I-KCKT allows dissection-free RNA profiling of adult Drosophila intestinal progenitor cells

Abstract

The adult Drosophila intestinal epithelium is a model system for stem cell biology, but its utility is limited by current biochemical methods that lack cell type resolution. Here, we describe a new proximity-based profiling method that relies upon a GAL4 driver, termed intestinal-kickout-GAL4 (I-KCKT-GAL4), exclusively expressed in intestinal progenitor cells. This method used UV cross-linked whole animal frozen powder as its starting material to immunoprecipitate the RNA cargoes of transgenic epitope-tagged RNA binding proteins driven by I-KCKT-GAL4. When applied to the general mRNA-binder, poly(A)-binding protein, the RNA profile obtained by this method identified 98.8% of transcripts found after progenitor cell sorting, and had low background noise despite being derived from whole animal lysate. We also mapped the targets of the more selective RNA binder, Fragile Mental Retardation Protein, using enhanced CLIP, and report for the first time its binding motif in Drosophila cells. This method will therefore enable the RNA profiling of wildtype and mutant intestinal progenitor cells from intact flies exposed to normal and altered environments, as well as the identification of RNA-protein interactions critical for stem cell function.