Targeting editing of tomato <i>SPEECHLESS</i> cis-regulatory regions generates plants with altered stomatal density in response to changing climate conditions

Abstract

Abstract Flexible developmental programs enable plants to customize their organ size and cellular composition. In leaves of eudicots, the stomatal lineage produces two essential cell types, stomata and pavement cells, but the total numbers and ratio of these cell types can vary. Central to this flexibility is the stomatal lineage initiating transcription factor, SPEECHLESS (SPCH). Here we show, by multiplex CRISPR/Cas9 editing of SlSPCH cis- regulatory sequences in tomato, that we can identify variants with altered stomatal development responses to light and temperature cues. Analysis of tomato leaf development across different conditions, aided by newly-created tools for live-cell imaging and translational reporters of SlSPCH and its paralogues SlMUTE and SlFAMA, revealed the series of cellular events that lead to the environmental change-driven responses in leaf form. Plants bearing the novel SlSPCH variants generated in this study are powerful resources for fundamental and applied studies of tomato resilience in response to climate change. Significance statement Plants can change their shape, size and cellular composition in response to environmental cues. Here, by precise gene editing of a core stomatal development regulator gene in tomato, we generate new alleles with enhanced or dampened responses to light and temperature cues. Combined with live imaging of development, we show the genetic and cellular pathways that contribute to customization of the leaf epidermis, and how this could lead to better climate-adapted varieties.