Cytokinin regulates apical hook development via the coordinated actions of EIN3/EIL1 and PIF transcription factors in Arabidopsis.

The apical hook is indispensable for protecting the delicate shoot apical meristem while dicot seedlings emerge from soil after germination in darkness. The development of the apical hook is co-ordinately regulated by multiple phytohormones and environmental factors. Yet, a holistic understanding of the spatial-temporal interactions between different phytohormones and environmental factors remains to be achieved. Using a chemical genetic approach, we identified kinetin riboside, as a proxy of kinetin, which promotes apical hook development of Arabidopsis thaliana in a partially ethylene-signaling-independent pathway. Further genetic and biochemical analysis revealed that cytokinin is able to regulate apical hook development via post-transcriptional regulation of the PHYTOCHROME INTERACTING FACTORs (PIFs), together with its canonical roles in inducing ethylene biosynthesis. Dynamic observations of apical hook development processes showed that ETHYLENE INSENSITVE3 (EIN3) and EIN3-LIKE1 (EIL1) are necessary for the exaggeration of hook curvature in response to cytokinin, while PIFs are crucial for the cytokinin-induced maintenance of hook curvature in darkness. Furthermore, these two families of transcription factors display divergent roles in light-triggered hook opening. Our findings reveal that cytokinin integrates ethylene signaling and light signaling via EIN3/EIL1 and PIFs, respectively, to dynamically regulate apical hook development during early seedling development.

New Wine in an Old Bottle: Utilizing Chemical Genetics to Dissect Apical Hook Development.

The apical hook is formed by dicot seedlings to protect the tender shoot apical meristem during soil emergence. Regulated by many phytohormones, the apical hook has been taken as a model to study the crosstalk between individual signaling pathways. Over recent decades, the roles of different phytohormones and environmental signals in apical hook development have been illustrated. However, key regulators downstream of canonical hormone signaling have rarely been identified via classical genetics screening, possibly due to genetic redundancy and/or lethal mutation. Chemical genetics that utilize small molecules to perturb and elucidate biological processes could provide a complementary strategy to overcome the limitations in classical genetics. In this review, we summarize current progress in hormonal regulation of the apical hook, and previously reported chemical tools that could assist the understanding of this complex developmental process. We also provide insight into novel strategies for chemical screening and target identification, which could possibly lead to discoveries of new regulatory components in apical hook development, or unidentified signaling crosstalk that is overlooked by classical genetics screening.

Interplay between Cell Wall and Auxin Mediates the Control of Differential Cell Elongation during Apical Hook Development.

Differential growth plays a crucial role during morphogenesis [1-3]. In plants, development occurs within mechanically connected tissues, and local differences in cell expansion lead to deformations at the organ level, such as buckling or bending [4, 5]. During early seedling development, bending of hypocotyl by differential cell elongation results in apical hook structure that protects the shoot apical meristem from being damaged during emergence from the soil [6, 7]. Plant hormones participate in apical hook development, but not how they mechanistically drive differential growth [8]. Here, we present evidence of interplay between hormonal signals and cell wall in auxin-mediated differential cell elongation using apical hook development as an experimental model. Using genetic and cell biological approaches, we show that xyloglucan (a major primary cell wall component) mediates asymmetric mechanical properties of epidermal cells required for hook development. The xxt1 xxt2 mutant, deficient in xyloglucan [9], displays severe defects in differential cell elongation and hook development. Analysis of xxt1 xxt2 mutant reveals a link between cell wall and transcriptional control of auxin transporters PINFORMEDs (PINs) and AUX1 crucial for establishing the auxin response maxima required for preferential repression of elongation of the cells on the inner side of the hook. Genetic evidence identifies auxin response factor ARF2 as a negative regulator acting downstream of xyloglucan-dependent control of hook development and transcriptional control of polar auxin transport. Our results reveal a crucial feedback process between the cell wall and transcriptional control of polar auxin transport, underlying auxin-dependent control of differential cell elongation in plants.

SAUR53 regulates organ elongation and apical hook development in Arabidopsis.

SAUR53 is a member of SAUR (Small Auxin-Up RNA) gene family of primary auxin responsive genes. In Arabidopsis, SAUR gene family is represented by 81 genes including two pseudogenes; however, the functions of most of these genes are not fully characterized yet. In the present study, we show that SAUR53 expresses throughout the plant and localizes to both plasma membrane and the nucleus. Unlike most other SAUR genes, expression of SAUR53 is not induced in response to auxin. Ectopic expression of SAUR53 results in the elongation of cells and organs, and also interferes with normal apical hook development by accelerating the hook maintenance phase. Moreover, root growth of SAUR53 overexpression seedlings is significantly insensitive to IAA and 2,4-D, while showing wild-type sensitivity to NAA, suggesting that elevated level of SAUR53 may interfere with normal auxin transport. Collectively, this study indicates that while SAUR53 positively regulates cell and organ elongation, it probably negatively regulates auxin transport in Arabidopsis.

Real-Time Analysis of the Apical Hook Development.

Mechanisms for cell protection are essential for survival of multicellular organisms. In plants, the apical hook, which is transiently formed in darkness when the germinating seedling penetrates towards the soil surface, plays such protective role and shields the vitally important shoot apical meristem and cotyledons from damage. The apical hook is formed by bending of the upper hypocotyl soon after germination, and it is maintained in a closed stage while the hypocotyl continues to penetrate through the soil and rapidly opens when exposed to light in proximity of the soil surface. To uncover the complex molecular network orchestrating this spatiotemporally tightly coordinated process, monitoring of the apical hook development in real time is indispensable. Here we describe an imaging platform that enables high-resolution kinetic analysis of this dynamic developmental process.

A current perspective on the role of AGCVIII kinases in PIN-mediated apical hook development.

Despite their sessile lifestyle, seed plants are able to utilize differential growth rates to move their organs in response to their environment. Asymmetrical growth is the cause for the formation and maintenance of the apical hook-a structure of dicotyledonous plants shaped by the bended hypocotyl that eases the penetration through the covering soil. As predicted by the Cholodny-Went theory, the cause for differential growth is the unequal distribution of the phytohormone auxin. The PIN-FORMED proteins transport auxin from cell-to-cell and control the distribution of auxin in the plant. Their localization and activity are regulated by two subfamilies of AGCVIII protein kinases: the D6 PROTEIN KINASEs as well as PINOID and its two closely related WAG kinases. This mini-review focuses on the regulatory mechanism of these AGCVIII kinases as well as their role in apical hook development of Arabidopsis thaliana.