Elucidating Novel Factors in the Plant Guard Cell CO2 Signaling Pathway: A Complementary Forward Genetic Screen in Arabidopsis thaliana

Climate change mitigation is at the forefront of global discussion, thus the study of plant responses to atmospheric CO2 is of great ecophysiological importance. Stomata, the pores of the plant, open and close in response to varying atmospheric CO2 levels. Therefore, it is important to investigate transduction pathways that mediate stomatal responses to CO2 variation. Since elevated CO2 reduces stomatal apertures, this response is of particular interest considering the rapidly rising atmospheric CO2 levels. While plants are considered to be a promising organism for atmospheric CO2 mitigation, still some components of the CO2 response pathway are not well understood, specifically the CO2 sensor in guard cells for both stomatal movements and stomatal development. This study aims to elucidate novel factors in the plant guard cell CO2 signaling pathway. This project consists of a complementary forward genetic screen and preliminary characterization of a robust candidate identified from the screen. The complementary forward genetic screen, performed in Arabidopsis thaliana, was conducted using two tools: the gain-of-function FOX-hunting plant line approach and a homologous gene silencing artificial-microRNA (amiRNA) plant line approach. Both FOX-hunting and amiRNA lines were screened in conditions that specifically favored the identification of mutants involved in stomatal movement or stomatal development response to CO2. A total of 43 amiRNA and 19 FOX-line candidate plants were identified and sequenced. One candidate repeatedly identified in the screen was an amiRNA line targeting two leucine-rich repeat (LRR) family genes in the A. thaliana genome: AT3G43740 and AT5G21090. This LRR candidate showed a partially impaired gas exchange response and a lower stomatal count compared to the wild-type, indicating the observed phenotype is a product of a stomatal development mutation. The remaining candidate plants identified from the screen will be characterized in the future, using this preliminary characterization as precedent, with aim to further the understanding of the guard cell CO2 signal transduction pathway in plants