Gravitropic setpoint angle control in higher plants

Lead Research Organisation: University of Leeds
Department Name: Ctr for Plant Sciences

Abstract

The overall shape of plants, the space they occupy above and below ground, is determined principally by the number, length, and angle of their branches. Interestingly, the angles at which many lateral branches grow out are set and maintained relative to gravity rather than the main root-shoot axis. These angles are known as a gravitropic setpoint angles or GSAs. Despite being a fundamental component of the wonderful variation in plant architecture observed throughout nature until recently the mechanisms underlying GSA regulation were not known.

The maintenance of vertical GSAs in the main primary root-shoot axis, otherwise known as gravitropism, is well understood: displacement of the main stem or root is perceived within specialised gravity-sensing cells in roots and shoots. This leads to the active transport of a plant hormone called auxin from these cells and to the lower side of the displaced organ. In roots, auxin inhibits cell elongation, causing bending downwards to the vertical while in the shoot, auxin does the opposite, promoting upward curvature. In our previous work on GSA control we showed that the same underlying gravitropic response occurs in non-vertical root and shoot branches but is counteracted by another growth component, the antigravitropic offset. We showed that branches that are less vertical have a stronger antigravitropic offset and that the magnitude of this offset is also regulated directly within the gravity-sensing cell. We have also shown that like gravitropism, antigravitropism depends on auxin transport but in this case from the upper, rather than the lower side of the gravity-sensing cell. The molecular mechanisms underlying antigravitropic activity and how that activity is restricted to non-vertical branches is not known and is the subject of this study.

The project has three parts. In the first, we focus on the regulation auxin transporters called PIN proteins that are responsible for moving auxin out of cells. In gravity-sensing cells, two particular PINs, PIN3 and PIN7, appear to mediate both gravitropic and antigravitropic auxin fluxes. The subcellular localisation and activity of PINs has been shown to be regulated by their phosphorylation. Our preliminary work has shown that a mutated phosphomimic version of PIN3 has less vertical lateral roots while a mutated PIN3 that cannot be phosphorylated has more vertical lateral roots. The simplest interpretation of these data is that unphosphorylated PINs in gravity-sensing cells contribute to downward, gravitropic auxin flux while phosphorylated PIN mediates an upward, antigravitropic flux. Importantly, while the direction of the gravitropic flux out of the gravity-sensing cells can be reset very rapidly if the plant is reorientated, our data indicate that changes in the direction of auxin fluxes that drive antigravitropic activity take much longer to be re-established. Therefore, by tracking the molecular and cell biological events within gravity-sensing cells of lateral roots undergoing simple reorientations we can observe which PINs and PIN phosphoregulators are associated with the faces of the cell that were once transporting auxin to sustain gravitropic or antigravitropic growth and then watch as the new polarities for gravi- and antigravitropic auxin fluxes are re-established.

A central feature of gravitropism is that the magnitude of graviresponse increases the further an organ is moved away from the vertical. This phenomenon is crucial for GSA control and so in the second part of the project we use advanced microscopy to understand the biophysical and molecular basis of this response.

In the final part of the project we will screen a collection of mutated Arabidopsis plants to look for plants with little or no non-vertical growth in lateral branches. This will allow us to identify the genes that restrict antigravitropic activity to lateral branches and allow us to form a coherent understanding of GSA control in plants.

Technical Summary

Lateral root and shoot branches are often maintained at specific angles with respect to gravity, a quantity known as the gravitropic set-point angle (GSA). The GSA values of lateral shoots and roots are most often non-vertical, allowing the plant to optimise the capture of resources both above- and below-ground. Despite the importance of branch angle as a fundamental parameter of plant form, until now research has focused on the mechanisms controlling numbers of lateral roots and shoots, and work on gravitropism has been all but confined to the primary root-shoot axis. Our recent work has addressed the central question of how gravity-dependent non-vertical GSAs are set and maintained. We showed that gravity-dependent non-vertical growth of lateral organs is sustained by means of an antigravitropic offset (AGO) mechanism that operates in tension with gravitropic response to sustain stable growth at non-vertical angles. Further we showed that the activity of the AGO requires auxin transport and that auxin specifies the magnitude of the AGO specifically in the gravity-sensing cells of lateral root and shoot gravity-sensing cells.

These findings represent a significant advance in our understanding of the biology of lateral organ gravitropism and provide the basis for the work proposed here. We aim to identify the molecular machinery that underlies the antagonistic interactions between gravitropism and antigravitropism within gravity-sensing cells of lateral branches. We also propose a forward genetic screen to identify the factors that restrict antigravitropic offset activity to lateral root and shoot branches. The project involves a combination genetics, molecular genetics, cell biology and state-of-the-art imaging along with novel assays designed to dissect the biophysical and molecular events within gravity-sensing cells that give rise to the antagonism between gravitropism and antigravitropism.

