Biology subjects

Duplus-Bottin, H.

Publications and source records attributed to Duplus-Bottin, H..

6 recordsLinked to original sources

bioRxiv2026

ZmSWEET Sucrose transporters expressed in the endosperm adjacent to the maize embryo are necessary for carbon partitioning and embryo growth

In cereals such as maize, the kernel accumulates large quantities of storage compounds, including carbohydrates, lipids, and proteins, a process that requires tight regulation of nutrient transport. Seeds are composed of distinct tissues: the embryo, the endosperm, and maternal tissues that are symplastically isolated (not connected through plasmodesmata), necessitating specialized nutrient transfer mechanisms. In maize, nutrient transfer from maternal tissues to the endosperm via specialized basal endosperm transfer layer (BETL) cells is well characterized. However, nutrient transfer at the endosperm/embryo interface remains poorly understood. Consequently, the routes by which maternal carbon-derived sugars support embryo growth are still unclear. Our previous transcriptomic profiling uncovered a novel Endosperm domain Adjacent to the embryo Scutellum (EAS) with strong enrichment for transporter genes. Notably, genes encoding three sugar transporters from the SWEET (Sugars Will Eventually be Exported Transporters) family are highly and preferentially expressed in the EAS, suggesting the existence of a specialized sugar transfer mechanism at this interface. We show that the ZmSWEET proteins encoded by these genes are membrane-localized sucrose transporters and are functionally important for kernel development. A gene-edited triple zmsweet14a/14b/15a knock-out mutant exhibits reduced kernel weight and embryo size, significantly decreased embryo oil accumulation at maturity, and altered carbon partitioning within the kernel. In addition to these defects, mutant kernels display a significant reduction in primary root length during germination, indicating either lasting physiological consequences of disrupted sucrose transport during seed development or an additional role for these SWEET transporters during germination. Together, our findings demonstrate that sucrose transport at the endosperm/embryo interface is critical for proper carbon allocation, embryo development, and seed vigor, and identify the EAS as a key functional domain and potential target for improving seed composition.

plant biology↗

bioRxiv2025

Characterizing drug activity with sensitive interactomes in human living cells

A number of human diseases results from abnormal protein-protein interactions (PPIs) involving key regulatory proteins. Therefore, an important strategy in therapeutics consists in developing inhibitory molecules that should ideally be specific for the aberrant PPI. In this context, it is critical to evaluate the number of PPIs that could be affected by the candidate molecule and to analyze the inhibitory potential before and after the formation of the PPI. Surprisingly, these two molecular aspects are rarely considered, due to a lack of appropriate methodological approaches. In this study, we present a novel methodology that captures drug-sensitive PPIs by considering drug-induced cellular functions in live cell conditions. As a proof-of-concept, we identified interactions of the human core signaling protein ERK1 that are specifically affected by two different inhibitory molecules. In addition, we used a complementary set of innovative tools that allowed visualizing the inhibitory effect on ERK1/cofactor protein complexes after their assembly in living cells. Overall, our work establishes a unique methodological approach for deciphering drug activity for potentially any target bait protein of interest.

molecular biology↗

bioRxiv2025

Trans-acting mutations reveal non-nuclear modulators of both intrinsic and extrinsic gene expression noise in a eukaryote.

Gene expression regulation is a stochastic process that can be modified by mutations not only in a deterministic manner but also in a probabilistic way, for example by changing the extent of cell-to-cell variability in gene expression (also called "gene expression noise"). Although systematic studies have successfully analyzed the properties of cis-acting mutations that modulate expression noise at their own locus, less is known about the type of genetic changes that alter expression noise in trans (at loci distant from the mutation). Here, we applied genetic mapping on yeast strains (Saccharomyces cerevisiae) generated by random mutagenesis and identified three mutations (in chs1, yme2 and msh1 genes) that changed the expression noise of a reporter gene in the nuclear genome regulated by the yeast TDH3 promoter. These mutations affected either extrinsic noise (variability due to cell-specific factors), intrinsic noise (inherent variability within each cell) or both, and their effects were found to not be specific to the TDH3 promoter. Surprisingly, all three mutations targeted proteins located outside of the nucleus: Yme2 and Msh1 being involved in the maintenance of mitochondrial genome integrity, and Chs1 being necessary for the repair of cell-wall defects in freshly-born daughter cells. Our results reveal that mitochondrial state can modulate the extent of intrinsic expression noise of eukaryotic nuclear genes.

