Spatiotemporal control of cell growth by CUC3 shapes leaf margins.

How a shape arises from the coordinated behavior of cells is one of the most fascinating questions in developmental biology. In plants, fine spatial and temporal controls of cell proliferation and cell expansion sustain differential growth that defines organ shape and size. At the leaf margin of Arabidopsis thaliana, interplay between auxin transport and transcription factors named CUP SHAPED COTYLEDON (CUCs), which are involved in the establishment of boundary domain identity, were reported to trigger differential growth, leading to serration. Cellular behaviors behind these differential growths remain scarcely described. Here, we used 3D and time lapse imaging on young leaves at different stages of development to determine the sequence of cellular events resulting in leaf serrations. In addition, we showed that the transcription factor CUC3 is a negative regulator of cell growth and that its expression dynamics in a small number of cells at the leaf margin is tightly associated with the control of differential growth.

Heterogeneity and its multiscale integration in plant morphogenesis.

Heterogeneity is observed at all levels in living organisms, but its role during the development of an individual is not well understood. Heterogeneity has either to be limited to ensure robust development or can be an actor of the biological processes leading to reproducible development. Here we review the sources of heterogeneity in plants, stress the interplay between noise in elementary processes and regulated biological mechanisms, and highlight how heterogeneity is integrated at multiple scales during plant morphogenesis.

Strong morphological defects in conditional Arabidopsis abp1 knock-down mutants generated in absence of functional ABP1 protein.

The Auxin Binding Protein 1 (ABP1) is one of the most studied proteins in plants. Since decades ago, it has been the prime receptor candidate for the plant hormone auxin with a plethora of described functions in auxin signaling and development. The developmental importance of ABP1 has recently been questioned by identification of Arabidopsis thaliana abp1 knock-out alleles that show no obvious phenotypes under normal growth conditions. In this study, we examined the contradiction between the normal growth and development of the abp1 knock-outs and the strong morphological defects observed in three different ethanol-inducible abp1 knock-down mutants ( abp1-AS, SS12K, SS12S). By analyzing segregating populations of abp1 knock-out vs. abp1 knock-down crosses we show that the strong morphological defects that were believed to be the result of conditional down-regulation of ABP1 can be reproduced also in the absence of the functional ABP1 protein. This data suggests that the phenotypes in  abp1 knock-down lines are due to the off-target effects and asks for further reflections on the biological function of ABP1 or alternative explanations for the missing phenotypic defects in the abp1 loss-of-function alleles.

Inhibition of cell expansion by rapid ABP1-mediated auxin effect on microtubules.

The prominent and evolutionarily ancient role of the plant hormone auxin is the regulation of cell expansion. Cell expansion requires ordered arrangement of the cytoskeleton but molecular mechanisms underlying its regulation by signalling molecules including auxin are unknown. Here we show in the model plant Arabidopsis thaliana that in elongating cells exogenous application of auxin or redistribution of endogenous auxin induces very rapid microtubule re-orientation from transverse to longitudinal, coherent with the inhibition of cell expansion. This fast auxin effect requires auxin binding protein 1 (ABP1) and involves a contribution of downstream signalling components such as ROP6 GTPase, ROP-interactive protein RIC1 and the microtubule-severing protein katanin. These components are required for rapid auxin- and ABP1-mediated re-orientation of microtubules to regulate cell elongation in roots and dark-grown hypocotyls as well as asymmetric growth during gravitropic responses.

AUXIN BINDING PROTEIN1 links cell wall remodeling, auxin signaling, and cell expansion in arabidopsis.

