Unite du Prof. Oates

Laboratoire

Publications associées (98)

Waiting on the Fringe: cell autonomy and signaling delays in segmentation clocks

Andrew Charles Oates

The rhythmic and sequential segmentation of the vertebrate body axis into somites during embryogenesis is governed by a multicellular, oscillatory patterning system called the segmentation clock. Despite many overt similarities between vertebrates, differences in genetic and dynamic regulation have been reported, raising intriguing questions about the evolution and conservation of this fundamental patterning process. Recent studies have brought insights into two important and related issues: (1) whether individual cells of segmentation clocks are autonomous oscillators or require cell–cell communication for their rhythm; and (2) the role of delays in the cell–cell communication that synchronizes the population of genetic oscillators. Although molecular details differ between species, conservation may exist at the level of the dynamics, hinting at rules for evolutionary trajectories in the system.

2020

Chargement

The body axis of vertebrates is subdivided into repetitive compartments called somites, which give rise primarily to the segmented architecture of the musculoskeletal system in the adult body. Somites form in a sequential and rhythmic manner in embryos and a physical boundary separates each somite from the rest of the unsegmented tissue and adjoining somites. Precise positioning of somite boundaries and determination of boundary cell fate in a select group of cells is thought to be driven by gene expression patterns and morphogen gradients. This pre-patterning step is followed by a mechanical process involving actomyosin activation in boundary cells and formation of an extracellular matrix that results in morphological boundary formation. While genes involved in somite boundary formation have been identified, there are many open questions about the underlying pre-patterning dynamics and mechanics and how these processes are coupled to generate a morphological boundary. Here, focusing on segmentation of zebrafish embryos as a model, we review pre-patterning processes critical for boundary formation and how cytoskeletal activity drives tissue separation. Our outlook is that this system holds exciting new avenues for unearthing general principles of boundary formation in developing embryos.

2020

Chargement

Transformation of the Drosophila Sex-Manipulative Endosymbiont Spiroplasma poulsonii and Persisting Hurdles for Functional Genetic Studies

Chloé Christiane Laurence Jollivet, Bruno Lemaitre, Florent François Masson

Insects are frequently infected by bacterial symbionts that greatly affect their physiology and ecology. Most of these endosymbionts are, however, barely tractable outside their native host, rendering functional genetics studies difficult or impossible. Spiroplasma poulsonii is a facultative bacterial endosymbiont of Drosophila melanogaster that manipulates the reproduction of its host by killing its male progeny at the embryonic stage. S. poulsonii, although a very fastidious bacterium, is closely related to pathogenic Spiroplasma species that are cultivable and genetically modifiable. In this work, we present the transformation of S. poulsonii with a plasmid bearing a fluorescence cassette, leveraging techniques adapted from those used to modify the pathogenic species Spiroplasma citri. We demonstrate the feasibility of S. poulsonii transformation and discuss approaches for mutant selection and fly colonization, which are persisting hurdles that must be overcome to allow functional bacterial genetics studies of this endosymbiont in vivo. IMPORTANCE Dozens of bacterial endosymbiont species have been described and estimated to infect about half of all insect species. However, only a few them are tractable in vitro, which hampers our understanding of the bacterial determinants of the host-symbiont interaction. Developing a transformation method for S. poulsonii is a major step toward genomic engineering of this symbiont, which will foster basic research on endosymbiosis. This could also open the way to practical uses of endosymbiont engineering through paratransgenesis of vector or pest insects.

AMER SOC MICROBIOLOGY2020

Chargement

What are you synching about? Emerging complexity of Notch signaling in the segmentation clock

