The development and application of methods to track the dynamics of cellular RNAs. Our work has contributed to our understanding of RNA population dynamics. RNAs are continuously transcribed and degraded, yet methods such as RNA-sequencing provide only a snapshot of RNA abundance. To reveal the rich regulated dynamics of the transcriptome, we have developed chemical approaches that improve on the use of 4-thiouridine metabolic labeling (Duffy et al, Mol Cell 2015). Through collaboration, we used this improved disulfide chemistry to study the role of m6A modification in the immune response (Li et al, Nature, 2017). Most recently we have developed a nucleoside recoding strategy to convert s4U to cytosine analogues (Schofield et al. Nature Methods, 2018). This mutational mapping approach allows analysis of RNA population dynamics without the need for biochemical enrichment. Our work has uncovered isoform-specific differences in the turnover of a disease related transcript that encodes for the polycomb/trithorax protein, ASXL1.
(Figure adapted from Duffy et al. (2018) Wiley Interdiscip Rev)
Schofield, J. A., Duffy, E. E., Kiefer, L., Sullivan, M. C. and Simon, M. D., TimeLapse-seq: Chemistry to Add a Temporal Dimension to RNA-seq. (2018) Nature Methods, doi:10.1038/nmeth.4582. PMID: 29355846
Duffy, E. E., Rutenberg-Schoenberg, M., Stark, C. D., Kitchen, R. R., Gerstein, M. B., and Simon, M. D., Tracking distinct RNA populations using efficient and reversible covalent chemistry. (2015) Mol Cell, 59(5), 858-66. PMID: 26340425.
Li, H. B., Tong, J., Zhu, S., Batista, P. J., Duffy, E. E., Zhao, J., Bailis, W., Cao, G.; Kroehling, L., Chen, Y., Wang, G., Broughton, J. P., Chen, Y. G., Kluger, Y.,Simon, M. D., Chang, H. Y., Yin, Z. and Flavell, R. A., m(6)A mRNA methylation controls T cell homeostasis by targeting the IL-7/STAT5/SOCS pathways. (2017) Nature, 548(7667), 338-342.