France

Seedling de-etiolation involves massive change in genome expression, many light-responsive genes undergoing pioneering rounds of transcription within minutes. In previous studies we reported that Arabidopsis photomorphogenesis indeed associates with a ~3-fold increase of global RNA Polymerase II elongation activity in most cotyledon nuclei (1), which, together with more recent studies (2), suggests that light induces a switch from a relatively quiescent to a more active transcriptional status during the transition. Studies from others and us focusing on nuclear and chromatin dynamics also unveiled that de-etiolation involves genome-wide variations nuclear architecture and in chromatin hallmarks of transcription elongation status at thousands&#39; protein-coding genes (1, 3-4), which are controlled by DET1 and other light signaling components (3). Yet, because of data normalization practices, the functional consequences of these multilevel adaptations of the chromatin status to an elevated transcriptional regime are not observed when analyzing transcriptomic profiles obtained with classical microarray or RNA-seq analyses. We therefore developed a new methodological setup enabling an absolute quantification of RNA-seq data to explore the functional impact of transcription intensification on gene expression. We will present how this approach can be used to assess to which extent, and for which genes, a global increase of RNA Pol 2 activity is reflected at the transcript level during photomorphogenesis. This new standpoint should help revisiting how signaling paths and chromatin machineries modulate plant genome expression in response to the environment.

References:
1- Bourbousse et al, PNAS (2015) 112(21) e2836-44
2- Godoy Herz, et al. Mol Cell (2019) PMID: 30661982
3- Nassrallah, Rougee et al, eLife (2018) e37892
4- Charron et al, Plant Cell (2009) 21:3732-48

SEB Annual Conference 2022

Global increase of the nuclear transcriptional regime during Arabidopsis photomorphogenesis: a chromatin perspective

Discover the art of creating extraordinary science at SEB’s Annual Conference at Le Corum Conference Center     in Montpellier, France from 5 to  8 July 2022.

SEB’s Annual Conference is celebrated for creating the perfect composition of research, new discoveries and collaborative connections. Be creative and innovative by exchanging ideas and knowledge with masters of biology from all over the world.

2022 will find over 1500 experimental biologists from across the globe converging on the third-largest French city near the Mediterranean coast and the fastest-growing city in the country over the past 25 years.
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Please register for the conference by clicking on this link:

https://www.sebiology.org/events/seb-annual-conference-2022/registration.html

The site will be open for registered participants closer to the event.

2022 will find over 1500 experimental biologists from across the globe converging on the third-largest French city near the Mediterranean coast and the fastest-growing city in the country over the past 25 years - Montpellier, France

Seedling de-etiolation involves massive change in genome expression, many light-responsive genes undergoing pioneering rounds of transcription within minutes. In previous studies we reported that Arabidopsis photomorphogenesis indeed associates with a ~3-fold increase of global RNA Polymerase II elongation activity in most cotyledon nuclei (1), which, together with more recent studies (2), suggests that light induces a switch from a relatively quiescent to a more active transcriptional status during the transition. Studies from others and us focusing on nuclear and chromatin dynamics also unveiled that de-etiolation involves genome-wide variations nuclear architecture and in chromatin hallmarks of transcription elongation status at thousands' protein-coding genes (1, 3-4), which are controlled by DET1 and other light signaling components (3). Yet, because of data normalization practices, the functional consequences of these multilevel adaptations of the chromatin status to an elevated transcriptional regime are not observed when analyzing transcriptomic profiles obtained with classical microarray or RNA-seq analyses. We therefore developed a new methodological setup enabling an absolute quantification of RNA-seq data to explore the functional impact of transcription intensification on gene expression. We will present how this approach can be used to assess to which extent, and for which genes, a global increase of RNA Pol 2 activity is reflected at the transcript level during photomorphogenesis. This new standpoint should help revisiting how signaling paths and chromatin machineries modulate plant genome expression in response to the environment.