Andrew Charles Oates, Olivier François Venzin

The Segmentation clock is a population of cellular genetic oscillators, located in the posterior of the elongating vertebrate embryo, that governs the rhythmic and sequential segmentation of the body axis into somites. Somites are blocks of cells that give rise to the segmented anatomy of the adult, including the backbone, muscles and skin. Malfunction of the segmentation clock results in malformations of these structures, a condition termed congenital scoliosis in the clinic. In all vertebrates, the oscillating cells of the segmentation clock are coordinated in a wave pattern, such that each new wave corresponds to a new segment. Maintenance of this wave pattern is important for precise segmentation and requires the local synchronization of the cellular oscillators. Existing models of the segmentation clock have explored the role of the Delta-Notch intercellular signaling pathway primarily as a coupling mechanism between neighboring autonomous oscillators. Recent work challenges several aspects of this simplification, suggesting that the mechanism of synchronization is more complex and may differ between species, and that Notch signaling may do more than synchronize cells. Here, we first examine evidence and models concerning the role of Notch signaling in driving, maintaining and synchronizing the mouse clock, highlighting results emerging from ex vivo culture systems of mouse segmentation clock cells. We then compare this to synchronization in the zebrafish, where accumulating evidence suggests that Notch signaling impacts the amplitude of the oscillating signal, and discuss whether the amplitude itself is meaningful for segmentation. Finally, we review work showing that multiple Delta ligands are active in segmentation, and consider how an interplay between these ligands could confer effective Notch functions in the segmentation clock. These lines of enquiry suggest that synchronization and Notch signaling are more complex than previously described, and reveal exciting new avenues for investigation into the coordination and precision of patterning the early embryo.

ACADEMIC PRESS INC ELSEVIER SCIENCE2020

Chargement

cis-regulatory variation modulates susceptibility to enteric infection in the Drosophila genetic reference panel

Tommaso Andreani, Maroun Bou Sleiman, Virginie Sandra Braman, Riccardo Dainese, Bart Deplancke, Michael Vincent Frochaux, Vincent Roland Julien Gardeux, Brian Dennis Hollis, Maria Litovchenko, Dani Osman

Background: Resistance to enteric pathogens is a complex trait at the crossroads of multiple biological processes. We have previously shown in the Drosophila Genetic Reference Panel (DGRP) that resistance to infection is highly heritable, but our understanding of how the effects of genetic variants affect different molecular mechanisms to determine gut immunocompetence is still limited. Results: To address this, we perform a systems genetics analysis of the gut transcriptomes from 38 DGRP lines that were orally infected with Pseudomonas entomophila. We identify a large number of condition-specific, expression quantitative trait loci (local-eQTLs) with infection-specific ones located in regions enriched for FOX transcription factor motifs. By assessing the allelic imbalance in the transcriptomes of 19 F1 hybrid lines from a large round robin design, we independently attribute a robust cis-regulatory effect to only 10% of these detected local-eQTLs. However, additional analyses indicate that many local-eQTLs may act in trans instead. Comparison of the transcriptomes of DGRP lines that were either susceptible or resistant to Pseudomonas entomophila infection reveals nutcracker as the only differentially expressed gene. Interestingly, we find that nutcracker is linked to infection-specific eQTLs that correlate with its expression level and to enteric infection susceptibility. Further regulatory analysis reveals one particular eQTL that significantly decreases the binding affinity for the repressor Broad, driving differential allele-specific nutcracker expression. Conclusions: Our collective findings point to a large number of infection-specific cis- and trans-acting eQTLs in the DGRP, including one common non-coding variant that lowers enteric infection susceptibility.

2020

Chargement

Mitochondrial haplotypes affect metabolic phenotypes in the Drosophila Genetic Reference Panel

Johan Auwerx, Roel Paulus Josephus Bevers, Virginie Sandra Braman, Bart Deplancke, Brian Dennis Hollis, Adamantia Kapopoulou, Maria Litovchenko

The nature and extent of mitochondrial DNA variation in a population and how it affects traits is poorly understood. Here we resequence the mitochondrial genomes of 169 Drosophila Genetic Reference Panel lines, identifying 231 variants that stratify along 12 mitochondrial haplotypes. We identify 1,845 cases of mitonuclear allelic imbalances, thus implying that mitochondrial haplotypes are reflected in the nuclear genome. However, no major fitness effects are associated with mitonuclear imbalance, suggesting that such imbalances reflect population structure at the mitochondrial level rather than genomic incompatibilities. Although mitochondrial haplotypes have no direct impact on mitochondrial respiration, some haplotypes are associated with stress- and metabolism-related phenotypes, including food intake in males. Finally, through reciprocal swapping of mitochondrial genomes, we demonstrate that a mitochondrial haplotype associated with high food intake can rescue a low food intake phenotype. Together, our findings provide new insight into population structure at the mitochondrial level and point to the importance of incorporating mitochondrial haplotypes in genotype-phenotype relationship studies.

