The Rna Binding Protein Msi2 Has Increased Rna Binding Activity In Leukemic Stem Cells Compared To Normal Hematopoietic Stem Cells

The RNA binding protein (RBP) MSI2 has been demonstrated to regulate the self-renewal activity and differentiation program of normal hematopoietic stem and progenitor cells (HSPCs), as well as leukemic stem cells (LSCs) (Kharas et al, Nature Medicine 2010, Park et al, 2013 Journal of Experimental Medicine, Park et al JCI 2015). Yet, the extent of MSI2 targets and their differential activity in these rare cells are not known. Standard techniques for identifying RBP targets, such as RNA-IP and CLIP, require large cell numbers (>10 million) and are technically challenging. Here we employed HyperTRIBE, a recently discovered method (McMahon et al. 2016 and Xu et al., 2017), to globally map the MSI2 targeting network in HSPCs and LSCs. In HyperTRIBE, the catalytic domain of the Drosophila ADAR (adenosine deaminase enzyme) is fused with an RBP. This fusion protein leaves a “fingerprint” on the RBP RNA targets by marking the binding sites with an editing A-to-G event. HyperTRIBE was originally developed in Drosophila, but it is not known whether it can be used as a tool for mapping RBP targets in mammalian systems.