References:
1- Bourbousse et al, PNAS (2015) 112(21) e2836-44
2- Godoy Herz, et al. Mol Cell (2019) PMID: 30661982
3- Nassrallah, Rougee et al, eLife (2018) e37892
4- Charron et al, Plant Cell (2009) 21:3732-48

technical paper

The basket of nuclear pore complex (NPC) is an assembly of flexible protein filaments and acts as a dynamic scaffold at the nucleoplasmic side that regulates various essential cellular processes, including nuclear transport, RNA processing, chromatin organization, and gene expression. However, the mechanisms behind the regulation are still poorly understood, especially in plants. Here, we identified a guanylate-binding protein (GBP)-like GTPase (GBPL3) as a specific component of the NPC basket in Arabidopsis. GBPL3 is located at the nuclear rim and confers robust physical and genetic interactions with various NPC and nucleoskeleton constituents. Proximity labeling proteomics experiment suggests that GBPL3 is predominantly distributed at the NPC basket and that it is intimately associated with chromatin remodelers, transcription regulators, and RNA processing machinery. Importantly, we found that GBPL3 promotes the formation of biomolecular condensates at the NPC basket, which is capable of recruiting its interactors. Loss of GBPL3 led to severely stunted plant growth, aberrant transcription of abiotic and biotic stress-related genes, and retention of total mRNA in the nucleus. These results suggest that GBPL3 is a plant-specific NPC basket component and can recruit transcription and RNA processing machinery through condensate formation to modulate gene expression and subsequent RNA processing, which is required for plant growth and stress responses.

GBPL3 is a novel plant nucleoporin that recruits RNA processing and transcription machinery at the nuclear basket by promoting condensates formation

Plants must cope with the highly dynamic nature of light to survive and thrive, a process referred to as acclimation. Although acclimation to different light intensities has been well studied, there has been little research into the effects of dynamic lighting regimes on this process. Natural fluctuating light in field grown crops, can limit photosynthesis through slow stomatal opening in response to increasing light, whilst slow stomatal closure can erode water use efficiency (McAusland et al, 2016). It has been reported that both biochemical and stomata limitation on photosynthesis can lead to wheat yield losses of upto 21% (Taylor and Long, 2017).Previously dynamic growth light has been shown to effect the photosynthetic abilities of Arabidopsis thaliana, reducing maximum photosynthetic capacity and light absorption and altering light use efficiency along with changes in photoprotection (Vialet-Chabrand et al., 2017). To understand changes in gene expression resulting from acclimation to fluctuating light, Whole Genome Bisulfite Sequencing (WGBS) was utilised to examine changes in DNA methylation. Genome-wide changes were noted, with differentially methylated regions observed across a range of protein coding genes, transposable elements, and non-coding RNAs, as well as multiple regulatory sequences. RNAseq analysis demonstrated that some of these genes with changed methylation state impacted transcription, although many more changes in transcript levels were noted beyond the WGBS data. This could provide a new set of gene targets for improved photosynthetic efficiency and ultimately crop yield, which could be incorporated into on-going crop breeding programmes.

The Ups and Downs of Being an Outdoor Plant

Decision making have hitherto been investigated almost exclusively in multicellular neural organisms. Yet, evidence for decision making  have been described in single celled organisms such as acellular slime molds. In the last 20 years, slime molds have become a model system, with multiple experiments seeking to reproduce behavioural predictions from neuroscience, animal behaviour and psychology In this talk, I will  focus on decision making in slime molds and explore various frameworks: nutritional geometry, speed versus accuracy trade-off, magnitude-sensitive reaction times, Weber's law and social influence.  Hopefully by the end of my talk, you will be convinced that slime molds are an ideal model system in which to investigate fundamental mechanisms underlying the ground-floor of decision making abilities. 

