Changes in Gene Expression in Space and Time Orchestrate Environmentally-Mediated Shaping of Root Architecture

Shaping of root architecture is a quintessential developmental response that involves the concerted action of many different cell types, is highly dynamic and underpins root plasticity. To determine to what extent the environmental regulation of lateral root development is a product of cell type preferential activities, we tracked transcriptomic responses to two different treatments that both change root development in Arabidopsis thaliana, at an unprecedented level of temporal detail. We found that individual transcripts are expressed with a very high degree of temporal and spatial specificity, yet biological processes are commonly regulated, in a mechanism we term response non-redundancy. Using causative gene network inference to compare the genes regulated in different cell types and during responses to nitrogen and a biotic interaction we found that common transcriptional modules often regulate the same gene families, but control different individual members of these families, specific to response and cell type. This reinforces that the activity of a gene cannot be defined simply as molecular function; rather, it is a consequence of spatial location, expression timing and environmental responsiveness. The response of cortical and pericycle cells, isolated using Fluorescence Activated Cell Sorting (FACS) was analysed 0 (before treatment) and 1, 2, 4, 6, 7, 10, 12, 14, 16, 20, 24 and 48 hours after treatment with 5mM KNO3 or S. meliloti in Arabidopsis thaliana Col0 roots; both genotypes carried GFP reporters marking either pericycle or cortical root cells. Gene expression was analysed using NimbleGen 12 x 135K microarrays, designed for the TAIR10 genome. Seedlings were grown on low nitrogen (0.1mM KNO3) agar plates for 9 days, transferred to fresh plates containing either 0.1 mM KNO3 (untreated and S. meliloti).or 5 mM KNO3 plates. At the end of the treatment, roots were harvested, protoplasts generated and GFP-marked epidermal and cortical cells isolated according to Gifford et al 2008 PNAS. All experiments were carried out in at least triplicate, with QC analysis used to select a complete array dataset.