Title: Molecular Mechanisms of RNA Processing in the TRAMP RNA Surveillance Complex

Abstract:

The TRAMP (Trf4-Air2-Mtr4 Polyadenylation) complex promotes RNA turnover by delivering targeted substrates to the nuclear RNA exosome. TRAMP contains a poly(A) polymerase (Trf4) that polyadenylates the 3’ end of targeted RNAs and an RNA helicase (Mtr4) that translocates across the RNA in a 3’-5’ direction, presenting the polyadenylated 3’ end of the RNA substrate to the exosome for degradation or processing. The precise molecular mechanisms of TRAMP assembly and RNA shuttling between TRAMP catalytic sites are poorly understood.

Our efforts to understand TRAMP interactions combine multiple biochemical, biophysical, and structural biology approaches. We recently completed solution hydrogen-deuterium exchange (HDX) experiments alongside thermodynamic and functional assays to clarify details of TRAMP assembly in S. cerevisiae. This study reveals a critical role for the Mtr4 Arch domain in complex assembly and function. We further show that tRNA binding by single active-site subunits, Mtr4 and Trf4-Air2, is altered upon TRAMP assembly. Additionally, we describe a working model for the TRAMP complex based on available biochemical and structural data, combined with AlphaFold predictions. Deletion mutagenesis and domain swapping experiments demonstrate species dependent differences and commonalities in TRAMP assembly and helicase activation. Overall, this work demonstrates that a wide range of temporal and conformational dynamics are associated with TRAMP interactions and RNA processing.

Bios:

Sean Johnson earned a Bachelor of Science in Chemistry with a Biochemistry emphasis from Utah State University in 1996. He obtained a Ph.D. in Biochemistry from Duke University in 2003 under the mentorship of Dr. Lorena Beese. Following this, he conducted postdoctoral research in Dr. Chris Hill’s lab at the University of Utah. In 2006, he joined Utah State University as the R. Gaurth Hansen Assistant Professor in the Department of Chemistry & Biochemistry. He currently serves as an Associate Professor of Chemistry and Biochemistry and Associate Dean of Graduate Studies in the College of Science at Utah State University.