My lab is creating technology to image gene expression in real time and with single-molecule
precision in living cells. Using tandem-repeat mRNA and protein tags, genetically
encoded intrabodies, and single-molecule microscopy, we now regularly image the live-cell
translation dynamics of single mRNAs. In this talk, I will introduce our technology and describe
how it can be used to amplify fluorescent signals within single translation sites. I will show how
we quantify these signals to determine the size, shape, subcellular localization, and mobilities of
translation sites, as well as their ribosomal densities and translational initiation and elongation
kinetics. I will then highlight a few recent applications of our technology, including the imaging
of translation shutdown during cellular stress, HIV-1 translational frameshifting, and
non-canonical translation initiation at internal ribosomal entry sites. I will conclude by
discussing new intrabodies my lab has engineered that bind the classic HA and FLAG epitopes in
living cells. As these intrabodies can be encoded on plasmids, they can easily be adapted
by other labs to image translation in multiple colors and in diverse living systems.
Timothy J. Stasevich is an Associate Professor in the Department of Biochemistry and Molecular
Biology at Colorado State University (CSU). His lab uses a combination of advanced fluorescence
microscopy, genetic engineering, and computational modeling to study the dynamics of
gene regulation in living mammalian cells. His lab helped pioneer the imaging of
real-time single- mRNA translation dynamics in living cells¹. Dr. Stasevich received his
B.S. in Physics and Mathematics from the University of Michigan, Dearborn, and his Ph.
D. in Physics from the University of Maryland, College Park. He transitioned into
experimental biophysics as a post- doctoral research fellow in the laboratory of Dr. James G.
McNally at the National Cancer Institute. During this time, he developed technology based on
fluorescence microscopy to help establish gold-standard measurements of live-cell protein dynamics.
Dr. Stasevich next moved to Osaka University, where he worked with Dr. Hiroshi Kimura as a Japan
Society for the Promotion of Science Foreign Postdoctoral Research Fellow. While there, he helped
create technology to image endogenous proteins and their post-translation modifications in vivo.
This allowed him to image the live-cell dynamics of epigenetic histone modifications during gene
activation for the first time². Before joining the faculty at CSU, Dr. Stasevich spent a year as a
Visiting Fellow at the HHMI Janelia Research Campus, where he applied superresolution fluorescence
microscopy to improve the spatiotemporal resolution of endogenous protein imaging in live cells.
1. Morisaki, T. et al. Real-time quantification of single RNA translation dynamics in living cells.Science 352, 1425–1429 (2016).
2. Stasevich, T. J. et al. Regulation of RNA polymerase II activation by histone acetylation in
single living cells. Nature 516, 272–275 (2014).
This lecture will be held via ZOOM:
Meeting ID: 996 8526 5039, Password: 480845
Please contact Sharlene Denos at denos@illinois.edu with questions or for more information.
