Two phylogenetically unrelated peptide-receptor modules jointly regulate lateral root initiation via a partially shared signaling pathway in Arabidopsis thaliana.

Peptide-receptor signaling is an important system for intercellular communication, regulating many developmental processes. A single process can be controlled by several distinct signaling peptides. However, since peptide-receptor modules are usually studied separately, their mechanistic interactions remain largely unexplored. Two phylogenetically unrelated peptide-receptor modules, GLV6/GLV10-RGI and TOLS2/PIP2-RLK7, independently described as inhibitors of lateral root initiation, show striking similarities between their expression patterns and gain- and loss-of-function phenotypes, suggesting a common function during lateral root spacing and initiation. The GLV6/GLV10-RGI and TOLS2/PIP2-RLK7 modules trigger similar transcriptional changes, likely in part via WRKY transcription factors. Their overlapping set of response genes includes PUCHI and PLT5, both required for the effect of GLV6/10, as well as TOLS2, on lateral root initiation. Furthermore, both modules require the activity of MPK6 and can independently trigger MPK3/MPK6 phosphorylation. The GLV6/10 and TOLS2/PIP2 signaling pathways seem to converge in the activation of MPK3/MPK6, leading to the induction of a similar transcriptional response in the same target cells, thereby regulating lateral root initiation through a (partially) common mechanism. Convergence of signaling pathways downstream of phylogenetically unrelated peptide-receptor modules adds an additional, and hitherto unrecognized, level of complexity to intercellular communication networks in plants.

Receptor kinase module targets PIN-dependent auxin transport during canalization.

Spontaneously arising channels that transport the phytohormone auxin provide positional cues for self-organizing aspects of plant development such as flexible vasculature regeneration or its patterning during leaf venation. The auxin canalization hypothesis proposes a feedback between auxin signaling and transport as the underlying mechanism, but molecular players await discovery. We identified part of the machinery that routes auxin transport. The auxin-regulated receptor CAMEL (Canalization-related Auxin-regulated Malectin-type RLK) together with CANAR (Canalization-related Receptor-like kinase) interact with and phosphorylate PIN auxin transporters. camel and canar mutants are impaired in PIN1 subcellular trafficking and auxin-mediated PIN polarization, which macroscopically manifests as defects in leaf venation and vasculature regeneration after wounding. The CAMEL-CANAR receptor complex is part of the auxin feedback that coordinates polarization of individual cells during auxin canalization.

Maternal auxin supply contributes to early embryo patterning in Arabidopsis.

The angiosperm seed is composed of three genetically distinct tissues: the diploid embryo that originates from the fertilized egg cell, the triploid endosperm that is produced from the fertilized central cell, and the maternal sporophytic integuments that develop into the seed coat1. At the onset of embryo development in Arabidopsis thaliana, the zygote divides asymmetrically, producing a small apical embryonic cell and a larger basal cell that connects the embryo to the maternal tissue2. The coordinated and synchronous development of the embryo and the surrounding integuments, and the alignment of their growth axes, suggest communication between maternal tissues and the embryo. In contrast to animals, however, where a network of maternal factors that direct embryo patterning have been identified3,4, only a few maternal mutations have been described to affect embryo development in plants5-7. Early embryo patterning in Arabidopsis requires accumulation of the phytohormone auxin in the apical cell by directed transport from the suspensor8-10. However, the origin of this auxin has remained obscure. Here we investigate the source of auxin for early embryogenesis and provide evidence that the mother plant coordinates seed development by supplying auxin to the early embryo from the integuments of the ovule. We show that auxin response increases in ovules after fertilization, due to upregulated auxin biosynthesis in the integuments, and this maternally produced auxin is required for correct embryo development.

WRKY23 is a component of the transcriptional network mediating auxin feedback on PIN polarity.

Auxin is unique among plant hormones due to its directional transport that is mediated by the polarly distributed PIN auxin transporters at the plasma membrane. The canalization hypothesis proposes that the auxin feedback on its polar flow is a crucial, plant-specific mechanism mediating multiple self-organizing developmental processes. Here, we used the auxin effect on the PIN polar localization in Arabidopsis thaliana roots as a proxy for the auxin feedback on the PIN polarity during canalization. We performed microarray experiments to find regulators of this process that act downstream of auxin. We identified genes that were transcriptionally regulated by auxin in an AXR3/IAA17- and ARF7/ARF19-dependent manner. Besides the known components of the PIN polarity, such as PID and PIP5K kinases, a number of potential new regulators were detected, among which the WRKY23 transcription factor, which was characterized in more detail. Gain- and loss-of-function mutants confirmed a role for WRKY23 in mediating the auxin effect on the PIN polarity. Accordingly, processes requiring auxin-mediated PIN polarity rearrangements, such as vascular tissue development during leaf venation, showed a higher WRKY23 expression and required the WRKY23 activity. Our results provide initial insights into the auxin transcriptional network acting upstream of PIN polarization and, potentially, canalization-mediated plant development.

