Biology subjects

Gilroy, S.

Publications and source records attributed to Gilroy, S..

5 recordsLinked to original sources

bioRxiv2025

Mutations adjacent to the nucleotide-binding cleft of Arabidopsis thaliana ACTIN7 confer resistance to the actin-disrupting compound latrunculin B

A forward genetic screen identified a dominant-negative Arabidopsis thaliana mutant resistant to growth inhibition caused by the actin-disrupting compound latrunculin B (LatB). Map-based cloning combined with whole-genome sequencing revealed that the mutant referred to here as lbr1 for LatB resistant1 had a point mutation in the AT5G09810 gene, which encodes the vegetative actin (ACT) isoform ACT7. The cytosine to thymine mutation in the second exon of ACT7 of lbr1 led to substitution of proline to serine at position 34 (P34S) adjacent to the nucleotide-binding cleft of the ACT7 protein. Confirmation that ACT7 is the causal gene for the lbr1 phenotype was achieved through transgenic complementation with ACT7 wild type (ACT7WT) and ACT7 P34S (ACT7P34S) constructs. ACT7P34 also rescued the seedling developmental defects and conferred partial resistance to LatB in the recessive act7-5 mutant. Furthermore, expressing a P34S mutation in ACT2 (ACT2P34S), another vegetative ACT isoform, conferred partial LatB resistance to wild type. Finally, site-directed mutagenesis of ACT7 amino acid residues forming putative hydrogen bonds with LatB, based on yeast and mammalian actin docking and structural analyses, reveals domains adjacent to the actin nucleotide-binding cleft crucial for LatBs effects on the plant actin cytoskeleton. HighlightCharacterization of the dominant-negative lbr1 mutant uncovers amino acid residues in the actin protein crucial for latrunculins mechanism of action in plants.

cell biology↗

bioRxiv2024

GLARE: DISCOVERING HIDDEN PATTERNS IN SPACEFLIGHT TRANSCRIPTOME USING REPRESENTATION LEARNING

AO_SCPLOWBSTRACTC_SCPLOWSpaceflight studies present novel insights into biological processes through exposure to stressors outside the evolutionary path of terrestrial organisms. Despite limited access to space environments, numerous transcriptomic datasets from spaceflight experiments are now available through NASAs GeneLab data repository, which allows public access to these datasets, encouraging further analysis. While various computational pipelines and methods have been used to process these transcriptomic datasets, learning-model-driven analyses have yet to be applied to a broad array of such spaceflight-related datasets. In this study, we propose an open-source pipeline, GLARE: GeneLAb Representation learning pipelinE, which consists of training different representation learning approaches from manifold learning to self-supervised learning that enhance the performance of downstream analytical tasks. We illustrate the utility of GLARE by applying it to gene-level transcriptional values from the results of the CARA spaceflight experiment, an Arabidopsis root tip transcriptome dataset that spanned light, dark, and microgravity treatments. We show that GLARE not only substantiated the findings of the original study concerning cell wall remodeling but also revealed additional patterns of gene expression affected by the treatments, including evidence of hypoxia. This work suggests there is great potential to supplement the insights drawn from initial studies on spaceflight omics-level data through further machine-learning-enabled analyses.

plant biology↗

bioRxiv2023

Glycome profiling and immunohistochemistry uncover spaceflight-induced changes in non-cellulosic cell wall components in Arabidopsis thaliana seedling roots

A large and diverse library of glycan-directed monoclonal antibodies (mAbs) was used to determine if plant cell walls are modified by low-gravity conditions encountered during spaceflight. This method called glycome profiling (glycomics) revealed global differences in non-cellulosic cell wall epitopes in Arabidopsis thaliana root extracts recovered from RNA purification columns between seedlings grown on the International Space Station-based Vegetable Production System and paired ground (1-g) controls. Immunohistochemistry on 11-day-old seedling primary root sections showed that ten of twenty-two mAbs that exhibited spaceflight-induced increases in binding through glycomics, labeled space-grown roots more intensely than those from the ground. The ten mAbs recognized xyloglucan, xylan, and arabinogalactan epitopes. Notably, three xylem-enriched unsubstituted xylan backbone epitopes were more intensely labeled in space-grown roots than in ground-grown roots, suggesting that the spaceflight environment accelerated root secondary cell wall formation. This study highlights the feasibility of glycomics for high-throughput evaluation of cell wall glycans using only root high alkaline extracts from RNA purification columns, and subsequent validation of these results by immunohistochemistry. This approach will benefit plant space biological studies because it extends the analyses possible from the limited amounts of samples returned from spaceflight and help uncover microgravity-induced tissue-specific changes in plant cell walls.

plant biology↗

bioRxiv2020

NASA GeneLab RNA-Seq Consensus Pipeline: Standardized Processing of Short-Read RNA-Seq Data

With the development of transcriptomic technologies, we are able to quantify precise changes in gene expression profiles from astronauts and other organisms exposed to spaceflight. Members of NASA GeneLab and GeneLab-associated analysis working groups (AWGs) have developed a consensus pipeline for analyzing short-read RNA-sequencing data from spaceflight-associated experiments. The pipeline includes quality control, read trimming, mapping, and gene quantification steps, culminating in the detection of differentially expressed genes. This data analysis pipeline and the results of its execution using data submitted to GeneLab are now all publicly available through the GeneLab database. We present here the full details and rationale for the construction of this pipeline in order to promote transparency, reproducibility and reusability of pipeline data, to provide a template for data processing of future spaceflight-relevant datasets, and to encourage cross-analysis of data from other databases with the data available in GeneLab.

bioinformatics↗

bioRxiv2018

A novel role for Cyclic Nucleotide-Gated Ion Channel 2 (DND1) in auxin signaling

Cyclic Nucleotide Gated Ion Channels (CNGCs) have been firmly established as Ca2+-conducting ion channels that regulate a wide variety of physiological responses in plants. CNGC2 has been implicated in plant immunity and Ca2+ signaling due to the autoimmune phenotypes exhibited by null mutants of CNGC2. However, cngc2 mutants display additional phenotypes that are unique among autoimmune mutants, suggesting that CNGC2 has functions beyond defense and generates distinct Ca2+ signals in response to different triggers. In this study we found that cngc2 mutants showed reduced gravitropism, consistent with a defect in auxin signaling. This was mirrored in the diminished auxin response detected by the auxin reporters DR5::GUS and DII-VENUS and in a strongly impaired auxin-induced Ca2+ response. Moreover, the cngc2 mutant exhibits higher levels of the endogenous auxin indole-3-acetic acid (IAA), indicating that excess auxin in cngc2 causes its pleiotropic phenotypes. These auxin signaling defects and the autoimmunity syndrome of cngc2 could be suppressed by loss-of-function mutations in the auxin biosynthesis gene YUCCA6 (YUC6), as determined by identification of the cngc2 suppressor mutant repressor of cngc2 (rdd1) as an allele of YUC6. A loss-of-function mutation in the upstream auxin biosynthesis gene TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS (TAA1, WEAK ETHYLENE INSENSITIVE8) also suppressed the cngc2 phenotypes, further supporting the tight relationship between CNGC2 and the TAA-YUC-dependent auxin biosynthesis pathway. Taking these results together, we propose that the Ca2+ signal generated by CNGC2 is a part of the negative feedback regulation of auxin homeostasis in which CNGC2 balances cellular auxin perception by influencing auxin biosynthesis.

plant biology↗