Abstract

Stomata are microscopic valves in the plant epidermis that efficiently exchange gas while minimizing water loss. Their patterning and differentiation are influenced by various signals, including the phytohormone abscisic acid (ABA). ABA is crucial for stomatal closure during stress conditions, particularly drought. While ABA's role in stomatal movement is well-studied, our understanding of ABA’s role in controlling stomatal development and the mechanisms underlying its interaction with the stomatal development pathway are limited.
In this study, the characterization of ABA-deficient and ABA-signaling mutants indicated that ABA regulation of stomatal development is mediated by upstream signaling components, including PYR/PYL/RCAR (ABA receptors), ABI1/ABI2 (ABA phosphatases), and SnRK2.2/2.3/2.6 (SnRK2 kinases), but is likely independent of the transcription factors in the core ABA signaling pathway. We analyzed the epidermal phenotype of ABA signaling mutants. The abi1-2 abi2-2 mutants, lacking two negative ABA signaling regulators, showed remarkably fewer stomatal and non-stomatal cells on the plant epidermis. Among nine ABA PP2Cs, we further identified ABI1 and ABI2 phosphatases as key regulators of stomatal development. To investigate epidermal defect generated in ABA signaling mutants, I examined the epidermal phenotype of wild type, abi1-2 abi2-2, and snrk2.2 snrk2.3 mutants using the TMMpro: GUS, a stomatal lineage cell-specific marker. The abi1-2 abi2-2 mutant showed fewer epidermal cells entering stomatal lineages, while the snrk2.2 snrk2.3 mutant exhibited an increased number. This finding indicates ABA restricts stomatal cell lineages. Using high-order ABA and stomatal mutants, we next investigated the molecular link between ABA signaling and stomatal developmental pathways. Our genetic interaction studies showed that ABIs act upstream of the stomatal receptors TOO MANY MOUTHS (TMM) and ERECTA family members (ER, ERL1, ERL2), the MAP kinases MPK3 and MPK6, and the transcription factor MUTE in the stomatal development pathway. Based on our genetic interaction data, we analyzed the physical interactions between ABI1/2 and ERECTA family receptors and TMM. Our data demonstrated that ABI1, but not ABI2, interacts with stomatal receptors, ER, ERL1, and ERL2.
Our data reveal novel interactions between ABA signaling and stomatal development pathways, expanding our understanding of ABA's role beyond stomatal movement regulation. These findings suggest potential strategies plants may employ to optimize growth and adapt to environmental stresses like drought. This research establishes a framework for future investigations into the intricate links between ABA and stomatal developmental pathways.