Experimental validation and characterization of putative targets of Escargot and STAT, two master regulators of the intestinal stem cells in Drosophila melanogaster.

Many organs contain adult stem cells (ASCs) to replace cells due to damage, disease, or normal tissue turnover. ASCs can divide asymmetrically, giving rise to a new copy of themselves (self-renewal) and a sister that commits to a specific cell type (differentiation). Decades of research have led to the identification of pleiotropic genes whose loss or gain of function affect diverse aspects of normal ASC biology. Genome-wide screens of these so-called genetic "master regulator" (MR) genes, have pointed to hundreds of putative targets that could serve as their downstream effectors. Here, we experimentally validate and characterize the regulation of several putative targets of Escargot (Esg) and the Signal Transducer and Activator of Transcription (Stat92E, a.k.a. STAT), two known MRs in Drosophila intestinal stem cells (ISCs). Our results indicate that regardless of bioinformatic predictions, most experimentally validated targets show a profile of gene expression that is consistent with co-regulation by both Esg and STAT, fitting a rather limited set of co-regulatory modalities. A bioinformatic analysis of proximal regulatory sequences in specific subsets of co-regulated targets identified additional transcription factors that might cooperate with Esg and STAT in modulating their transcription. Lastly, in vivo manipulations of validated targets rarely phenocopied the effects of manipulating Esg and STAT, suggesting the existence of complex genetic interactions among downstream targets of these two MR genes during ISC homeostasis.

Heterochromatin Protein 1 (HP1) inhibits stem cell proliferation induced by ectopic activation of the Jak/STAT pathway in the Drosophila testis.

Stem cells can divide asymmetrically with respect to cell fate, producing a copy of themselves (self-renewal), while giving rise to progeny that will differentiate along a specific lineage. Mechanisms that bias the balance towards self-renewal or extend the proliferative capacity of the differentiating progeny can result in tissue overgrowth and, eventually, the formation of tumors. Recent work has explored the role of heterochromatin and heterochromatin-associated proteins in the regulation of stem cell behavior under homeostatic conditions, but less is known about their possible roles in potentiating or suppressing stem cell overproliferation. Here we used ectopic activation of the Jak/STAT pathway in germline and somatic stem cells of the D. melanogaster testis as an in vivo model to probe the function of Heterochromatin Protein 1 (HP1) in stem cell overproliferation. Forced expression of HP1 in either early germ or somatic cells suppressed the overgrowth of testes in response to ectopic Jak/STAT activation. Interestingly, HP1 expression led to distinct phenotypes, depending on whether it was overexpressed in somatic or germ cells, possibly reflecting different cell-autonomous and non-autonomous effects in each cell type. Our results provide a new framework for further in vivo studies aimed at understanding the interactions between heterochromatin and uncontrolled stem cell proliferation, as well as the complex cross-regulatory interactions between the somatic and germline lineages in the Drosophila testis.