Characterizing the regulatory logic of transcriptional control at the DNA sequence level by ensembles of thermodynamic models

Characterizing how the genome encodes the regulatory logic of

transcription is a main challenge of the post-genomic era, which

can be overcome with the aid of theoretical physics tools. Gene

regulation is crucial for embryo development robustness. E.g. pattern

formation along the antero-posterior axis (AP) of Drosophila

embryos happens with striking single nuclei precision during

syncytial blastoderm stage. The pair-rule gene even-skipped (eve)

participates of this process forming seven stripes transverse to

AP observable ~3 hours after egg deposition. Under proper scaling, each stripe is

described by four well-conserved parameters determining its

location, width, intensity, and time of formation. That

simplicity contrasts with the intricate combination of

experimentally observed regulatory mechanisms encoded in specific

enhancers within eve's 16kb loci and challenges us to formulate general models about regulation of expression in metazoans. In this talk we discuss how an

ensemble of fits to data produced by application of simulated

annealing to optimize the parameters of a thermodynamics-based

sequence-level model aids understanding transcriptional

regulation. Quantitative experimental data on reporters driven by

the whole locus of the eve gene in the blastoderm of Drosophila

embryos was used for validating our approach. The fits are

clustered accordingly with their intrinsic regulatory logic. A

multiscale analysis enables visualization of quantitative

features resulting from the deconvolution of the regulatory

profile emergent from the interaction of multiple transcription

factors with the locus of eve. A few clusters of highly active

DNA binding sites within the enhancers collectively modulate

eve's transcription. Analysis of variable enhancers’

length shows the importance of DNA-bound protein-protein interactions

for transcriptional regulation. The interplay between activation

and quenching enables function conservation of enhancers in different species of Drosophila.