An auxin signaling network translates low-sugar-state input into compensated cell enlargement in the fugu5 cotyledon

Author summary How leaf size is determined is a longstanding question in biology. In the simplest scenario, leaf size would be a function of cell number and size. Yet, accumulating evidence on the model plant Arabidopsis thaliana suggested the presence of compensatory mechanisms, so that when the leaf contains fewer cells, the size of each cell is unusually increased (the so-called compensated cell enlargement (CCE)). While decreased cell numbers in the compensation exhibiting fugu5 mutants have been ascribed to reduced sugar biosynthesis from seed oil reserves, molecular mechanisms underlying CCE remain ill-known. Recently, IBA (a precursor of the phytohormone auxin) has emerged as a potential regulator of CCE. Here, to further illuminate the role of IBA in CCE, we used a series of high-order mutants and analyzed their genetic interaction with fugu5. We found that while CCE in fugu5 was promoted by IBA, defects in IBA-to-auxin conversion, auxin response, or the vacuolar V-ATPase activity alone cancelled CCE. This provides a scenario in which following seed germination, the low-sugar-state triggers auxin synthesis, leading to CCE through the activation of the V-ATPase, illustrating how fine-tuning cell and organ size regulation depend on interplays between metabolism and auxin levels in plants. In plants, the effective mobilization of seed nutrient reserves is crucial during germination and for seedling establishment. The Arabidopsis H+-PPase-loss-of-function fugu5 mutants exhibit a reduced number of cells in the cotyledons. This leads to enhanced post-mitotic cell expansion, also known as compensated cell enlargement (CCE). While decreased cell numbers have been ascribed to reduced gluconeogenesis from triacylglycerol, the molecular mechanisms underlying CCE remain ill-known. Given the role of indole 3-butyric acid (IBA) in cotyledon development, and because CCE in fugu5 is specifically and completely cancelled by ech2, which shows defective IBA-to-indoleacetic acid (IAA) conversion, IBA has emerged as a potential regulator of CCE. Here, to further illuminate the regulatory role of IBA in CCE, we used a series of high-order mutants that harbored a specific defect in IBA-to-IAA conversion, IBA efflux, IAA signaling, or vacuolar type H+-ATPase (V-ATPase) activity and analyzed the genetic interaction with fugu5-1. We found that while CCE in fugu5 was promoted by IBA, defects in IBA-to-IAA conversion, IAA response, or the V-ATPase activity alone cancelled CCE. Consistently, endogenous IAA in fugu5 reached a level 2.2-fold higher than the WT in 1-week-old seedlings. Finally, the above findings were validated in icl-2, mls-2, pck1-2 and ibr10 mutants, in which CCE was triggered by low sugar contents. This provides a scenario in which following seed germination, the low-sugar-state triggers IAA synthesis, leading to CCE through the activation of the V-ATPase. These findings illustrate how fine-tuning cell and organ size regulation depend on interplays between metabolism and IAA levels in plants.

Automated summary

This study investigated the role of IBA in regulating cell and organ size in plants, demonstrating that IBA can promote CCE. However, defects in IBA-to-IAA conversion, IAA response, or V-ATPase activity cancel CCE, highlighting the crucial interplay between metabolism and auxin levels in fine-tuning cell and organ size regulation in plants.