Targets and functions of the mammalian snoRNAome

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Project Abstract: Small nucleolar RNA (snoRNA) are natural guide non-coding RNAs derived from over 900 annotated snoRNA genes in the human genome. The major known function of about one-third of snoRNAs is to install 2’O-methyl and pseudouridine modifications in the ribosomal RNA. These rRNA modifications guide ribosome assembly and maturation, and fine-tune translation in a cell type and cell state dependent manner. However, the majority of snoRNAs do not have a well described function; they are called orphans because their cellular RNA targets are not known. How snoRNA interacts with the human transcriptome and the functional consequences of these potential non-canonical snoRNA-RNA interactions in gene expression regulation and human diseases remain to be determined. We recently developed several new sequencing technologies that are directly relevant to studying snoRNA biology. They include methods to robustly identify inter-molecular RNA-RNA interactions in cells and transcriptome-wide sequencing of 2’O-methyl and pseudouridine modifications. These new tools will allow us to address mysteries of snoRNA biology. Aim 1 will identify the cellular RNA targets of the snoRNAome through an advanced version of kethoxal-assisted RNA-RNA interaction sequencing (KARR-seq) approach to enable comprehensive capture and identification of snoRNA-RNA interactions at the transcriptomic scale. These results will be used to identify the rules of snoRNA-guided targeting of mRNA sequences. Aim 2 will investigate two types of snoRNA-mRNA interactions transcriptome-wide and the consequences on regulating gene expression through the application of new sequencing methods to map 2’O-methyl and pseudouridine modifications in pre-mRNA and mature mRNA, and associate these results with the snoRNA-mRNA interactome. We will also selectively delete snoRNA genes or knockdown snoRNA levels, alter the expression of snoRNP components to investigate the consequence of specific snoRNA-mRNA interaction. All together these results will provide the first comprehensive snoRNA-RNA interactome and derive the guiding principles of snoRNA-mRNA interactions and the associated biological functions.