The main interest of the Lenstra lab is to understand the mechanisms of transcription regulation in eukaryotic cells. In the last half century, our knowledge of gene expression regulation has greatly advanced, starting from pioneering molecular biology experiments on the lac operon and culminating in the explosion of genome-wide data collection. The majority of these measurements are coming from large populations of cells, providing a static snapshot of the average behavior of a biological process. However, the transcriptional process is highly dynamic, with transcription complexes assembling and disassembling on DNA with very rapid time scales. This stochastic behavior of individual macromolecules results in heterogeneity in transcriptional responses and cellular decisions, even for genetically identical cells in the same environment.
We use cutting-edge single-molecule microscopy techniques to visualize the behavior of individual protein and RNA molecules in living cells, providing direct quantitative measurements of the regulatory steps of transcription. These imaging assays are combined with genetic perturbations or novel gene-specific targeting approaches to modulate transcription of endogenous genes. For example, we used time-lapse single-molecule imaging of sense and antisense (non-coding) transcription in living cells in tandem with strand-specific transcriptional blocking of the ncRNA by CRISPR/dCas9 to interrogate the role of ncRNA transcription in the yeast galactose response. By studying transcription dynamics in single cells we aim to understand the molecular mechanisms of transcription regulation, and how stochasticity in RNA synthesis modulates cell-to-cell variability and contributes to disease progression.