Recent studies have provided unprecedented insights into how individual molecular mechanisms are correlated to changes in gene expression levels. It is now recognized that substantial transcriptome complexity arises from decisions involved in the processing of nascent mRNAs. However, by and large, the many studies of gene expression have focused on characterizing steady-state mRNA levels, even though living systems are inherently dynamic. Thus, a key open question in functional genomics is: how to the dynamics of individual molecular mechanisms combine to influence the cellular transcriptome? Our overarching hypothesis is that the fate of an mRNA is governed by kinetic interactions underlying molecular efficiency across disparate steps in the mRNA lifecycle ? at the core, transcription elongation, mRNA splicing, and 3? end cleavage. Current knowledge of spatiotemporal coordination across these processes likely represents the tip of the iceberg. In this proposal, we will combine genetic, molecular, and cellular genomic techniques with high-dimensional computational analyses to address two specific themes of mechanistic efficiency. First, we aim to dissect the kinetic basis for efficient mRNA biogenesis and maturation. Work from my research and others has suggested that there is substantial variability in kinetic actions and rate-limiting steps for mature mRNA production across genes. Determining the molecular logic underlying this variability will inform our understanding of the molecular basis for transcriptome complexity in both homeostatic and diseased cells. We will address this issue by answering three questions. 1) What are the kinetics of each individual step in mRNA processing?; 2) What are the factors that govern rate-limiting steps of mature mRNA production; and 3) How does the RNA processing machinery balance speed and accuracy? The goal of the second theme of our research is to determine how RNA processing kinetics underlie the regulation of cellular transitions. The kinetics of gene regulatory processes likely play a crucial role in cellular responses or transitions, by altering the order of molecular events or speed of processes to rapidly regulate gene expression dynamics. To dissect the kinetic plasticity underlying cellular response, we will answer related questions in two disparate systems: 4) How do immune cells rapidly change their transcriptome upon stimulation?; and 5) How is transcriptome complexity regulated upon neuronal differentiation? Our research will result in insights crucial to uncovering the cascade of molecular events that cumulatively establish steady- state gene expression levels, with implications for improved therapeutic designs or strategies that target the early progression of cellular shifts rather than only the final outcome.
Phenotypic complexity across cells, tissues, and disease states is driven by coordinated changes in gene regulation that allow a continuum of gene expression levels to arise from the same genome sequence. Though profiles of steady-state gene expression levels are central to our understanding of the molecular underpinnings of developmental processes, immune response, cancer, or the efficiency of drug targeting, they do not fully capture the dynamic nature of cellular processes. This proposal focuses on understanding the kinetic interplay between steps of RNA biogenesis and maturation, in particular, the speed and accuracy of mechanisms underlying responsible for establishing mRNA levels in cells.
