Live Imaging of Bicoid-Dependent Transcription in Drosophila Embryos

Abstract

The early Drosophila embryo is an ideal model to understand the transcriptional regulation of well-defined patterns of gene expression in a developing organism. In this system, snapshots of transcription measurements obtained by RNA FISH on fixed samples cannot provide the temporal resolution needed to distinguish spatial heterogeneity from inherent noise. Here, we used the MS2-MCP system to visualize in living embryos nascent transcripts expressed from the canonical hunchback (hb) promoter under the control of Bicoid (Bcd). The hb-MS2 reporter is expressed as synchronously as endogenous hb in the anterior half of the embryo, but unlike hb it is also active in the posterior, though more heterogeneously and more transiently than in the anterior. The length and intensity of active transcription periods in the anterior are strongly reduced in absence of Bcd, whereas posterior ones are mostly Bcd independent. This posterior noisy signal decreases progressively through nuclear divisions, so that the MS2 reporter expression mimics the known anterior hb pattern at cellularization. We propose that the establishment of the hb pattern relies on Bcd-dependent lengthening of transcriptional activity periods in the anterior and may require two distinct repression mechanisms in the posterior.

Modulation of temporal dynamics of gene transcription by activator potency in theDrosophilaembryo

“…There are two aspects of the temporal dynamics of a gene's transcription in a developmental system: onset and duration (Garcia et al, 2013;Liu and Ma, 2013b;Lucas et al, 2013). Proper control of the temporal dynamics of transcription is particularly important for developmental systems that progress rapidly, such as the early Drosophila embryo (Foe and Alberts, 1983;Guilgur et al, 2014).…”

Mitosis-associated repression in development

“…Different mechanisms have been documented, including competition between DNA-binding proteins, "quenching" of activators bound to adjacent sites within target enhancers, or direct repression of the promoter (Levine and Manley 1989;Gray et al 1994;Gray and Levine 1996a,b). Newly developed live-imaging methods permit the first opportunity to explore the temporal dynamics of these repression mechanisms in living embryos (Garcia et al 2013;Lucas et al 2013).…”

Experimental and Modeling Approaches for Understanding the Effect of Gene Expression Noise in Biological Development

“…The MCP signal was sampled every 30-60 s and calibrated to the number of active PolII molecules. Peak transcription (up to 100 transcripts being made) can occur within 1-2 min after nuclear division [52]. hb output corresponds well to transcriptional initiation rates up through interphase 13, but the large increase in output entering interphase 14 indicates an additional contribution from whether a nucleus is active or not [51].…”

“…Time series were first measured in Drosophila for hb [51,52]. The MCP signal was sampled every 30-60 s and calibrated to the number of active PolII molecules.…”