Sarah E Harris, Yue Hu, Kaitlin Bridges, Francisco F Cavazos, Justin G Martyr, Bryan B Guzmán, Jernej Murn, Maria M Aleman, Daniel Dominguez
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引用次数: 0
Abstract
RNA-protein interactions are pivotal to proper gene regulation. Many RNA-binding proteins possess multiple RNA-binding domains; however, how these domains interplay to select and regulate RNA targets remains poorly understood. Here, we investigate three multi-domain proteins, Musashi-1, Musashi-2, and unkempt, which share a high degree of RNA specificity, a common feature across RNA-binding proteins. We used massively parallel in vitro assays with unprecedented depth with random or naturally-derived RNA sequences and find that individual domains within a protein can have differing affinities, specificities, and motif spacing preferences. We conducted large scale competition assays between these proteins and determined how individual protein specificities and affinities influence competitive binding. Integration of binding and regulation in cells with in vitro specificities showed that target selection involves a combination of the protein intrinsic specificities described here, but cellular context is critical to drive these proteins to motifs in specific transcript regions. Finally, evolutionarily conserved RNA regions displayed evidence of binding multiple RBPs in cultured cells, and these RNA regions represent the highest affinity targets. This work emphasizes the importance of in vitro and in cultured cells studies to fully profile RNA-binding proteins and highlights the complex modes of RNA-protein interactions and the contributing factors in target selection.
期刊介绍:
The Journal of Biological Chemistry welcomes high-quality science that seeks to elucidate the molecular and cellular basis of biological processes. Papers published in JBC can therefore fall under the umbrellas of not only biological chemistry, chemical biology, or biochemistry, but also allied disciplines such as biophysics, systems biology, RNA biology, immunology, microbiology, neurobiology, epigenetics, computational biology, ’omics, and many more. The outcome of our focus on papers that contribute novel and important mechanistic insights, rather than on a particular topic area, is that JBC is truly a melting pot for scientists across disciplines. In addition, JBC welcomes papers that describe methods that will help scientists push their biochemical inquiries forward and resources that will be of use to the research community.