Precision of readout at the hunchback gene: analyzing short transcription time traces in living fly embryos

The simultaneous expression of the hunchback gene in the numerous nuclei of the developing fly embryo gives us a unique opportunity to study how transcription is regulated in living organisms. A recently developed MS2-MCP technique for imaging nascent messenger RNA in living Drosophila embryos allows us to quantify the dynamics of the developmental transcription process. The initial measurement of the morphogens by the hunchback promoter takes place during very short cell cycles, not only giving each nucleus little time for a precise readout, but also resulting in short time traces of transcription. Additionally, the relationship between the measured signal and the promoter state depends on the molecular design of the reporting probe. We develop an analysis approach based on tailor made autocorrelation functions that overcomes the short trace problems and quantifies the dynamics of transcription initiation. Based on live imaging data, we identify signatures of bursty transcription initiation from the hunchback promoter. We show that the precision of the expression of the hunchback gene to measure its position along the anterior-posterior axis is low both at the boundary and in the anterior even at cycle 13, suggesting additional post-transcriptional averaging mechanisms to provide the precision observed in fixed embryos

Synthetic reconstruction of the hunchback promoter specifies the role of Bicoid, Zelda and Hunchback in the dynamics of its transcription

For over 40 years, the Bicoid-hunchback (Bcd-hb) system in the fruit fly embryo has been used as a model to study how positional information in morphogen concentration gradients is robustly translated into step-like responses. A body of quantitative comparisons between theory and experiment have since questioned the initial paradigm that the sharp hb transcription pattern emerges solely from diffusive biochemical interactions between the Bicoid transcription factor and the gene promoter region. Several alternative mechanisms have been proposed, such as additional sources of positional information, positive feedback from Hb proteins or out-of-equilibrium transcription activation. By using the MS2-MCP RNA-tagging system and analysing in real time, the transcription dynamics of synthetic reporters for Bicoid and/or its two partners Zelda and Hunchback, we show that all the early hb expression pattern features and temporal dynamics are compatible with an equilibrium model with a short decay length Bicoid activity gradient as a sole source of positional information. Meanwhile, Bicoid’s partners speed-up the process by different means: Zelda lowers the Bicoid concentration threshold required for transcriptional activation while Hunchback reduces burstiness and increases the polymerase firing rate.publishedVersionPeer reviewe

La proteine E2 des papillomavirus: de la fixation a l'ADN aux mecanismes d'activation et de repression de la transcription

SIGLEAvailable from INIST (FR), Document Supply Service, under shelf-number : T 84316 / INIST-CNRS - Institut de l'Information Scientifique et TechniqueFRFranc

Exploration moléculaire de l' activité du gradient morphogénétique Bicoid dans l' embryon précoce de drosophile

PARIS-BIUSJ-Thèses (751052125) / SudocPARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF

Le facteur d'assemblage de la chromatine CAF-1 (analyse fonctionnelle chez la drosophile)

La chromatine joue un rôle essentiel dans la stabilité du génome et lors de la différentiation. Mon travail a principalement consisté à caractériser les phénotypes d un mutant pour la grande sous-unité du facteur d assemblage de la chromatine CAF-1 chez la drosophile. Les larves mutantes meurent à cause de défauts s accumulant dans leurs cellules endocyclantes, alors que la progression de leur cycle cellulaire est normale. Ces cellules présentent des défauts d organisation des nucléosomes, d efficacité de la réplication des régions d euchromatine et d intégrité du génome. Cette étude a permis de corréler l assemblage des nucléosomes avec l efficacité du processus de réplication in vivo. Des résultats préliminaires indiquent aussi que les cellules mitotiques pourraient présenter des besoins variables en CAF-1, selon leur différentiation ou la nature de leur division. Enfin, j ai participé à la caractérisation de la fonction de l interaction entre CAF-1 et la protéine HP1a in vivo.PARIS-BIUSJ-Thèses (751052125) / SudocPARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF

Précision du gradient morphogénétique Bicoid dans l'embryon précoce de drosophile

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Constraints and limitations on decoding positional information: the Bicoid case-study

The regulation of expression of the hunchback promoter by the maternal Bicoid gradient has been studied as a model system in development for many years. Yet, at the level of quantitative agreement between data and theoretical models, even the first step of this regulation, transcription, continues to be challenging. This situation is slowly progressing, thanks to quantitative live-imaging techniques coupled to advanced statistical data analysis and modelling. Here we outline the current state our knowledge of this apparently “simple” step, highlighting the newly appreciated role of bursty transcription dynamics and its regulation

The replicative histone chaperone CAF1 is essential for the maintenance of identity and genome integrity in adult stem cells

International audienc

Precision in a rush: Trade-offs between reproducibility and steepness of the hunchback expression pattern

International audienceFly development amazes us by the precision and reproducibility of gene expression, especially since the initial expression patterns are established during very short nuclear cycles. Recent live imaging of hunchback promoter dynamics shows a stable steep binary expression pattern established within the three minute interphase of nuclear cycle 11. Considering expression models of different complexity, we explore the trade-off between the ability of a regulatory system to produce a steep boundary and minimize expression variability between different nuclei. We show how a limited readout time imposed by short developmental cycles affects the gene’s ability to read positional information along the embryo’s anterior posterior axis and express reliably. Comparing our theoretical results to real-time monitoring of the hunchback transcription dynamics in live flies, we discuss possible regulatory strategies, suggesting an important role for additional binding sites, gradients or non-equilibrium binding and modified transcription factor search strategies

Precision in a rush: Trade-offs between reproducibility and steepness of the hunchback expression pattern.

Fly development amazes us by the precision and reproducibility of gene expression, especially since the initial expression patterns are established during very short nuclear cycles. Recent live imaging of hunchback promoter dynamics shows a stable steep binary expression pattern established within the three minute interphase of nuclear cycle 11. Considering expression models of different complexity, we explore the trade-off between the ability of a regulatory system to produce a steep boundary and minimize expression variability between different nuclei. We show how a limited readout time imposed by short developmental cycles affects the gene's ability to read positional information along the embryo's anterior posterior axis and express reliably. Comparing our theoretical results to real-time monitoring of the hunchback transcription dynamics in live flies, we discuss possible regulatory strategies, suggesting an important role for additional binding sites, gradients or non-equilibrium binding and modified transcription factor search strategies