Problem solving in acellular slime molds

Extensive efforts have revealed genetic programs plants use to control their development in response to the environment. Much less is known about how the output of these programs is impacted by the multicellular context they operate within. In order to better understand how collections of cells in plants process information, we are taking a computational perspective of development and examining parallels between these naturally evolved biological organisms and engineered computational systems. More specifically, whether the control principles of distributed computation apply to plants. In an effort to address this, we are viewing plant organs as integrated systems of interacting cells, and mapping intercellular connectivity into networks. This provides a quantitative approach to understanding cellular patterning, while mapping the multicellular “circuitry” plants use to perform computation. By integrating these topological templates with mathematical models describing the genetic programs that operate within individual cells, the impact of each cell organization and communication on developmental outputs is being examined. This is providing insight into the extent to which development is controlled beyond intracellular genetic mechanisms.  

Information processing in plant organs

Fate decisions in developing tissues involve cells transitioning between a set of discrete cell states. Geometric models, often referred to as Waddington landscapes, are an appealing way to describe differentiation dynamics and developmental decisions. We consider the differentiation of neural and mesodermal cells from pluripotent mouse embryonic stem cells exposed to different combinations and durations of signalling factors. We developed a principled statistical approach using flow cytometry data to quantify differentiating cell states. Then, using a framework based on Catastrophe Theory and approximate Bayesian computation, we constructed the corresponding dynamical landscape. The result was a quantitative model that accurately predicted the proportions of neural and mesodermal cells differentiating in response to specific signalling regimes. Taken together, the approach we describe is broadly applicable for the quantitative analysis of differentiation dynamics and for determining the logic of developmental cell fate decisions.

Gene-free landscape models for development

Capturing the rules governing developmental decisions in multicellular organisms is key to study cellular functions, including  reprogramming of cell differentiation and function. To understand how different genes coordinately regulate developmental decisions towards cell function, we used a multi-scale hybrid model integrating ordinary differential equations (ODEs) and agent-based modeling. This model allowed us to gain a deeper understanding in the coordinate function of key developmental genes, including WUSCHEL-RELATED HOMEOBOX 5 (WOX5) and SHORTROOT (SHR), to regulate stem cell divisions. We predicted cell-type-specific expression dynamics of SHR, SCARECROW, WOX5, AN3 and CYCLIND6;1, and showed an interdependency between CEI and QC divisions. At present, we know these stem cell populations divide, differentiate and display functions in response to relative positional information. A major challenge is to accurately capture the multicellular 3 dimensional (3D) microenvironment. To further study systemic behavior of these stem cells within a defined microenvironment, we established the framework of 3D bioprinting with plant cells to study cell viability, cell division, and cell identity. Within our designed microenvironment, the deposited Arabidopsis and soybean cells re-entered the cell cycle, which led to the formation of microcalli. Furthermore, we showed that the identity of isolated cells of Arabidopsis roots expressing endodermal markers maintained longer periods of time. We believe that the 3D bioprinting framework paves the way for a general use of 3D bioprinting for studying the rules governing developmental decisions.

Coordinated regulation of gene expression among stem cells

The plant hormone gibberellin (GA) is involved in many developmental processes, however, how GA is distributed within plant tissues is not yet understood. In this talk, we present a multi-cellular mathematical model for GA transport within the Arabidopsis root. We consider both passive and facilitated modes of transport through the symplast and the apoplast. In particular, we model the effects of the NPF cytoplasm importers, localised in different layers of the root. In addition, we consider the effect of a vacuole as a possible storage mechanism for GA within each cell and model the role of transport across the tonoplast. We assume an effective GA source coming from the stele and constant degradation rate of GA within the whole root. We also present results on the values of the various transporter permeabilities, estimated from a model and oocyte experimental data. We discuss the governing equations and show numerical results predicting the spatial and temporal distribution of GA. Our results provide insights into the role of the vacuole in regulating the amount of GA in the cell, and how this contributes to GA distributions and cellular decision making.

