Antoine Coulon (Institut Curie, Paris). Biophysics seminar ESPCI-ENS

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6 April 13:00 » 14:00 — Conf IV, Ecole Normale Supérieure

Probing the spatiotemporal kinetics of transcription at the single-RNA level

The synthesis of mature messenger RNA molecules from the DNA sequence of a gene is a central step in gene expression and is regulated at many levels (transcription initiation, elongation, splicing...). Yet, at the single-gene level, the stochastic kinetics of such regulatory processes and their coordination in space and time remain poorly understood.

Using the RNA-labeling techniques MS2 and PP7, we can now image transcription “in 4D” to probe the processes involved in mRNA synthesis. More specifically, with this approach, we are able to visualize and monitor in real-time the position and the amount of nascent transcripts from individual genes of interest, with high temporal resolution and single-molecule sensitivity. The possibility of using two colors (e.g. to label either two distinct parts of a single gene or two different genes) opens up a vast range of possibilities to study the kinetics and coordination between the molecular mechanisms involved in transcriptional regulation. A methodological challenge is the analysis and interpretation of the resulting datasets, i.e. stochastic and noisy transcriptional time traces. To this end, we also develope a method based on auto/cross-correlation analysis to reveal the signatures of the underlying molecu­lar processes and extract quantitative information about single-RNA transcription kinetics.

I will present these imaging and data analysis approaches and how we applied them to understand the kinetics of single-RNA synthesis, sense/antisense transcription and gene bursting, as well as our future directions to study transcriptional coordination in four dimensions.

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