Cell expansion is an increase in cell size and thus plays an essential role in plant growth and development. Phytohormones and the primary plant cell wall play major roles in the complex process of cell expansion. In shoot tissues, cell expansion requires the auxin receptor AUXIN BINDING PROTEIN1 (ABP1), but the mechanism by which ABP1 affects expansion remains unknown. We analyzed the effect of functional inactivation of ABP1 on transcriptomic changes in dark-grown hypocotyls and investigated the consequences of gene expression on cell wall composition and cell expansion. Molecular and genetic evidence indicates that ABP1 affects the expression of a broad range of cell wall-related genes, especially cell wall remodeling genes, mainly via an SCF(TIR/AFB)-dependent pathway. ABP1 also functions in the modulation of hemicellulose xyloglucan structure. Furthermore, fucosidase-mediated defucosylation of xyloglucan, but not biosynthesis of nonfucosylated xyloglucan, rescued dark-grown hypocotyl lengthening of ABP1 knockdown seedlings. In muro remodeling of xyloglucan side chains via an ABP1-dependent pathway appears to be of critical importance for temporal and spatial control of cell expansion.

Auxin-binding protein 1 is a negative regulator of the SCF(TIR1/AFB) pathway.

Auxin is a major plant hormone that controls most aspects of plant growth and development. Auxin is perceived by two distinct classes of receptors: transport inhibitor response 1 (TIR1, or auxin-related F-box (AFB)) and auxin/indole-3-acetic acid (AUX/IAA) coreceptors, that control transcriptional responses to auxin, and the auxin-binding protein 1 (ABP1), that controls a wide variety of growth and developmental processes. To date, the mode of action of ABP1 is still poorly understood and its functional interaction with TIR1/AFB-AUX/IAA coreceptors remains elusive. Here we combine genetic and biochemical approaches to gain insight into the integration of these two pathways. We find that ABP1 is genetically upstream of TIR1/AFBs; ABP1 knockdown leads to an enhanced degradation of AUX/IAA repressors, independently of its effects on endocytosis, through the SCF(TIR1/AFB) E3 ubiquitin ligase pathway. Combining positive and negative regulation of SCF ubiquitin-dependent pathways might be a common mechanism conferring tight control of hormone-mediated responses.

AUXIN RESPONSE FACTOR8 regulates Arabidopsis petal growth by interacting with the bHLH transcription factor BIGPETALp.

Plant organ growth and final size are determined by coordinated cell proliferation and expansion. The BIGPETALp (BPEp) basic helix-loop-helix (bHLH) transcription factor was shown to limit Arabidopsis thaliana petal growth by influencing cell expansion. We demonstrate here that BPEp interacts with AUXIN RESPONSE FACTOR8 (ARF8) to affect petal growth. This interaction is mediated through the BPEp C-terminal domain (SD(BPEp)) and the C-terminal domain of ARF8. Site-directed mutagenesis identified an amino acid consensus motif in SD(BPEp) that is critical for mediating BPEp-ARF8 interaction. This motif shares sequence similarity with motif III of ARF and AUXIN/INDOLE-3-ACETIC ACID proteins. Petals of arf8 mutants are significantly larger than those of the wild type due to increased cell number and increased cell expansion. bpe arf8 double mutant analyses show that during early petal development stages, ARF8 and BPEp work synergistically to limit mitotic growth. During late stages, ARF8 and BPEp interact to limit cell expansion. The alterations in cell division and cell expansion observed in arf8 and/or bpe mutants are associated with a change in expression of early auxin-responsive genes. The data provide evidence of an interaction between an ARF and a bHLH transcription factor and of its biological significance in regulating petal growth, with local auxin levels likely influencing such a biological function.

Cell surface- and rho GTPase-based auxin signaling controls cellular interdigitation in Arabidopsis.

Auxin is a multifunctional hormone essential for plant development and pattern formation. A nuclear auxin-signaling system controlling auxin-induced gene expression is well established, but cytoplasmic auxin signaling, as in its coordination of cell polarization, is unexplored. We found a cytoplasmic auxin-signaling mechanism that modulates the interdigitated growth of Arabidopsis leaf epidermal pavement cells (PCs), which develop interdigitated lobes and indentations to form a puzzle-piece shape in a two-dimensional plane. PC interdigitation is compromised in leaves deficient in either auxin biosynthesis or its export mediated by PINFORMED 1 localized at the lobe tip. Auxin coordinately activates two Rho GTPases, ROP2 and ROP6, which promote the formation of complementary lobes and indentations, respectively. Activation of these ROPs by auxin occurs within 30 s and depends on AUXIN-BINDING PROTEIN 1. These findings reveal Rho GTPase-based auxin-signaling mechanisms, which modulate the spatial coordination of cell expansion across a field of cells.

