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Center for Nonlinear Studies |
In the cell, two classes of macromolecules, RNA and protein, interact to form ribonucleoprotein complexes (RNPs) that have vital structural, catalytic and regulatory roles. Despite their functional importance, the mechanistic details and dynamics of RNPs are poorly understood. For example, alternative splicing is a highly regulated biological process that plays a crucial role in proteomic diversity in eukaryotes, but lacks a detailed mechanistic framework. Recently, single molecule techniques have been developed to investigate inter- and intramolecular interactions using Fluorescence Resonance Energy Transfer (FRET). Single-molecule FRET (smFRET) can provide information about heterogeneity within a population of molecules and dynamic behavior of single molecules, which cannot be observed directly from ensemble measurement. The goal of my research is to investigate protein-RNA interactions in alternative splicing as well as ribosome assembly, another protein-RNA interaction, by using FRET in combination with smFRET. The data show that Polypyrimidine tract binding protein (PTB) causes RNA looping upon binding, supporting a looping mechanism of exon exclusion. Furthermore, a PTB antagonist, Feminizing locus on X (Fox), competes with PTB to reduce looping and promote exon inclusion. Studies of a central junction in 16S rRNA and the primary binding protein that triggers a conformational change in rRNA, S15, show that mutations that drastically alter the junction dynamics at the S15 binding site affect 30S ribosome assembly.
Host: Brian Munsky (CNLS/CCS3) and Jim Werner (CINT)