GSE126716
PARIS analysis of mouse Xist RNA structure
Long noncoding RNAs (lncRNAs) are an important class of transcripts that regulate gene expression on many levels, yet their mechanisms of action remain poorly understood. Previous studies have indicated that these RNAs can serve as modular scaffolds to recruit a variety of protein complexes and coordinate their distinct functions. XIST, a founding member of the lncRNA family, controls the inactivation of an entire X chromosome in placental mammals. Here we develop and integrate several orthogonal structure-interaction analysis methods to demonstrate that XIST RNA-protein complex folds into a modular architecture that is conserved in evolution. The discrete XIST RNA domains interact with distinct sets of effector proteins to orchestrate the X chromosome inactivation (XCI). The modular architecture plays an essential role, in addition to the sequence motifs, in determining the specificity of RBP binding and m6A modification. Together, this work builds a comprehensive structure-function model for the XIST RNA-protein complex, and establishes a paradigm for mechanistic studies of lncRNA functions. Overall design: Cells are crosslinked with AMT and RNA is extracted from cells by RNase digestion. Crosslinked RNA fragments are selected from total RNA by 2D gel electrophoresis. Purified crosslinked RNA duplexes are proximity ligated and crosslinking is reversed. Then the chimeric RNA molecules are converted into cDNA libraries for high-throughput sequencing.