Planned Impact

This project addresses a fundamental question in plant developmental biology that is also of great agronomic importance. The primary beneficiaries of the research, outside of the academic plant science community, will be commercial plant breeding companies. We hope that in the long term our research in Arabidopsis will be used to inform crop improvement strategies that lead to crop species with improved water and nutrient acquisition capability. Although most of the proposed work here will be carried out on Arabidopsis, the fact that the general principles of GSA control we have discovered have been shown to apply to a range of broadleaf (common bean, pea, soy bean) and cereal species (maize, wheat, rice, barley, and oats) indicates that these approaches are likely to be of broad relevance and utility to a range of plant and crop scientists.

Bayer Cropscience have already expressed their support of the work and we are currently engaging with additional potential commercial partners, including BASF and Syngenta. They recognise the potential for the manipulation of root and shoot angles for the optimisation of water- and nutrient-use efficiency and photosynthetic performance respectively. Particularly in the USA, market research carried out by Leeds University has shown that improvement in nitrogen assimilation would be of significant importance in the case of crops such as maize and soybean. The importance of growth angle as a essential yield-determining component of plant architecture has further been emphasized through several studies that have shown increases in shoot biomass and nutrient assimilation in shallower rooting varieties of maize and bean (Reviewed in Lynch, 2013, doi:10.1093/aob/mcs293) These groups have also attempted to identify QTL controlling branching angle in these species. This highlights the desirability of tools for controlled manipulation of structural traits to maximise performance at lower rates of fertiliser application and reduced reliance on irrigation and other resources. We will closely remain in close contact with Bayer Cropscience, BASF, and Syngenta in this regard and will seek opportunities to collaborate with them under guidance from the University Research and Innovation Services.

While taking care not to damage the commercial potential of our work, we will disseminate aspects of our work both within the academic community and also to non-specialist audiences through outreach opportunities such as Cafe Scientifique and Discovery Zone events and other invited public lectures and discussions. Academic beneficiaries will be mainly in international crop improvement, auxin biology, and developmental biology communities. The proposed research would be of great interest to these researchers because identification of the auxin-regulated mechanisms that control branch angle will provide new targets and approaches for the manipulation of plant architectural traits.

Publications

10 25 50
 
Description We are using molecular and genetic tools to dissect the mechanisms controlling GSA in the Arabidopsis lateral root. thus far, we have shown that gravitropism in lateral roots is angle-dependent. Further, we show that the polarity of the previously-identified antigravitropic offset (AGO) mechanism is, like that of gravitropic response, set and resettable relative to gravity. Critically, the kinetics of resetting the AGO are slow relative to alterations in the polarity of gravitropic response. This combination of a slow-resetting and angle-independent AGO acting in tension with angle-dependent gravitropic response underpins the maintenance of non-vertical GSAs. We show that both stable growth at GSA and upward and downward graviresponse in lateral roots are dependent on auxin transport and the generation of auxin symmetry and auxin asymmetry respectively. These patterns of auxin distribution in the root tip are reflected in the control of subcellular localisation of auxin-transporting PIN3 and PIN7 proteins in the lateral root columella, the regulation of which is dependent on the PIN protein phosphatase subunit RCN1. Finally, we show that in addition to driving tropic response, auxin acts to induce more vertical growth by increasing RCN1 levels within the lateral root columella.
Exploitation Route We expect that our findings will provide technological approaches to alter root growth angle, a important trait that can affect the ability of root systems to forage for nutrients and water.
Sectors Agriculture, Food and Drink

 
Description BBSRC responsive mode
Amount £576,766 (GBP)
Funding ID BB/N010124/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 07/2016 
End 06/2019
 
Description Friml Collaboration 
Organisation Institute of Science and Technology Austria
Country Austria 
Sector Academic/University 
PI Contribution We share an interest in gravitropism and PIN biology with this group. We have supplied the Friml group we molecular genetic tools.
Collaborator Contribution The Friml lab has hosted PDRAs from my group to allow them to use the vertical-stage confocal microscope they have at IST, Vienna
Impact Papers are being prepared
Start Year 2013
 
Description Weijers lab collaboration 
Organisation University of Wageningen
Country Netherlands 
Sector Academic/University 
PI Contribution We have shared research materials and data ahead of publication. We also performed SPR analysis for the paper listed below.
Collaborator Contribution They have shared research materials and data ahead of publication.
Impact Boer, D.R., Freire-Rios, A., van den Berg W.A., Saaki, T., Manfield, I.W., Kepinski, S., López- Vidrieo, I., Franco-Zorrilla, J.M., de Vries, S.C., Solano, R., Weijers, D., Coll, M. (2014) Structural basis for DNA binding specificity by the auxin-dependent ARF transcription factors. Cell 30;156(3):577-89.
Start Year 2010