De-Problematizing 'GMOs': Suggestions for Communicating about Genetic Engineering.

The public debates concerning genetic engineering (GE) involve many non-scientific issues. The ensuing complexity is one reason why biotechnologists are reluctant to become involved. By sharing our personal experiences in science communication and suggesting ways to de-problematize GE, we aim to inspire our colleagues to engage with the public.

Plant embryogenesis requires AUX/LAX-mediated auxin influx.

The plant hormone auxin and its directional transport are known to play a crucial role in defining the embryonic axis and subsequent development of the body plan. Although the role of PIN auxin efflux transporters has been clearly assigned during embryonic shoot and root specification, the role of the auxin influx carriers AUX1 and LIKE-AUX1 (LAX) proteins is not well established. Here, we used chemical and genetic tools on Brassica napus microspore-derived embryos and Arabidopsis thaliana zygotic embryos, and demonstrate that AUX1, LAX1 and LAX2 are required for both shoot and root pole formation, in concert with PIN efflux carriers. Furthermore, we uncovered a positive-feedback loop between MONOPTEROS (ARF5)-dependent auxin signalling and auxin transport. This MONOPTEROS-dependent transcriptional regulation of auxin influx (AUX1, LAX1 and LAX2) and auxin efflux (PIN1 and PIN4) carriers by MONOPTEROS helps to maintain proper auxin transport to the root tip. These results indicate that auxin-dependent cell specification during embryo development requires balanced auxin transport involving both influx and efflux mechanisms, and that this transport is maintained by a positive transcriptional feedback on auxin signalling.

Tightly controlled WRKY23 expression mediates Arabidopsis embryo development.

The development of a multicellular embryo from a single zygote is a complex and highly organized process that is far from understood. In higher plants, apical-basal patterning mechanisms are crucial to correctly specify root and shoot stem cell niches that will sustain and drive post-embryonic plant growth and development. The auxin-responsive AtWRKY23 transcription factor is expressed from early embryogenesis onwards and the timing and localization of its expression overlaps with the root stem cell niche. Knocking down WRKY23 transcript levels or expression of a dominant-negative WRKY23 version via a translational fusion with the SRDX repressor domain affected both apical-basal axis formation as well as installation of the root stem cell niche. WRKY23 expression is affected by two well-known root stem cell specification mechanisms, that is, SHORTROOT and MONOPTEROS-BODENLOS signalling and can partially rescue the root-forming inability of mp embryos. On the basis of these results, we postulate that a tightly controlled WRKY23 expression is involved in the regulation of both auxin-dependent and auxin-independent signalling pathways towards stem cell specification.

SCF(TIR1/AFB)-auxin signalling regulates PIN vacuolar trafficking and auxin fluxes during root gravitropism.

The distribution of the phytohormone auxin regulates many aspects of plant development including growth response to gravity. Gravitropic root curvature involves coordinated and asymmetric cell elongation between the lower and upper side of the root, mediated by differential cellular auxin levels. The asymmetry in the auxin distribution is established and maintained by a spatio-temporal regulation of the PIN-FORMED (PIN) auxin transporter activity. We provide novel insights into the complex regulation of PIN abundance and activity during root gravitropism. We show that PIN2 turnover is differentially regulated on the upper and lower side of gravistimulated roots by distinct but partially overlapping auxin feedback mechanisms. In addition to regulating transcription and clathrin-mediated internalization, auxin also controls PIN abundance at the plasma membrane by promoting their vacuolar targeting and degradation. This effect of elevated auxin levels requires the activity of SKP-Cullin-F-box(TIR1/AFB) (SCF(TIR1/AFB))-dependent pathway. Importantly, also suboptimal auxin levels mediate PIN degradation utilizing the same signalling pathway. These feedback mechanisms are functionally important during gravitropic response and ensure fine-tuning of auxin fluxes for maintaining as well as terminating asymmetric growth.

Spatiotemporal regulation of lateral root organogenesis in Arabidopsis by cytokinin.

