title: Deciphering Transcriptional Dynamics In Vivo by Counting Nascent RNA Molecules.
abstract: Deciphering how the regulatory DNA sequence of a gene dictates its expression in response to intra and extracellular cues is one of the leading challenges in modern genomics. The development of novel single-cell sequencing and imaging techniques, as well as a better exploitation of currently available single-molecule imaging techniques, provides an avenue to interrogate the process of transcription and its dynamics in cells by quantifying the number of RNA polymerases engaged in the transcription of a gene (or equivalently the number of nascent RNAs) at a given moment in time. In this paper, we propose that measurements of the cell-to-cell variability in the number of nascent RNAs provide a mostly unexplored method for deciphering mechanisms of transcription initiation in cells. We propose a simple kinetic model of transcription initiation and elongation from which we calculate nascent RNA copy-number fluctuations. To demonstrate the usefulness of this approach, we test our theory against published nascent RNA data for twelve constitutively expressed yeast genes. Rather than transcription being initiated through a single rate limiting step, as it had been previously proposed, our single-cell analysis reveals the presence of at least two rate limiting steps. Surprisingly, half of the genes analyzed have nearly identical rates of transcription initiation, suggesting a common mechanism. Our analytical framework can be used to extract quantitative information about dynamics of transcription from single-cell sequencing data, as well as from single-molecule imaging and electron micrographs of fixed cells, and provides the mathematical means to exploit the quantitative power of these technologies.
authors.affiliation: Department of Physics, Brandeis University, Waltham, Massachusetts, United States of America.
authors.forename: Sandeep
authors.initials: S
authors.lastname: Choubey
authors.affiliation: Department of Physics, Brandeis University, Waltham, Massachusetts, United States of America.
authors.forename: Jane
authors.initials: J
authors.lastname: Kondev
authors.affiliation: Rowland Institute at Harvard, Harvard University, Cambridge, Massachusetts, United States of America.
authors.forename: Alvaro
authors.initials: A
authors.lastname: Sanchez
mesh_terms: D000465:Algorithms; D019295:Computational Biology / Q000379:methods*; D003593:Cytoplasm / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D008957:Models, Genetic; D012313:RNA / Q000032:analysis* / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic / Q000235:genetics*; D015003:Yeasts / Q000235:genetics / Q000378:metabolism
chemical_list: D012313:RNA; D012321:DNA-Directed RNA Polymerases