Title : Growth directions and stiffness across cell layers determine whether tissues stay smooth or buckle
Abstract:
The link between organ morphology and function is well-established, yet what factors coordinate cell growth across tissue layers to form distinct organ shapes remains an unresolved open question. We aimed to address this question using Arabidopsis sepals. While their smooth shape plays a critical role in properly enclosing and protecting developing flower buds, what growth factors underlie this smooth morphology remains elusive. We used an Arabidopsis gain-of-function mutant of ASYMMETRIC LEAVES 2 (AS2) transcription factor, named as2-7D, as a tool to investigate how the wild-type (WT) forms smooth sepals. In as2-7D, AS2 is expressed on both the abaxial and adaxial epidermal layers, unlike the WT where AS2 is expressed only on the inner epidermis. Our mathematical model suggests that the overgrowth of the outer surface, coupled with reduced stiffness, could theoretically lead to buckling. We show that during early sepal development, ectopic AS2 expression disrupts cell growth patterns on both surfaces, resulting in mechanical stress-induced buckling. We also found the cells on the outer epidermis of as2-7D grow in a misdirected manner, initiating buckling in the proximal-distal direction. Through osmotic treatment assays, we show that the outer epidermis of as2-7D is significantly softer than the inner surface, whereas both layers of the WT have similar stiffness. Therefore, both conditions required for buckling are met in as2-7D but not in wild type sepals. Additionally, overexpression of KRP1, a cyclin-dependent kinase inhibitor in as2-7D limits sideways growth and increases outer epidermal stiffness, thereby substantially reducing buckling and thus supporting our model. We found ectopic PIN1 convergence sites especially where outgrowths begin to develop, and blocking auxin transport significantly reduces outgrowths. This suggests that buckling is sufficient to generate PIN1 convergence mediated outgrowth formation. Our work shows that plants coordinate growth and tissue stiffness throughout the organ to promote smooth organ formation.