2019

Chargement

Commensal Gut Bacteria Buffer the Impact of Host Genetic Variants on Drosophila Developmental Traits under Nutritional Stress

Maroun Bou Sleiman, Virginie Sandra Braman, Bart Deplancke, Michael Vincent Frochaux, Maria Litovchenko

Eukaryotic genomes encode several buffering mechanisms that robustly maintain invariant phenotypic outcome despite fluctuating environmental conditions. Here we show that the Drosophila gut-associated commensals, represented by a single facultative symbiont, Lactobacillus plantarum (Lp(WJL)), constitutes a so far unexpected buffer that masks the contribution of the host's cryptic genetic variation (CGV) to developmental traits while the host is under nutritional stress. During chronic under-nutrition, Lp(WJL) consistently reduces variation in different host phenotypic traits and ensures robust organ patterning during development; Lp(WJL) also decreases genotype-dependent expression variation, particularly for development-associated genes. We further provide evidence that Lp(WJL) buffers via reactive oxygen species (ROS) signaling whose inhibition impairs microbiota-mediated phenotypic robustness. We thus identified a hitherto unappreciated contribution of the gut facultative symbionts to host fitness that, beyond supporting growth rates and maturation timing, confers developmental robustness and phenotypic homogeneity in times of nutritional stress.

2019

Chargement

Detection of mRNA by Whole Mount in situ Hybridization and DNA Extraction for Genotyping of Zebrafish Embryos

Rachna Narayanan, Andrew Charles Oates

In situ hybridization is used to visualize the spatial distribution of gene transcripts in tissues and in embryos, providing important information about disease and development. Current methods involve the use of complementary riboprobes incorporating non-radioactive labels that can be detected by immunohistochemistry and coupled to chromogenic or fluorescent visualization. Although recent fluorescent methods have allowed new capabilities such as single-molecule counting, qualitative chromogenic detection remains important for many applications because of its relative simplicity, low cost and high throughput, and ease of imaging using transmitted light microscopy. A remaining challenge is combining high contrast signals with reliable genotyping after hybridization. Dextran sulfate is commonly added to the hybridization buffer to shorten development times and improve contrast, but this reagent inhibits PCR-based genotyping. This paper describes a modified protocol for in situ hybridization in fixed whole mount zebrafish embryos using digoxigenin (DIG) labeled riboprobes that are detected with alkaline phosphatase conjugated anti-DIG antibodies and nitroblue tetrazolium (NBT)/5-bromo-4-chloro-3-indolyl-phosphate (BCIP) chromogenic substrates. To yield embryos compatible with downstream genotyping after hybridization without sacrificing contrast of the signal, this protocol omits dextran sulfate and utilizes a lower hybridization temperature.

BIO-PROTOCOL2019

Chargement

Cellular checkerboard patterns are observed at many stages of embryonic development. We study an analytically tractable model for lateral inhibition and show that the steady states are analogous to optical phonons at the Gamma point, which have the wave number k = 0. We study the cases of cells arranged in linear and hexagonal lattices. To determine how the final pattern is selected it is necessary to take into account the granularity of the pattern and, analogously to solid-state physics, to redefine the basis and lattice sites in terms of a periodic crystal. The sites and basis are determined by looking at the symmetries of inhibitory interactions between cells. The redefined basis for cells placed in a linear lattice is composed by two cells which are embedded in another linear lattice, while for cells placed in a hexagonal lattice the redefined basis consists of three cells embedded in another hexagonal lattice. The pattern in hexagonal lattices can be driven into three different states: two of those states are periodic checkerboards and a third in which both periodic states coexist. These observations provides new predictions for experiments.

2019

Chargement

One of the great challenges in biology is to understand the mechanisms by which morphogenetic processes arise from molecular activities. We investigated this problem in the context of actomyosin-based cortical flow in C. elegans zygotes, where large-scale flows emerge from the collective action of actomyosin filaments and actin binding proteins (ABPs). Large-scale flow dynamics can be captured by active gel theory by considering force balances and conservation laws in the actomyosin cortex. However, which molecular activities contribute to flow dynamics and large-scale physical properties such as viscosity and active torque is largely unknown. By performing a candidate RNAi screen of ABPs and actomyosin regulators we demonstrate that perturbing distinct molecular processes can lead to similar flow phenotypes. This is indicative for a 'morphogenetic degeneracy' where multiple molecular processes contribute to the same large-scale physical property. We speculate that morphogenetic degeneracies contribute to the robustness of bulk biological matter in development.

ELIFE SCIENCES PUBLICATIONS LTD2018

Chargement