AUXIN BINDING PROTEIN 1: functional and evolutionary aspects.

In this review, we examine the role of AUXIN BINDING PROTEIN 1 (ABP1) in mediating growth and developmental responses. ABP1 is involved in a broad range of cellular responses to auxin, acting either as the main regulator of the response, such as seen for entry into cell division or, as a fine-tuning device as for the regulation of expression of early auxin response genes. Phylogenetic analysis has revealed that ABP1 is an ancient protein that was already present in various algae and has acquired a motif of retention in the endoplasmic reticulum only recently. An evaluation of the evidence for ABP1 function according to its cellular localization supports the plasma membrane as a starting point for ABP1-mediated auxin signaling.

The RNA binding protein Tudor-SN is essential for stress tolerance and stabilizes levels of stress-responsive mRNAs encoding secreted proteins in Arabidopsis.

Tudor-SN (TSN) copurifies with the RNA-induced silencing complex in animal cells where, among other functions, it is thought to act on mRNA stability via the degradation of specific dsRNA templates. In plants, TSN has been identified biochemically as a cytoskeleton-associated RNA binding activity. In eukaryotes, it has recently been identified as a conserved primary target of programmed cell death-associated proteolysis. We have investigated the physiological role of TSN by isolating null mutations for two homologous genes in Arabidopsis thaliana. The double mutant tsn1 tsn2 displays only mild growth phenotypes under nonstress conditions, but germination, growth, and survival are severely affected under high salinity stress. Either TSN1 or TSN2 alone can complement the double mutant, indicating their functional redundancy. TSN accumulates heterogeneously in the cytosol and relocates transiently to a diffuse pattern in response to salt stress. Unexpectedly, stress-regulated mRNAs encoding secreted proteins are significantly enriched among the transcripts that are underrepresented in tsn1 tsn2. Our data also reveal that TSN is important for RNA stability of its targets. These findings show that TSN is essential for stress tolerance in plants and implicate TSN in new, potentially conserved mechanisms acting on mRNAs entering the secretory pathway.

Cellular responses to auxin: division versus expansion.

The phytohormone auxin is a major regulator of plant growth and development. Many aspects of these processes depend on the multiple controls exerted by auxin on cell division and cell expansion. The detailed mechanisms by which auxin controls these essential cellular responses are still poorly understood, despite recent progress in the identification of auxin receptors and components of auxin signaling pathways. The purpose of this review is to provide an overview of the present knowledge of the molecular mechanisms involved in the auxin control of cell division and cell expansion. In both cases, the involvement of at least two signaling pathways and of multiple targets of auxin action reflects the complexity of the subtle regulation of auxin-mediated cellular responses. In addition, it offers the necessary flexibility for generating differential responses within a given cell depending on its developmental context.

Recent progress in auxin biology.

Like animals, plants have evolved into complex organisms. Developmental cohesion between tissues and cells is possible due to signaling molecules (messengers) like hormones. The first hormone discovered in plants was auxin. This phytohormone was first noticed because of its involvement in the response to directional light. Nowadays, auxin has been established as a central key player in the regulation of plant growth and development and in responses to environmental changes. At the cellular level, auxin controls division, elongation, and differentiation as well as the polarity of the cell. Auxin, to integrate so many different signals, needs to be regulated at many different levels. A tight regulation of auxin synthesis, activity, degradation as well as transport has been demonstrated. Another possibility to modulate auxin signaling is to modify the capacity of response of the cells by expressing differentially the signaling components. In this review, we provide an overview of the present knowledge in auxin biology, with emphasis on root development.

The AUXIN BINDING PROTEIN 1 is required for differential auxin responses mediating root growth.