Modelling GA transport within the Arabidopsis root

The hormone gibberellin (GA) controls plant growth and regulates growth responses to environmental stress. In roots and monocotyledonous leaves, GA controls growth by regulating the size of the division zone - regulating the cell's decision to cease division and begin rapid elongation. We used a systems approach to investigate the establishment of the GA distribution in the maize leaf growth zone to understand how drought and cold alter leaf growth. By developing and parameterizing a multiscale computational model that includes cell movement, growth-induced dilution and metabolic activities, we revealed that the GA distribution is predominantly determined by variations in GA metabolism. Considering wild-type and UBI::GA20-OX-1 leaves, the model predicted the peak in GA concentration, which has been shown to determine division-zone size. Drought and cold modified enzyme transcript levels, although the model revealed that this did not explain the observed GA distributions. Instead, the model predicted that GA distributions are also mediated by post-transcriptional modifications increasing the activity of GA20oxidase in drought and of GA2oxidase in cold, which we confirmed by enzyme activity measurements. This work provides a new, mechanistic understanding of the GA metabolism in plant growth regulation.

Modelling reveals post-transcriptional regulation of GA metabolism enzymes in response to drought and cold

During in vitro tissue culture, plants form callus and regenerate organs by reprograming developmental pathways. In addition to hormonal signals, stress-induced cues, such as those from wounding, can trigger and promote cellular reprogramming. Conjugation of the small ubiquitin-related modifier (SUMO) to target transcriptional regulators is thought to rapidly and precisely modulate their activity in response to stress. We previously investigated if SUMOylation is involved in wound-induced reprograming and showed that loss of the E3 ligase SIZ1 causes a hyper-active wound response and enhances shoot regeneration (Coleman et al, 2020). Based on these observations, we hypothesized that SUMOylation suppresses wounding-induced regulator(s) and searched for early acting players underlying the stress-induced cellular reprograming. By analyzing the transcriptome of wounded plants, we found that HEAT SHOCK FACTOR A1s (HSFA1s) are activated after wounding. Mutant and overexpression analysis showed that HSFA1s, also known to be activated by heat stress, promote shoot regeneration in Arabidopsis. Furthermore, the regulation of HSFA1s appeared independent of transcription and biochemical analysis revealed that they are subject to SUMOylation in response to stress. Here, I will discuss how key stress-induced transcription factors regulate cellular reprograming and how SUMOylation modulates this regenerative response in plants.

The role of SUMOylation during shoot regeneration in Arabidopsis

SUMO conjugation is emerging as an important mechanism to transduce environmental cues into cellular signaling. We present evidence to show that SUMOylation of key transcriptional regulators such as DELLA and BZR1, provides a conduit for environmental influence of root growth and development during salt stress. SUMOylation stabilizes BZR1 in the nucleus by inhibiting its interaction with BIN2 kinase. During salt stress, Arabidopsis plants arrest growth through deSUMOylation of BZR1 in the cytoplasm by promoting the accumulation of the BZR1 targeting SUMO protease, ULP1a. ULP1a mutants are salt tolerant and insensitive to the BR inhibitor, brassinazole with regards to root growth. BR treatment stimulates ULP1a degradation, allowing SUMOylated BZR1 to accumulate and promote growth. This study uncovers a mechanism for integrating environmental cues into BR signaling to shape root growth.
Intriguingly an interacting partner of BZR1, the growth repressing DELLA proteins are also SUMOylated. We demonstrate that a proportion of DELLAs are conjugated to SUMO, the extent of conjugation increases during salt stress. We identify a SUMO interacting motif (SIM) in the GA receptor GID1 and demonstrate that SUMO-conjugated DELLA binds to this motif in a GA-independent manner. The consequent sequestration of GID1 by SUMO-conjugated DELLAs leads to an accumulation of non-SUMOylated DELLAs resulting in repression of root growth in salinity.  We present a model depicting how SUMO decodes environmental cues to shape root growth and development. 

Global increase of the nuclear transcriptional regime during Arabidopsis photomorphogenesis: a chromatin perspective

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GBPL3 is a novel plant nucleoporin that recruits RNA processing and transcription machinery at the nuclear basket by promoting condensates formation

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