Dynamic growth re-orientation orchestrates flatness in the Arabidopsis leaf

Abstract

The growth and division of cells in plant leaves is highly dynamic in time and space, all while the cells cannot move relative to their neighbors. Given these constraints, models predict that long range regulatory systems must exist to maintain flat forms. Juxtaposed microRNA (miRNA) networks could serve as one of these regulatory systems. One of these miRNAs, miR319 is thought to be expressed from the base of leaves and to promote growth by degrading class II TCP transcription factor mRNAs. A miR319 overexpression mutant,jagged and wavy (jaw-D)exhibits rippling and undulating leaves, consistent with biomechanical predictions that without genetic spatial coordination, tissues will deform. It has been theorized thatjaw-Drippling results from overgrowth at the margins, however this does not fully address how miR319 expression from the base of wild-type (WT) leaves allows them to flatten. Here, we track the growth, cell division and cell maturation in live WT andjaw-Dleaves to ask how miR319 expression in WT promotes flattening. This data revealed the importance of spatially distinct growth, division and differentiation patterns in WT leaves, which are missing injaw-D. We propose that WT leaf cells respond to differentiation cues to dynamically re-orient growth in specific tissue locations and regulate flattening.

“An optimized pipeline for live imaging whole Arabidopsis leaves at cellular resolution”

“…This uncovered that, at 7 DAS, the cells in jaw-D leaves exhibit greater average areal growth amongst cells and less variability between cells than WT (Figure 7A-C, Student’s t-test p < .0001, CV asymptotic test p < 1.578488e-19). In other work, we have shown that jaw-D petioles are shorter than WT (Harline et al 2022). Our method suggests that this final organ level phenotype could be created by homogeneity and misregulation of growth patterns in the jaw-D petiole.…”

“…Combining our imaging techniques with our custom MorphoGraphX scripts enables us to capture a large dataset encompassing the early development of WT and jaw-D leaves (Figure 6). From this dataset, we could analyze cell growth, division and morphology characteristics between different tissue regions, like the petiole and the margin (Figure 7 and Harline et al 2022).…”

“…By labeling the cells of the petiole through the script, we were able to compare the average In other work, we have shown that jaw-D petioles are shorter than WT (Harline et al 2022). Our method suggests that this final organ level phenotype could be created by homogeneity and misregulation of growth patterns in the jaw-D petiole.…”

“…From this dataset, we could analyze cell growth, division and morphology characteristics between different tissue regions, like the petiole and the margin (Figure 7 and Harline et al 2022).…”

“…Our samples contain a small plasma membrane-localized protein tagged with a fluorescent protein, so we are able to get high resolu- tion images of every cell border in these images. This allows us to track the creation of recognizable leaf tissue containing thousands of cells from an initial unrecognizable nub of tens of cells (Harline et al 2022). Our samples grow from hundreds of micrometers in area to millimeters in area, so they quickly exceed the single 20x imaging window at which the plasma membrane marker is resolvable (~5 DAS).…”