Modulating transcriptional noise through genetic modification of eukaryotic promoters

SCHOR IE; IUNGMAN M

Salta

Congreso; Joint LV Annual SAIB Meeting and XIV PABMB Conference; 2019

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular - Panamerican Association for Biochemistry and Molecular Biology

The promoter regions of eukaryotic genes control both the average expression levels and their cell-to-cell variability, a quality known as intrinsic transcription noise. We are interested in understanding if this transcriptional noise can affect cellular processes, such as the coupling between transcription and RNA processing or the robustness of regulatory networks directing cellular processes like differentiation. In order to study this, we need to create tools in relevant model systems allowing the modulation of noise independently from transcript levels. In this project we will try to dissect the regulation of both parameters in mammalian promoters, through engineering synthetic variants based on evidence from molecular and genetic studies in yeast and flies. For the synthetic variants, we will test the effects of two features: the number and affinity of the binding site of an activating transcription factor and the addition of nucleosomal positioning sequences. We will use a dual fluorescent reporter to assess the functional properties of promoter variants, transfected in human and murine cells. The readout will be the relative fluorescence in single cells, measured through analytic flow cytometry. Finally, since the chromatin structure regulates both transcription levels and noise, we will study its impact on the different variants, comparing our constructs in episomal vs. genome-integrated context.