Robert H. Singer, Timothée Lionnet, Hye Yoon Park, Bin Wu, Tatjana Trcek, Sami Hocine and Daniel Larson
Department of Anatomy and Structural Biology and Gruss Lipper Biophotonics Center, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY
Imaging has been an essential tool to analyze the dynamic properties of RNA. New technologies in optical microscopy and fluorescent probe development to detect and track individual mRNA molecules in single living cells have yielded insights that could not have been obtained through any ensemble measurement, for instance in measuring the time for transcription elongation or nuclear pore transport. We have been dedicated to developing and implementing these technologies to further the understanding of dynamics of mRNA regulation in cells. We have utilized various computational approaches to analyze the components of real-time transcription of endogenous genes in living tissues of a transgenic mouse where the mRNAs are labeled by a fluorescent RNA binding protein. These approaches employ a plethora of imaging techniques, ranging from confocal and multiphoton microscopy, long-term cell imaging, high-speed real-time widefield microscopy, single molecule tracking, super-registration microscopy and fluorescence fluctuation analysis. The kinetics of the key elements of RNA metabolism: initiation, elongation, termination, as well as nuclear pore export, cytoplasmic trafficking, localization, translation and decay are now tractable at the single molecule level in vivo. By using mathematical modeling to extract the quantitative parameters of these kinetics, we can describe these processes precisely in living cells.
Supported by NIH GMS Grants to RH Singer