Alternate Grainy head isoforms regulate Drosophila midgut intestinal stem cell differentiation

Regeneration of the Drosophila midgut epithelium depends upon differential expression of transcription factors in intestinal stem cells and their progeny. The grainy head locus produces multiple splice forms that result in production of two classes of transcription factor, designated Grh.O and Grh.N. grainy head expression is associated with epithelial tissue and has roles in epidermal development and regeneration but had not been examined for a function in the midgut epithelium. Here we show that null mutant clones had a limited effect on intestinal stem cell (ISC) maintenance and proliferation but surprisingly specific loss of all Grh.O isoforms results in loss of ISCs from the epithelium. This was confirmed by generation of a new Grh.O class mutant to control for genetic background effects. Grh.O mutant ISCs were not lost due to cell death but were forced to differentiate. Ectopic expression of a Grh.N isoform also resulted in ISC differentiation similar to loss of Grh.O function. Grh.O expression must be tightly regulated as high level ectopic expression of a member of this isoform class in enteroblasts, but not ISCs, resulted in cells with confused identity and promoted excess proliferation in the epithelium. Thus, midgut regeneration is not only dependent upon signalling pathways that regulate transcription factor expression, but also upon regulated mRNA splicing of these genes.