BACKGROUND: In plants, the phytohormone auxin is a crucial regulator sustaining growth and development. At the cellular level, auxin is interpreted differentially in a tissue- and dose-dependent manner. Mechanisms of auxin signalling are partially unknown and the contribution of the AUXIN BINDING PROTEIN 1 (ABP1) as an auxin receptor is still a matter of debate. METHODOLOGY/PRINCIPAL FINDINGS: Here we took advantage of the present knowledge of the root biological system to demonstrate that ABP1 is required for auxin response. The use of conditional ABP1 defective plants reveals that the protein is essential for maintenance of the root meristem and acts at least on the D-type CYCLIN/RETINOBLASTOMA pathway to control entry into the cell cycle. ABP1 affects PLETHORA gradients and confers auxin sensitivity to root cells thus defining the competence of the cells to be maintained within the meristem or to elongate. ABP1 is also implicated in the regulation of gene expression in response to auxin. CONCLUSIONS/SIGNIFICANCE: Our data support that ABP1 is a key regulator for root growth and is required for auxin-mediated responses. Differential effects of ABP1 on various auxin responses support a model in which ABP1 is the major regulator for auxin action on the cell cycle and regulates auxin-mediated gene expression and cell elongation in addition to the already well known TIR1-mediated ubiquitination pathway.

Conditional repression of AUXIN BINDING PROTEIN1 reveals that it coordinates cell division and cell expansion during postembryonic shoot development in Arabidopsis and tobacco.

AUXIN BINDING PROTEIN1 (ABP1) has long been characterized as a potentially important mediator of auxin action in plants. Analysis of the functional requirement for ABP1 during development was hampered because of embryo lethality of the null mutant in Arabidopsis thaliana. Here, we used conditional repression of ABP1 to investigate its function during vegetative shoot development. Using an inducible cellular immunization approach and an inducible antisense construct, we showed that decreased ABP1 activity leads to a severe retardation of leaf growth involving an alteration in cell division frequency, an altered pattern of endocycle induction, a decrease in cell expansion, and a change in expression of early auxin responsive genes. In addition, local repression of ABP1 activity in the shoot apical meristem revealed an additional role for ABP1 in cell plate formation and cell shape. Moreover, cells at the site of presumptive leaf initiation were more sensitive to ABP1 repression than other regions of the meristem. This spatial context-dependent response of the meristem to ABP1 inactivation and the other data presented here are consistent with a model in which ABP1 acts as a coordinator of cell division and expansion, with local auxin levels influencing ABP1 effectiveness.

Dissecting the molecular basis of the regulation of wood formation by auxin in hybrid aspen.

Indole acetic acid (auxin) is a key regulator of wood formation, and an observed overlap between auxin concentration gradient and developing secondary xylem cells has led to the hypothesis that auxin regulates wood formation by acting as a morphogen. We dissected the role of auxin in wood formation by identifying the auxin-responsive transcriptome in wood-forming tissues and investigating alterations in wood formation in transgenic hybrid aspen plants (Populus tremula x Populus tremuloides) with perturbed auxin signaling. We showed that auxin-responsive genes in wood-forming tissues respond dynamically to changes in cellular auxin levels. However, the expression patterns of most of the auxin-responsive genes displayed limited correlation with the auxin concentration across this developmental zone. Perturbing auxin signaling by reducing auxin responsiveness reduced the cambial cell division activity, caused spatial deregulation of cell division of the cambial initials, and led to reductions in not only radial but also axial dimensions of fibers and vessels. We propose that, instead of acting as a morphogen, changes in auxin concentration in developing secondary xylem cells may provide important regulatory cues that modulate the expression of a few key regulators; these, in turn, may control the global gene expression patterns that are essential for normal secondary xylem development.

The auxin-binding protein 1 is essential for the control of cell cycle.