The architecture of a plant's root system, established postembryonically, results from both coordinated root growth and lateral root branching. The plant hormones auxin and cytokinin are central endogenous signaling molecules that regulate lateral root organogenesis positively and negatively, respectively. Tight control and mutual balance of their antagonistic activities are particularly important during the early phases of lateral root organogenesis to ensure continuous lateral root initiation (LRI) and proper development of lateral root primordia (LRP). Here, we show that the early phases of lateral root organogenesis, including priming and initiation, take place in root zones with a repressed cytokinin response. Accordingly, ectopic overproduction of cytokinin in the root basal meristem most efficiently inhibits LRI. Enhanced cytokinin responses in pericycle cells between existing LRP might restrict LRI near existing LRP and, when compromised, ectopic LRI occurs. Furthermore, our results demonstrate that young LRP are more sensitive to perturbations in the cytokinin activity than are developmentally more advanced primordia. We hypothesize that the effect of cytokinin on the development of primordia possibly depends on the robustness and stability of the auxin gradient.

Cell polarity: stretching prevents developmental cramps.

Initiation and successive development of organs induce mechanical stresses at the cellular level. Using the tomato shoot apex, a new study now proposes that mechanical strain regulates the plasma membrane abundance of the PIN1 auxin transporter, thereby reinforcing a positive feed-back loop between growth and auxin accumulation.

Transcription factor WRKY23 assists auxin distribution patterns during Arabidopsis root development through local control on flavonol biosynthesis.

Gradients of the plant hormone auxin, which depend on its active intercellular transport, are crucial for the maintenance of root meristematic activity. This directional transport is largely orchestrated by a complex interaction of specific influx and efflux carriers that mediate the auxin flow into and out of cells, respectively. Besides these transport proteins, plant-specific polyphenolic compounds known as flavonols have been shown to act as endogenous regulators of auxin transport. However, only limited information is available on how flavonol synthesis is developmentally regulated. Using reduction-of-function and overexpression approaches in parallel, we demonstrate that the WRKY23 transcription factor is needed for proper root growth and development by stimulating the local biosynthesis of flavonols. The expression of WRKY23 itself is controlled by auxin through the Auxin Response Factor 7 (ARF7) and ARF19 transcriptional response pathway. Our results suggest a model in which WRKY23 is part of a transcriptional feedback loop of auxin on its own transport through local regulation of flavonol biosynthesis.

Arabidopsis α Aurora kinases function in formative cell division plane orientation.

To establish three-dimensional structures/organs, plant cells continuously have to adapt the orientation of their division plane in a highly regulated manner. However, mechanisms underlying switches in division plane orientation remain elusive. Here, we characterize a viable double knockdown mutant in Arabidopsis thaliana group α Aurora (AUR) kinases, AUR1 and AUR2, (aur1-2 aur2-2), with a primary defect in lateral root formation and outgrowth. Mutant analysis revealed that aur1-2 aur2-2 lateral root primordia are built from randomly oriented cell divisions instead of distinct cell layers. This phenotype could be traced back to cytokinesis defects and misoriented cell plates during the initial anticlinal pericycle cell divisions that give rise to lateral root primordia. Complementation assays showed that the Arabidopsis α group Aurora kinases are functionally divergent from the single ß group member AUR3 and that AUR1 functions in division plane orientation prior to cytokinesis. In addition to defective lateral root patterning, aur1-2 aur2-2 plants also show defects in orienting formative divisions during embryogenesis, divisions surrounding the main root stem cell niche, and divisions surrounding stomata formation. Taken together, our results put forward a central role for α Aurora kinases in regulating formative division plane orientation throughout development.

A novel aux/IAA28 signaling cascade activates GATA23-dependent specification of lateral root founder cell identity.

BACKGROUND: Lateral roots are formed at regular intervals along the main root by recurrent specification of founder cells. To date, the mechanism by which branching of the root system is controlled and founder cells become specified remains unknown. RESULTS: Our study reports the identification of the auxin regulatory components and their target gene, GATA23, which control lateral root founder cell specification. Initially, a meta-analysis of lateral root-related transcriptomic data identified the GATA23 transcription factor. GATA23 is expressed specifically in xylem pole pericycle cells before the first asymmetric division and is correlated with oscillating auxin signaling maxima in the basal meristem. Also, functional studies revealed that GATA23 controls lateral root founder cell identity. Finally, we show that an Aux/IAA28-dependent auxin signaling mechanism in the basal meristem controls GATA23 expression. CONCLUSIONS: We have identified the first molecular components that control lateral root founder cell identity in the Arabidopsis root. These include an IAA28-dependent auxin signaling module in the basal meristem region that regulates GATA23 expression and thereby lateral root founder cell specification and root branching patterns.