genetics↗

bioRxiv2024

Kinetic properties of optogenetic DNA editing by LiCre-loxP

Advances in optogenetics now allow to specifically modify the DNA of live cells with light. However, using these technologies successfully requires to know their properties in terms of sensitivity, efficiency, kinetics and mechanism. We previously developed an optogenetic tool made of a single chimeric protein called LiCre that enables the induction of specific changes in the genome with blue light via DNA recombination between loxP sites [1]. Here, we used in vitro and in vivo experiments combined with kinetic modeling to provide a deeper characterization of the photo-activated LiCre-loxP recombination reaction. We find that LiCre binds DNA with high affinity in absence of light stimulus and that this binding is cooperative although not as much as for the Cre recombinase from which LiCre was derived. In yeast, addition of riboflavin to the culture medium had no effect on LiCres efficiency, even when cells over-expressed riboflavin kinase, suggesting that abundance of the flavin mono-nucleotide co-factor is not limiting for the reaction. However, LiCres efficiency in yeast gradually increased when raising temperature from 20{degrees}C to 37{degrees}C. The recombination kinetics observed in live cells are best explained by a model where photo-activation of two or more DNA-bound LiCre units (happening in seconds) can produce (in several minutes) a functional recombination synapse. This model was able to capture the effect of a point mutation altering LiCres light cycle. This deeper understanding of the LiCre-loxP system provides additional knowledge for designing experiments where specific genetic changes are induced in live cells with light. AUTHOR SUMMARYUsing a technology called optogenetics, scientists are now able to change the DNA sequence of live cells by illuminating them with light. In theory, they can trigger in genetically-engineered organisms a mutation of interest in specific cells at a specific time. This practice, however, is not common because optogenetics relies on light-controlled enzymes that are recent and not well characterized. We previously developed one of these enzymes, called LiCre, which recognizes a specific piece of DNA. Following illumination with blue light, LiCre can switch on or off whatever gene is close by. Here, we combined experiments and computational modeling to better understand how, and how fast, LiCre works. We find that, although it is inactive in the dark, LiCre does not need photo-activation to bind to its DNA partner. We estimated the speed at which LiCre gets deactivated in the dark and the speed at which active LiCre molecules modify DNA. We also showed the effect of temperature and illumination dynamics on LiCres efficiency. These results will help design strategies where LiCre can be used to conduct genetic studies at high spatio-temporal resolution, or implement it in industrial and biomedical applications.

bioengineering↗

bioRxiv2020

A monogenic and fast-responding Light-Inducible Cre recombinase as a novel optogenetic switch

ABSTRACTOptogenetics enables genome manipulations with high spatiotemporal resolution, opening exciting possibilities for fundamental and applied biological research. Here, we report the development of LiCre, a novel light-inducible Cre recombinase. LiCre is made of a single flavin-containing protein comprising the asLOV2 photoreceptor domain of Avena sativa fused to a Cre variant carrying destabilizing mutations in its N-terminal and C-terminal domains. LiCre can be activated within minutes of illumination with blue light, without the need of additional chemicals. When compared to existing photoactivatable Cre recombinases based on two split units, LiCre displayed faster and stronger activation by light as well as a lower residual activity in the dark. LiCre was efficient both in yeast, where it allowed us to control the production of β-carotene with light, and in human cells. Given its simplicity and performances, LiCre is particularly suited for fundamental and biomedical research, as well as for controlling industrial bioprocesses.Competing Interest StatementThe authors declare the following competing interest: A patent application covering LiCre and its potential applications has been filed. Patent applicant: CNRS; inventors: Helene Duplus-Bottin, Martin Spichty and Gael Yvert.View Full Text

synthetic biology↗

bioRxiv2017

Assigning Function To Natural Allelic Variation Via Dynamic Modeling Of Gene Network Induction

More and more natural DNA variants are being linked to physiological traits. Yet, understanding what differences they make on molecular regulations remains challenging. Important properties of gene regulatory networks can be captured by computational models. If model parameters can be personalized according to the genotype, their variation may then reveal how DNA variants operate in the network. Here, we combined experiments and computations to visualize natural alleles of the yeast GAL3 gene in a space of model parameters describing the galactose response network. Alleles altering the activation of Gal3p by galactose were discriminated from those affecting its activity (production/degradation or efficiency of the activated protein). The approach allowed us to correctly predict that a non-synonymous SNP would change the binding affinity of Gal3p with the Gal80p transcriptional repressor. Our results illustrate how personalizing gene regulatory models can be used for the mechanistic interpretation of genetic variants.

systems biology↗