NIH - Oxford
National Institutes of Health Investigator:
Primary Research Area:
Genetics and Epidemiology
Secondary Research Area:
Immunology and Infectious Disease
High-throughput identification of microRNA targets in T lymphocytes
We are interested in regulatory networks that govern how immune cells differentiate and function. Post-transcriptional regulation by microRNAs can be critical for a cell’s gene expression program. Unfortunately, deciphering the direct targets of microRNAs remain challenging. Phylogenomic analyses have been used to identify genes with 3’ UTRs that harbor evolutionarily conserved motifs that may serve as potential binding sites for microRNAs. However, in practice, such predictions have a high false positive rate. Although the microRNA-target interactions may in fact occur, they may not result in a detectable functional effect in a given cell under a given condition for a variety of reasons. Therefore, we will undertake systematic experimental approaches that take advantage of recent advances in genomics and proteomics to identify microRNA targets that change in expression in T cells. Following perturbation of a microRNA or pathway, we can perform RNA-seq and shotgun proteomics to identify changes in the whole transcriptome and proteome. Linear regression analysis of the data can be used to identify correlations between expression changes and sequence motifs within 3’UTRs (eg. microRNA seed binding sites). Additionally, two recently developed methods referred to as HITS-CLIP and PAR-CLIP can identify RNA sequences that directly interact with the Argonaute-containing miRNA-induced silencing complex (miRISC). Systematic integration of the resulting large data sets will provide a valuable network model of microRNA-mediated regulation.