Transcriptome profiling of Drosophila melanogaster midgut cell populations by mRNA sequencing. Devanjali Dutta, Bruce Edgar. CELL GROWTH AND PROLIFERATION, ZMBH-DKFZ, HEIDELBERG, BADEN-WÜRTTEMBERG, Germany.

   Drosophila midgut is a rapidly proliferating tissue and is maintained by a continuous supply of differentiated cells that originate from intestinal stem cells (ISCs).ISCs proliferate throughout life, renewing and generating transient cells called enteroblasts (EBs), which differentiate into enterocytes (ECs) or enteroendocrine cells (EEs). The proper regulation of intestinal stem cell maintenance, proliferation and differentiation is critical for maintaining gut homeostasis.Notch knockdown produces tumors,however the identity of these tumors is unclear.We have developed a method for isolating the intestinal stem cells, progenitors and all the differentiated cell types using Fluorescent activated cell sorting (FACS).Analysis of our comparitive cell-type transcriptome data suggests that the genetic differences between the intestinal cell types in the adult Drosophila midgut are not completely classified or graded.Interestingly only 17 genes were found to be specific to Myo1A+ ECs ,whereas the How + visceral muscle seemed to be expressing 911 specific genes.We observed that the Dl+ISCs, Su(H)+EBs and the Rab3+EEs had a reasonable 104, 319 and 168 cell type specific genes respectively. Presence of known cell type specific genes in our filtered dataset confirms that pure populations of cells were FACS sorted and that our expression profiling of the intestinal cell populations is accurate.mRNA sequencing of sorted esg+ tumor cells revealed that ISC tumors expressed a number of ISC markers such as Dl, spdo, esg, shg & EE specific markers like ase, sc, Ast, Pros and Rab3.We also found increased expression of the EGFR receptor,spi,Krn and factors like rhomboids 1, 4, 5 and Star.We compared the whole transcriptomes of esg+ tumor cells with those of Dl+ ISCs, SuH+ enteroblasts or esg+ cells from wild-type flies, and found that esg+ tumor cells were more similar to the esg+ population (83.8%) and to SuH+ enteroblasts (76.3%)than to wild-type Dl+ ISCs (61.8%). These comparisons suggest that these tumor cells are proliferative progenitors or precursors but not identical to wild-type Dl+ ISCs.