The root of Arabidopsis thaliana is used as a model system to unravel the molecular nature of cell elongation and its arrest. From a micro-array performed on roots that were treated with aminocyclopropane-1-carboxylic acid (ACC), the precursor of ethylene, a Small auxin-up RNA (SAUR)-like gene was found to be up regulated. As it appeared as the 76th gene in the family, it was named SAUR76. Root and leaf growth of overexpression lines ectopically expressing SAUR76 indicated the possible involvement of the gene in the division process. Using promoter::GUS and GFP lines strong expression was seen in endodermal and pericycle cells at the end of the elongation zone and during several stages of lateral root primordia development. ACC and IAA/NAA were able to induce a strong up regulation of the gene and changed the expression towards cortical and even epidermal cells at the beginning of the elongation zone. Confirmation of this up regulation of expression was delivered using qPCR, which also indicated that the expression quickly returned to normal levels when the inducing IAA-stimulus was removed, a behaviour also seen in other SAUR genes. Furthermore, confocal analysis of protein-GFP fusions localized the protein in the nucleus, cytoplasm and plasma membrane. SAUR76 expression was quantified in several mutants in ethylene and auxin-related pathways, which led to the conclusion that the expression of SAUR76 is mainly regulated by the increase in auxin that results from the addition of ACC, rather than by ACC itself.

In plants many developmental processes are regulated by auxin and its directional transport. PINOID (PID) kinase helps to regulate this transport by influencing polar recruitment of PIN efflux proteins on the cellular membranes. We investigated how altered auxin levels affect leaf growth in Arabidopsis thaliana. Arabidopsis mutants and transgenic plants with altered PID expression levels were used to study the effect on auxin distribution and leaf development. Single knockouts showed small pleiotropic growth defects. Contrastingly, several leaf phenotypes related to changes in auxin concentrations and transcriptional activity were observed in PID overexpression (PIDOE) lines. Unlike in the knockout lines, the leaves of PIDOE lines showed an elevation in total indole-3-acetic acid (IAA). Accordingly, enhanced DR5-visualized auxin responses were detected, especially along the leaf margins. Kinematic analysis revealed that ectopic expression of PID negatively affects cell proliferation and expansion rates, yielding reduced cell numbers and small-sized cells in the PIDOE leaves. We used PIDOE lines as a tool to study auxin dose effects on leaf development and demonstrate that auxin, above a certain threshold, has a negative affect on leaf growth. RNA sequencing further showed how subtle PIDOE-related changes in auxin levels lead to transcriptional reprogramming of cellular processes.

Drought plays a central role in increasing the incidence and severity of dry root rot (DRR) disease in chickpea. This is an economically devastating disease, compromising chickpea yields particularly severely in recent years due to erratic rainfall patterns. Macrophomina phaseolina (formerly Rhizoctonia bataticola) is the causal agent of DRR disease in the chickpea plant. The infection pattern in chickpea roots under well-watered conditions and drought stress are poorly understood at present. This study provides detailed disease symptomatology and the characteristics of DRR fungus at morphological and molecular levels. Using microscopy techniques, the infection pattern of DRR fungus in susceptible chickpea roots was investigated under well-watered and drought stress conditions. Our observations suggested that drought stress intensifies the progression of already ongoing infection by weakening the endodermal barrier and overall defense. Transcriptomic analysis suggested that the plant’s innate immune defense program is downregulated in infected roots when subjected to drought stress. Further, genes involved in hormonal regulation are differentially expressed under drought stress. These findings provide hints in terms of potential chickpea genes to target in crop improvement programs to develop climate change-resilient cultivars.

