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The maintenance and reformation of gene expression domains are the basis for the morphogenic processes of multicellular systems. In a leaf primordium of Arabidopsis thaliana, the expression of FILAMENTOUS FLOWER (FIL) and the activity of the microRNA miR165/166 are specific to the abaxial side. This miR165/166 activity restricts the target gene expression to the adaxial side. The adaxial and abaxial specific gene expressions are crucial for the wide expansion of leaf lamina. The FIL-expression and the miR165/166-free domains are almost mutually exclusive, and they have been considered to be maintained during leaf development. However, we found here that the position of the boundary between the two domains gradually shifts from the adaxial side to the abaxial side. The cell lineage analysis revealed that this boundary shifting was associated with a sequential gene expression switch from the FIL-expressing (miR165/166 active) to the miR165/166-free (non-FIL-expressing) states. Our genetic analyses using the enlarged fil expression domain2 (enf2) mutant and chemical treatment experiments revealed that impairment in the plastid (chloroplast) gene expression machinery retards this boundary shifting and inhibits the lamina expansion. Furthermore, these developmental effects caused by the abnormal plastids were not observed in the genomes uncoupled1 (gun1) mutant background. This study characterizes the dynamic nature of the adaxial-abaxial specification process in leaf primordia and reveals that the dynamic process is affected by the GUN1-dependent retrograde signal in response to the failure of plastid gene expression. These findings advance our understanding on the molecular mechanism linking the plastid function to the leaf morphogenic processes. [ABSTRACT FROM AUTHOR]

Auxin response factor (ARF) family genes play a central role in controlling sensitivity to the plant hormone auxin. We characterized the function ofARF8 inArabidopsisby investigating a T-DNA insertion line (arf8-1) and overexpression lines (ARF8OX) ofARF8.arf8-1showed a long-hypocotyl phenotype in either white, blue, red or far-red light conditions, in contrast toARF8OX that displayed short hypocotyls in the light. Stronger and weaker apical dominance, and promotion and inhibition of lateral root formation were observed inarf8-1andARF8OX respectively. Sensitivity to auxin was unaltered inarf8-1hypocotyls with respect to growth inhibition caused by exogenously applied auxin and growth promotion induced by higher temperatures.ARF8expression was observed constitutively in shoot and root apexes, and was induced in the light condition in hypocotyls. Free IAA contents were approximately 30% reduced in light-grown hypocotyls ofARF8OX, but were similar between those ofarf8-1and wild type. Expression of the threeGH3genes was reduced inarf8-1and increased inARF8OX, indicating that they are targets of ARF8 transcriptional control. Because the three GH3 proteins may be involved in the conjugation of IAA as suggested by, and because two of the threeGH3genes are auxin inducible, ARF8 may control the free IAA level in a negative feedback fashion by regulatingGH3gene expression.ARFfamily genes seem to control both auxin sensitivity and homeostasis inArabidopsis. [ABSTRACT FROM AUTHOR]

Plant survival depends on seed germination and progression through post-germinative developmental checkpoints. These processes are controlled by the stress phytohormone abscisic acid (ABA). ABA regulates the basic leucine zipper transcriptional factor ABI5, a central hub of growth repression, while the reactive nitrogen molecule nitric oxide (NO) counteracts ABA during seed germination. However, the molecular mechanisms by which seeds sense more favourable conditions and start germinating have remained elusive. Here we show that ABI5 promotes growth via NO, and that ABI5 accumulation is altered in genetic backgrounds with impaired NO homeostasis. S-nitrosylation of ABI5 at cysteine-153 facilitates its degradation through CULLIN4-based and KEEP ON GOING E3 ligases, and promotes seed germination. Conversely, mutation of ABI5 at cysteine-153 deregulates protein stability and inhibition of seed germination by NO depletion. These findings suggest an inverse molecular link between NO and ABA hormone signalling through distinct posttranslational modifications of ABI5 during early seedling development. [ABSTRACT FROM AUTHOR]

Multicellular organisms rely on intercellular communication systems to organize their cellular functions. In studies focusing on intercellular communication, the key experimental techniques include the generation of chimeric tissue using transgenic DNA recombination systems represented by the CRE/ loxP system. If an experimental system enables the induction of chimeras at highly targeted cell(s), it will facilitate the reproducibility and precision of experiments. However, multiple technical limitations have made this challenging. The stochastic nature of DNA recombination events, especially, hampers reproducible generation of intended chimeric patterns. Infrared laser-evoked gene operator (IR-LEGO), a microscopic system that irradiates targeted cells using an IR laser, can induce heat shock-mediated expression of transgenes, for example, CRE recombinase gene, in the cells. In this study, we developed a method that induces CRE/ loxP recombination in the target cell(s) of plant roots and leaves in a highly specific manner. We combined IR-LEGO, an improved heat-shock-specific promoter, and dexamethasone-dependent regulation of CRE. The optimal IR-laser power and irradiation duration were estimated via exhaustive irradiation trials and subsequent statistical modeling. Under optimized conditions, CRE/ loxP recombination was efficiently induced without cellular damage. We also found that the induction efficiency varied among tissue types and cellular sizes. The developed method offers an experimental system to generate a precisely designed chimeric tissue, and thus, will be useful for analyzing intercellular communication at high resolution in roots and leaves., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Tomoi, Tameshige, Betsuyaku, Hamada, Sakamoto, Uchida, Torii, Shimizu, Tamada, Urawa, Okada, Fukuda, Tatematsu, Kamei and Betsuyaku.)