INTACT-based guard cell transcriptomes from a progressive drought time course reveal targets for modifying stomatal responses

Drought is an important environmental stress that limits crop production. Guard cells (GCs) act to control the rate of water loss. To better understand how gene expression in GCs changes during progressive drought, we generated GC-specific RNA-seq transcriptomes during mild, moderate, and severe drought stress. Additionally, we sampled re-watered plants after severe drought. These transcriptomes were generated using the INTACT (isolation of nuclei tagged in specific cell types) system to capture the RNA from GC nuclei. We optimized the INTACT protocol for Arabidopsis thaliana leaf tissue, incorporating fixation to preserve RNA during nuclear isolation. To identify gene expression changes unique to GCs, we also generated INTACT transcriptomes from all leaf cell types, using the 35S viral promoter. These data sets highlight shared and unique gene expression changes between GCs and the bulk leaf tissue. Only GCs have detectable gene expression changes at the earliest drought time point, and a high percentage of moderate drought GC DEGs are not observed in severe drought, unlike the bulk leaf tissue, showing that GCs tailor their gene expression changes to drought severity. A thermal imaging screen of mutants of 80 candidate early drought-responsive genes revealed that 10 of these exhibit a cooler-than-wild-type phenotype under moderate drought conditions. The drought-responsive GC and leaf RNA-seq transcriptomes are available in the Arabidopsis ePlant at the Bio-Analytic Resource for Plant Biology website. These findings provide valuable insights into GC-specific drought responses and identify targets for enhancing drought tolerance in crops.