Under normal and stress conditions plant growth require a complex interplay between phytohormones and reactive oxygen species (ROS). However, details of the nature of this crosstalk remain elusive. Here, we demonstrate that PINOID (PID), a serine threonine kinase of the AGC kinase family, perturbs auxin homeostasis, which in turn modulates rosette growth and induces stress responses in Arabidopsis plants. Arabidopsis mutants and transgenic plants with altered PID expression were used to study the effect on auxin levels and stress-related responses. In the leaves of plants with ectopic PID expression an accumulation of auxin, oxidative burst and disruption of hormonal balance was apparent. Furthermore, PID overexpression led to the accumulation of antioxidant metabolites, while pid knockout mutants showed only moderate changes in stress-related metabolites. These physiological changes in the plants overexpressing PID modulated their response toward external drought and osmotic stress treatments when compared to the wild type. Based on the morphological, transcriptome, and metabolite results, we propose that perturbations in the auxin hormone levels caused by PID overexpression, along with other hormones and ROS downstream, cause antioxidant accumulation and modify growth and stress responses in Arabidopsis. Our data provide further proof for a strong correlation between auxin and stress biology.

High ambient temperature suppresses Arabidopsis (Arabidopsis thaliana) rosette leaf area and elongates the stem and petiole. While the mechanism underlying the temperature-induced elongation response has been extensively studied, the genetic basis of temperature regulation of leaf size is largely unknown. Here, we show that warm temperature inhibits cell proliferation in Arabidopsis leaves, resulting in fewer cells compared to the control condition. Cellular phenotyping and genetic and biochemical analyses established the key roles of PHYTOCHROME INTERACTING FACTOR4 (PIF4) and TEOSINTE BRANCHED1/CYCLOIDEA/PCF4 (TCP4) transcription factors in the suppression of Arabidopsis leaf area under high temperature by a reduction in cell number. We show that temperature-mediated suppression of cell proliferation requires PIF4, which interacts with TCP4 and regulates the expression of the cell cycle inhibitor KIP-RELATED PROTEIN1 (KRP1) to control leaf size under high temperature. Warm temperature induces binding of both PIF4 and TCP4 to the KRP1 promoter. PIF4 binding to KRP1 under high temperature is TCP4 dependent as TCP4 regulates PIF4 transcript levels under high temperature. We propose a model where a warm temperature-mediated accumulation of PIF4 in leaf cells promotes its binding to the KRP1 promoter in a TCP4-dependent way to regulate cell production and leaf size. Our finding of high-temperature-mediated transcriptional upregulation of KRP1 integrates a developmental signal with an environmental signal that converges on a basal cell regulatory process.

These authors contributed equally to this work[Abstract] The leaf is the major functional part of the shoot performing the bulk of photosynthetic activity. Its development is relatively plastic allowing the plant to adapt to environmental changes by modifying leaf size and anatomy. Moreover, a leaf is made up of various distinct cell layers, each having specialized functions. To understand functional adaptation and the development of the leaf it is essential to obtain cross sections throughout leaf development and at maturity (Kalve et al., 2014). Here, we describe a protocol for transverse sectioning of Arabidopsis thaliana leaves using resin embedding. This protocol provides a reliable platform to yield high quality images of cross sections allowing study of development of various tissue layers across the transversal axis of the leaf. As this method is an adaptation of the protocol developed for the Arabidopsis root tip by Viane (1999) andDe Smet et al. (2004), it can easily be modified to accommodate other organs and species. Materials and Reagents

Summary Cellular and genetic understanding of the rice leaf size regulation is limited, despite rice being the staple food of more than half of the global population. We investigated the mechanism controlling the rice leaf length using cultivated and wild rice accessions that remarkably differed for leaf size. Comparative transcriptomics, gibberellic acid (GA) quantification and leaf kinematics of the contrasting accessions suggested the involvement of GA, cell cycle and growth‐regulating factors (GRFs) in the rice leaf size regulation. Zone‐specific expression analysis and VIGS established the functions of specific GRFs in the process. The leaf length of the selected accessions was strongly correlated with GA levels. Higher GA content in wild rice accessions with longer leaves and GA‐induced increase in the leaf length via an increase in cell division confirmed a GA‐mediated regulation of division zone in rice. Downstream to GA, OsGRF7 and OsGRF8 function for controlling cell division to determine the rice leaf length. Spatial control of cell division to determine the division zone size mediated by GA and downstream OsGRF7 and OsGRF8 explains the leaf length differences between the cultivated and wild rice. This mechanism to control the rice leaf length might have contributed to optimizing leaf size during domestication.

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