The phytohormone auxin has been known for >50 years to be required for entry into the cell cycle. Despite the critical effects exerted by auxin on the control of cell division, the molecular mechanism by which auxin controls this pathway is poorly understood, and how auxin is perceived upstream of any change in the cell cycle is unknown. Auxin Binding Protein 1 (ABP1) is considered to be a candidate auxin receptor, triggering early modification of ion fluxes across the plasma membrane in response to auxin. ABP1 has also been proposed to mediate auxin-dependent cell expansion, and is essential for early embryonic development. We investigated whether ABP1 has a role in the cell cycle. Functional inactivation of ABP1 in the model plant cell system BY2 was achieved through cellular immunization via the conditional expression of a single-chain fragment variable (scFv). This scFv was derived from a well characterized anti-ABP1 monoclonal antibody previously shown to block the activity of the protein. We demonstrate that functional inactivation of ABP1 results in cell-cycle arrest, and provide evidence that ABP1 plays a critical role in regulation of the cell cycle by acting at both the G1/S and G2/M checkpoints. We conclude that ABP1 is essential for the auxin control of cell division and is likely to constitute the first step of the auxin-signalling pathway mediating auxin effects on the cell cycle.

Does auxin binding protein 1 control both cell division and cell expansion?

The Auxin-Binding Protein 1 (ABP1) was identified over 30 years ago thanks to it's high affinity for active auxins. ABP1 plays an essential role in plant life yet to this day, its function remains 'enigmatic.' A recent study by our laboratory shows that ABP1 is critical for regulation of the cell cycle, acting both in G(1) and at the G(2)/M transition. We showed that ABP1 is likely to mediate the permissive auxin signal for entry into the cell cycle. These data were obtained by studying a conditional functional knock-out of ABP1 generated by cellular immunization in the model tobacco cell line, Bright Yellow 2.

Auxins.

Auxin is a multifactorial phytohormone that is required for cell division. Fine gradients determine points of developmental change in time and space. It is associated intimately with the axiality of plant growth, and increasing doses lead to cell expansion or inhibition of cell expansion in different tissues. From embryonic patterning to fruit dehiscence every plant process has some involvement with auxin as a hormonal signal, including responses to wounding. Moreover, synthetic auxins have widespread uses as agrochemicals, particularly as selective herbicides. Despite the importance of auxin as a plant signal the pathways of its biosynthesis are still not clear. Much more is known about auxin perception and the mechanisms through which gene transcription is regulated. One receptor has been identified, and protein crystallography data has explained its auxin-binding capacity, but this is likely to control only a subset of auxin-mediated responses. Little is known of the signal transduction intermediates. A second receptor has been nominated and may be involved in controlling auxin-mediated gene transcription. A complex set of proteins comprising signalosome and proteasome contribute to the regulation of sets of transcription factors to confer regulation by derepression. A set of auxin transport proteins has been described with associated regulatory interactors, and these account for polar auxin flow and the control of auxin movements across cells, tissues, and around the plant. The gradients these transport systems build regulate the responses of growth and differentiation, including the plant's response to gravity. These areas are described and discussed by relating the physiology of the whole plant to the details of genetic and protein activities.

The tobacco bZIP transcription factor BZI-1 binds the GH3 promoter in vivo and modulates auxin-induced transcription.

Summary In order to establish the biological function of the tobacco basic leucine zipper (bZIP) transcription factor BZI-1 in hormone signalling, we have analysed transgenic plants which were altered with respect to the protein level or the activation potential of BZI-1. Overexpression of a dominant-negative derivative of BZI-1, lacking the N-terminal activation domain, resulted in plants displaying reduced internode size, enhanced lateral shoot formation and small, curly leaves. The response to auxin monitored with reference to root organogenesis, epinastic leaf curvature and transcription of the auxin-induced GH3 gene was reduced. In vitro, BZI-1 specifically binds to ACGT elements (ACEs) present in the GH3 promoter. In vivo, binding to the GH3 promoter was confirmed by chromatin immunoprecipitation (ChIP). Overexpression of BZI-1 in transgenic plants did not lead to a significant activation of the GH3 target gene. In contrast, plants expressing a VP16 (Herpes simplex virion protein 16)-BZI-1 fusion protein showed enhanced auxin-induced GH3 transcription. However, VP16-BZI-1 is insufficient to trigger GH3 expression independently of the auxin stimulus. Whereas auxin responsiveness has been shown to be mediated by ARF (auxin response factor) transcription factors, we discuss a function of BZI-1 assisting in fine-tuning of auxin-induced transcription.