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Bringing into focus the cellular dynamics of the plant growth hormone gibberellin

Periodic Reporting for period 4 - CELL HORMONE (Bringing into focus the cellular dynamics of the plant growth hormone gibberellin)

Período documentado: 2022-07-01 hasta 2022-12-31

Plants and animals use a suite of mobile small molecules, i.e. hormones, to coordinate myriad cellular activities into a multicellular whole.

This project works towards understanding the ‘logic’ of how phytohormone patterns and dynamics connect upstream cues to downstream developmental programs. In light of the breakthroughs in understanding hormone biosynthesis, perception, and signalling of the past three decades, the key challenge before us is to unravel how specific environmental and endogenous cues direct specific responses through spatiotemporally specific hormone accumulations and depletions. This challenge is deepened by a
fundamental problem in hormone research – hormone concentrations and responses are most often assessed at the organism or organ level. Furthermore, hormone perturbations are often genetic mutants or hormone treatments that are highly pleiotropic owing to the broad-based nature of hormone programming.
Our project aims to correct these problems by mapping spatiotemporal hormone patterns at high-resolution in vivo followed by perturbing the mapped hormone patterns at high-resolution to interrogate how these patterns are determined and how these hormone patterns relate to plant developmental programs relevant to agriculture.

Indeed, manipulation of the growth hormone gibberellin, the principal focus of this project, is already fundamental to global food security and thus deeper understanding of gibberellin signalling holds great promise for the future.
Prior to the current project period, we engineered Gibberellin Perception Sensor (GPS1), a fluorescent biosensor that detects gibberellin, and deployed it in Arabidopsis thaliana where we observed gradients of gibberellin in living plant tissues. We continue to map gibberellin patterns in Arabidopsis roots and hypocotyls and have now also perturbed these patterns in myriad ways to reveal how they are made and how the influence plant development.
Our key developmental questions were how are gibberellin levels controlled, what is their relationship to cell growth, and how does environmental change affect gibberellin and plasticity?

We studied in depth the mechanisms directing gibberellin levels in hypocotyls and roots. We have determined the dominant biochemical and light signalling steps determining how hypocotyl gibberellin gradients are drawn in darkness and redrawn in the light. During these studies, we also uncovered that a spatial correlation between gibberellin levels and cellular growth in the dark grown hypocotyl can be disrupted in certain mutant backgrounds, indicating that gibberellins alone are not responsible for setting cell growth patterns.

In roots, we previously observed a gibberellin gradient correlating with cell length and have now deciphered which enzymatic steps are most important for setting up this gradient. Interestingly, root meristematic cells and elongating cell types exhibited distinct rate-limiting biochemical steps. An unexplained gibberellin pattern in a third cell type led us to a new mode of regulation of gibberellin biosynthesis. Our results conflicted with previous multiscale mathematical models of gibberellin patterning and thus we collaborated the modellers that generated the models to develop new multiscale models that better capture in vivo hormone dynamics in roots. Finally, we are continuing to probe which gibberellin accumulations control which growth phenomena, including testing an emergent hypothesis that cell elongation requires two distinct phases of GA accumulation.

We further investigated in high resolution the hormonal and biophysical dynamics in hypocotyls exposed to light for the first time, i.e. apical hook opening, which is a model for differential growth. We have characterised how light triggers dynamics in growth, pH, microtubule orientations and gibberellin and auxin hormone levels at the cellular level and also developed a morphodynamic mathematical model describing apical hook opening. Working iteratively between experiments and modelling, we found evidence for several phenomena being critical for differential cell growth during apical hook opening. Some of the answer lies in innate physical stress patterns that emerge during the opening of a pressurised hooked object with interior expansion and exterior restriction. The model is improved with the addition of hormone differential mediated mechanical loosening along with dynamic and differential microtubule reorientations in response to stress. Another fascinating discovery is that the light induced hormone dynamics are themselves dependent on changes in physical mechanics such that a mechanochemical feeback loop is operating in apical hook opening.
• Specification of mechanisms controlling gibberellin gradients in roots and hypocotyls.
• Redevelopment of a multiscale model describing the mechanisms controlling gibberellin gradients in roots.
• Overturning standing hypotheses regarding the relationship between gibberellin distributions and cellular growth.
• Discovery of a new mechanism of regulation of gibberellin biosynthesis.
• Engineering of a novel biosensor, GPS2, that exhibits improved reversibility and orthogonality (i.e. reduced interference with endogenous signalling when expressed in plants).
• Development of a new multiscale model of hormone action and physical forces in differential growth in our hypocotyl system.
• Extensive library of gibberellin perturbation lines.
• Engineering of a optogenetic gene expression system for plants.

Expected impact – a deeper fundamental understanding of how hormone levels direct plant development and redirect plant development following environmental change will be critical for creating improved crops and cropping practices for food security and climate resilience. The fundamental advances we have accomplished will also impact on developmental biology through providing a way forward in the field’s movement towards quantitative, high-resolution information and increasingly accurate multiscale models of complex developmental processes. Additionally, our technological contributions of novel hormone biosensors and hormone perturbation lines will immediately be useful a range of researchers working in species that produce gibberellin, including bacteria, fungi and plants. The Highlighter system, once implemented in stable transgenics, has extremely broad applications in both fundamental research and agriculture.
GA patterns in vivo