Abstract
We used live imaging to visualize the transcriptional dynamics of the Drosophila melanogaster even-skipped gene at single-cell and high-temporal resolution as its seven stripe expression pattern forms, and developed tools to characterize and visualize how transcriptional bursting varies over time and space. We find that despite being created by the independent activity of five enhancers, even-skipped stripes are sculpted by the same kinetic phenomena: a coupled increase of burst frequency and amplitude. By tracking the position and activity of individual nuclei, we show that stripe movement is driven by the exchange of bursting nuclei from the posterior to anterior stripe flanks. Our work provides a conceptual, theoretical and computational framework for dissecting pattern formation in space and time, and reveals how the coordinated transcriptional activity of individual nuclei shapes complex developmental patterns.
Highlights
The patterns of gene expression that choreograph animal development are formed dynamically by an interplay between processes – transcription, mRNA decay and degradation, diffusion, directional transport and the migration of cells and tissues – that vary in both space and time
The classical view of transcription as a switch or a tunable rheostat has been replaced in recent years by the recognition that mRNA synthesis occurs in bursts, with promoters switching stochastically between an ON state where polymerases are loaded and begin elongating, and an OFF state where few or no new transcripts are initiated (Figure 1A; Zenklusen et al, 2008; Golding et al, 2005; Blake et al, 2006; Janicki et al, 2004; Chubb et al, 2006; Yunger et al, 2010; Raj et al, 2006; Lionnet et al, 2011; Muramoto et al, 2012; Little et al, 2013; Xu et al, 2015; Lenstra et al, 2015; Fukaya et al, 2016; Desponds et al, 2016; Hendy et al, 2017; Lammers et al, 2020)
We used recombineering (Warming et al, 2005) to modify a bacterial artificial chromosome (BAC) (Venken et al, 2006) containing the D. melanogaster eve gene and all of its enhancers and regulatory elements (Venken et al, 2009), replacing the coding region with an array of MS2 stem loops followed by the D. melanogaster yellow (y) gene (Figure 1D; Bothma et al, 2014)
Summary
The patterns of gene expression that choreograph animal development are formed dynamically by an interplay between processes – transcription, mRNA decay and degradation, diffusion, directional transport and the migration of cells and tissues – that vary in both space and time. The spatial aspects of transcription have dominated the study of developmental gene expression, with the role of temporal processes in shaping patterns receiving comparably little attention (Bothma and Levine, 2013; Garcia et al, 2020). Gene expression patterns are dynamic on many levels. They form, change and disappear over time, often as cells, tissues, and organs are forming and moving in the developing embryo (Lawrence, 1992). The transcriptional process that creates these patterns is itself highly dynamic. The classical view of transcription as a switch or a tunable rheostat has been replaced in recent years by the recognition that mRNA synthesis occurs in bursts, with promoters switching stochastically between an ON state where polymerases are loaded and begin elongating, and an OFF state where few or no new transcripts are initiated (Figure 1A; Zenklusen et al, 2008; Golding et al, 2005; Blake et al, 2006; Janicki et al, 2004; Chubb et al, 2006; Yunger et al, 2010; Raj et al, 2006; Lionnet et al, 2011; Muramoto et al, 2012; Little et al, 2013; Xu et al, 2015; Lenstra et al, 2015; Fukaya et al, 2016; Desponds et al, 2016; Hendy et al, 2017; Lammers et al, 2020)
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