr8B inversion is significantly older than the larger chromosomal inversion in which it resides. We also evaluated key candidate genes involved in gibberellin biosynthesis and anthocyanin regulation, which we hypothesize to underlie the adaptive phenotypic effects of the inv_chr8A inversion. Although little evidence was found to suggest that inversion breakpoint mutations drive adaptive phenotypic effects, our findings support the supergene mechanism of adaptive evolution as a dynamic and continuous process.

genomics↗

bioRxiv2018

Formation of periodic pigment spots by the reaction-diffusion mechanism

Many organisms exhibit visually striking spotted or striped pigmentation patterns. Turings reaction-diffusion model postulates that such periodic pigmentation patterns form when a local autocatalytic feedback loop and a long-range inhibitory feedback loop interact. At its simplest, this network only requires one self-activating activator that also activates a repressor, which inhibits the activator and diffuses to neighboring cells. However, the molecular activators and repressors fully fitting this versatile model remain elusive. Here, we characterize an R2R3-MYB activator and an R3-MYB repressor in monkeyflowers that correspond to Turings model and explain how periodic anthocyanin spots form. Notably, disrupting this pattern impacts pollinator visitation. Thus, subtle changes in simple reaction-diffusion networks are likely essential contributors to the evolution of the remarkable diversity of periodic pigmentation patterns in flowers.

plant biology↗

bioRxiv2018

Developmental genetics of corolla tube formation: role of the tasiRNA-ARF pathway

More than 80,000 angiosperm species produce flowers with petals fused into a corolla tube. As an important element of the tremendous diversity of flower morphology, the corolla tube plays a critical role in many specialized interactions between plants and animal pollinators (e.g., beeflies, hawkmoths, hummingbirds, nectar bats), which in turn drives rapid plant speciation. Despite its clear significance in plant reproduction and evolution, the corolla tube remains one of the least understood plant structures from a developmental genetics perspective. Through mutant analyses and transgenic experiments, here we show that the tasiRNA-ARF pathway is required for corolla tube formation in the monkeyflower species Mimulus lewisii. Loss-of-function mutations in the M. lewisii orthologs of ARGONAUTE7 and SUPPRESSOR OF GENE SILENCING 3 cause a dramatic decrease in abundance of TAS3-derived small RNAs and a moderate up-regulation of AUXIN RESPONSE FACTOR 3 (ARF3) and ARF4, which lead to inhibition of lateral expansion of the bases of petal primordia and complete arrest of the upward growth of the inter-primordial regions, resulting in unfused corollas. By using an auxin reporter construct, we discovered that auxin distribution is continuous along the petal primordium base and the inter-primordial region during the critical stage of corolla tube formation in the wild-type, and that this auxin distribution is much weaker and more restricted in the mutant. Together, these results suggest a new conceptual model highlighting the central role of auxin directed synchronized growth of the petal primordium base and the inter-primordial region in corolla tube formation.

plant biology↗

bioRxiv2017

A Tetratricopeptide Repeat Protein Regulates Carotenoid Biosynthesis and Chromoplast Development in Monkeyflowers (Mimulus)

The incredible diversity of floral color and pattern in nature is largely determined by the transcriptional regulation of anthocyanin and carotenoid biosynthetic genes. While the transcriptional control of anthocyanin biosynthesis is well understood, little is known about the factors regulating the carotenoid biosynthetic pathway in flowers. Here, we characterize the Reduced Carotenoid Pigmentation 2 (RCP2) locus from two monkeyflower (Mimulus) species, the bumblebee-pollinated M. lewisii and hummingbird-pollinated M. verbenaceus. We show that loss-of-function mutations of RCP2 cause drastic down-regulation of the entire carotenoid biosynthetic pathway in these species. Through bulk segregant analysis and transgenic experiments, we have identified the causal gene underlying RCP2, encoding a tetratricopeptide repeat (TPR) protein that is closely related to the Arabidopsis Reduced Chloroplast Coverage (REC) proteins. RCP2 appears to regulate carotenoid biosynthesis independently of RCP1, a previously identified R2R3-MYB master regulator of carotenoid biosynthesis. We show that RCP2 is required for chromoplast development and suggest that it most likely regulates the expression of carotenoid biosynthetic genes through chromoplast-to-nucleus retrograde signaling. Furthermore, we demonstrate that M. verbenaceus is just as amenable to chemical mutagenesis and in planta transformation as the more extensively studied M. lewisii, making these two species an excellent platform for comparative developmental genetics studies of two closely related species with dramatic phenotypic divergence.

genetics↗