Furthermore, mis-regulation of some of these direct targets contributes to the expanded stem cell phenotype observed in Lsd1 mutants. These experiments revealed that Lsd1 exhibits highly enriched binding to over one hundred genomic sites within a specific cell population. To determine direct regulatory targets of Lsd1, we employed chromatin immunoprecipitation coupled with massive parallel sequencing (ChIP-seq) using specific cell populations inside and outside of the GSC niche. Previous work has shown that loss of the highly conserved histone demethylase Lsd1 results in ectopic niche signaling and an expanded GSC phenotype. Studies of Drosophila germline stem cells (GSCs) have suggested that chromatin programming greatly influences the behavior of these cells and their progeny.

The mechanisms that govern the formation, size and signaling output of in vivo niches remain poorly understood. These results suggest that Lsd1 restricts the number of GSC-like cells by regulating a diverse group of genes and provide further evidence that escort cell function must be carefully controlled during development and adulthood to ensure proper germline differentiation. Knockdown of other potential direct Lsd1 target genes, not obviously linked to BMP signaling, also partially suppresses the Lsd1 mutant phenotype. Interestingly, over-expression of engrailed, but not hedgehog, results in an expansion of GSC cells, marked by the expansion of BMP signaling. Instead, Lsd1 regulates engrailed expression and disruption of engrailed and its putative downstream target hedgehog suppress the Lsd1 mutant phenotype. Lsd1 does not directly target the dpp locus in escort cells. These findings indicate that Lsd1 exhibits highly selective binding that depends greatly on specific cellular contexts. This analysis shows that Lsd1 associates with a surprisingly limited number of sites in escort cells and fewer, and often, different sites in cap cells. To better characterize the function of Lsd1 in different cell populations within the ovary, we performed Chromatin immunoprecipitation coupled with massive parallel sequencing (ChIP-seq). Here we present evidence that loss of Lsd1 also results in gradual changes in escort cell morphology and their eventual death. Loss of the highly conserved histone demethylase Lsd1 in Drosophila escort cells results in increased BMP signaling outside the cap cell niche and an expanded germline stem cell (GSC) phenotype. However the mechanisms that govern the formation, size and signaling of in vivo niches remain poorly understood. Specialized microenvironments called niches regulate tissue homeostasis by controlling the balance between stem cell self-renewal and the differentiation of stem cell daughters.

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