The march of the PINs: developmental plasticity by dynamic polar targeting in plant cells.

Development of plants and their adaptive capacity towards ever-changing environmental conditions largely depend on the spatial distribution of the plant hormone auxin. At the cellular level, various internal and external signals are translated into specific changes in the polar, subcellular localization of auxin transporters from the PIN family thereby directing and redirecting the intercellular fluxes of auxin. The current model of polar targeting of PIN proteins towards different plasma membrane domains encompasses apolar secretion of newly synthesized PINs followed by endocytosis and recycling back to the plasma membrane in a polarized manner. In this review, we follow the subcellular march of the PINs and highlight the cellular and molecular mechanisms behind polar foraging and subcellular trafficking pathways. Also, the entry points for different signals and regulations including by auxin itself will be discussed within the context of morphological and developmental consequences of polar targeting and subcellular trafficking.

NINJA connects the co-repressor TOPLESS to jasmonate signalling.

Jasmonoyl-isoleucine (JA-Ile) is a plant hormone that regulates a broad array of plant defence and developmental processes. JA-Ile-responsive gene expression is regulated by the transcriptional activator MYC2 that interacts physically with the jasmonate ZIM-domain (JAZ) repressor proteins. On perception of JA-Ile, JAZ proteins are degraded and JA-Ile-dependent gene expression is activated. The molecular mechanisms by which JAZ proteins repress gene expression remain unknown. Here we show that the Arabidopsis JAZ proteins recruit the Groucho/Tup1-type co-repressor TOPLESS (TPL) and TPL-related proteins (TPRs) through a previously uncharacterized adaptor protein, designated Novel Interactor of JAZ (NINJA). NINJA acts as a transcriptional repressor whose activity is mediated by a functional TPL-binding EAR repression motif. Accordingly, both NINJA and TPL proteins function as negative regulators of jasmonate responses. Our results point to TPL proteins as general co-repressors that affect multiple signalling pathways through the interaction with specific adaptor proteins. This new insight reveals how stress-related and growth-related signalling cascades use common molecular mechanisms to regulate gene expression in plants.

Bimodular auxin response controls organogenesis in Arabidopsis.

Like animals, the mature plant body develops via successive sets of instructions that determine cell fate, patterning, and organogenesis. In the coordination of various developmental programs, several plant hormones play decisive roles, among which auxin is the best-documented hormonal signal. Despite the broad range of processes influenced by auxin, how such a single signaling molecule can be translated into a multitude of distinct responses remains unclear. In Arabidopsis thaliana, lateral root development is a classic example of a developmental process that is controlled by auxin at multiple stages. Therefore, we used lateral root formation as a model system to gain insight into the multifunctionality of auxin. We were able to demonstrate the complementary and sequential action of two discrete auxin response modules, the previously described Solitary Root/indole-3-Acetic Acid (IAA)14-Auxin Response Factor (ARF)7-ARF19-dependent lateral root initiation module and the successive Bodenlos/IAA12-Monopteros/ARF5-dependent module, both of which are required for proper organogenesis. The genetic framework in which two successive auxin response modules control early steps of a developmental process adds an extra dimension to the complexity of auxin's action.

Manipulation of auxin transport in plant roots during Rhizobium symbiosis and nematode parasitism.

The plant rhizosphere harbors many different microorganisms, ranging from plant growth-promoting bacteria to devastating plant parasites. Some of these microbes are able to induce de novo organ formation in infected roots. Certain soil bacteria, collectively called rhizobia, form a symbiotic interaction with legumes, leading to the formation of nitrogen-fixing root nodules. Sedentary endoparasitic nematodes, on the other hand, induce highly specialized feeding sites in infected plant roots from which they withdraw nutrients. In order to establish these new root structures, it is thought that these organisms use and manipulate the endogenous molecular and physiological pathways of their hosts. Over the years, evidence has accumulated reliably demonstrating the involvement of the plant hormone auxin. Moreover, the auxin responses during microbe-induced de novo organ formation seem to be dynamic, suggesting that plant-associated microbes can actively modify their host's auxin transport. In this review, we focus on recent findings in auxin transport mechanisms during plant development and on how plant symbionts and parasites have evolved to manipulate these mechanisms for their own purposes.