Erika A. Bach

Aging tissues lose function in part because stem cells change in number and behavior, but how age-related changes in the stem cell niche drive these processes is not well understood. Using the fruit fly testis, we asked how aging of the niche microenvironment influences stem cell maintenance and competition. We show that levels of niche cell–derived bone morphogenetic protein (BMP) signals decline with age, leading to increased expression of the transcriptional corepressor Hairless in germline stem cells (GSCs). Elevated Hairless reduces the RNA binding protein Imp, causing loss of the stem cell factor Chinmo and triggering abnormal extracellular matrix accumulation, stem cell displacement, and niche deterioration. Overexpressing BMP signals in the niche, up-regulating Imp in GSCs, or depleting Hairless in GSCs ameliorate multiple aging-related defects. In contrast, GSC clones with low Imp or high Hairless outcompete their neighbors and take over the niche. These findings reveal how aging niche signals affect tissue decline and stem cell residence.

Maintenance of somatic sex identity is essential for adult tissue function. In the Drosophila testis, adult somatic stem cells known as cyst stem cells (CySCs) require the transcription factor Chinmo to preserve male identity. Loss of Chinmo leads to reprogramming of CySCs into their ovarian counterparts through induction of the female-specific RNA-binding protein Transformer ^F (Tra ^F ), though the underlying mechanism has remained unclear. Here, we identify the pioneer transcription factor Zelda (Zld) as a critical mediator of this sex reversal. In wild-type CySCs, zld mRNA is repressed by microRNAs (miRs), but following Chinmo loss, these miRs are downregulated, allowing zld mRNA to be translated. Zld is necessary for feminization of chinmo -mutant CySCs, and ectopic expression of Zld in wild-type CySCs is sufficient to induce Tra ^F and drive female reprogramming. Two Zld target genes, qkr58E-2 and Ecdysone receptor ( EcR ), are upregulated in chinmo -mutant CySCs and are normally female-biased in adult gonads. Qkr58E-2 facilitates Tra ^F production, while EcR promotes female gene expression programs. Zld overexpression feminizes otherwise wild-type CySCs by upregulating EcR, which in turn downregulates the chinmo gene. Strikingly, overexpression of Zld also feminizes adult male adipose tissue by inducing Tra ^F and downregulating Chinmo, indicating that Zld can override male identity in multiple adult XY tissues. Together, these findings uncover a post-transcriptional mechanism in which miRs-mediated repression of a pioneer factor safeguards male identity and prevents inappropriate activation of the female program in adult somatic cells.

Summary: Aging is associated with progressive tissue decline and shifts in stem cell clonality. The role of niche signals in driving these processes remains poorly understood. Using the Drosophila testis, we identify a regulatory axis in which age-related decline of niche signals (BMPs) lead to upregulation of the co-repressor Hairless, which downregulates the RNA-binding protein Imp in aged germline stem cells (GSCs). Reduced Imp causes loss of Chinmo, a key factor in GSC aging and competition. Reduced Chinmo causes ectopic Perlecan secretion which accumulates in the testis lumen and causes GSC loss. Aging of the testis is reversed by increasing BMPs in the niche, or by overexpressing Imp or depleting Hairless in GSCs. Furthermore, GSC clones with reduced Imp or increased Hairless are more competitive, expelling wild-type neighbors and monopolizing the niche. Thus, BMPs regulate testicular niche aging through the Hairless–Imp–Chinmo axis and “winning” GSCs usurp these aging mechanisms. Highlights: Aged niche cells produce less BMPs, resulting in more Hairless (H) in aged GSCs Elevated H represses Imp , resulting in less Chinmo and in ectopic ECM secretion Aging is prevented by higher BMP in niche cells, or by higher Imp or lower H in GSCs GSCs with low Imp or high H exploit these aging mechanisms to colonize the GSC pool

Summary: Somatic sex identity must be maintained throughout adulthood for tissue function. Adult somatic stem cells in the Drosophila testis (i.e., CySCs) lacking the transcription factor Chinmo are reprogrammed to their ovarian counterparts by induction of female-specific Tra ^F , but this is not mechanistically understood. Pioneer factors play central roles in direct reprogramming, and many upregulated genes in chinmo ^−/- CySCs contain binding sites for the pioneer factor Zelda (Zld). microRNAs repress zld mRNA in wild type CySCs, but they are downregulated after Chinmo loss, allowing for zld mRNA translation. Zld depletion from chinmo ^−/- CySCs suppresses feminization, and ectopic Zld induces Tra ^F and feminizes wild-type CySCs. qkr58E-2 and ecdysone receptor ( EcR ), direct Zld targets in the embryo, are female-biased in adult gonads and upregulated in chinmo ^−/- CySCs. The RNA-binding protein Qkr58E-2 produces Tra ^F , while EcR promotes female-biased gene expression. Ectopic Zld feminizes adult male adipose tissue, demonstrating that Zld can instruct female and override male identity in adult XY tissues. Highlights: zld mRNA is repressed by microRNAs in XY somatic gonadal cells Zld is upregulated in and required for sex reversal of XY chinmo ^−/- cells Zld induces Qkr58E-2 and EcR, which cause Tra ^F and female-biased transcription Zld feminizes XY adipose cells by inducing Tra ^F and downregulating Chinmo

In this review, we introduce the concept of cell competition, which occurs between heterogeneous neighboring cell populations. Cells with higher relative fitness become “winners” that outcompete cells of lower relative fitness (“losers”). We discuss the idea of super-competitors, mutant cells that expand at the expense of wild-type cells. Work on adult stem cells (ASCs) has revealed principles of neutral competition, wherein ASCs can be stochastically lost and replaced, and of biased competition, in which a winning ASC with a competitive advantage replaces its neighbors. Germline stem cells (GSCs) are ASCs that are uniquely endowed with the ability to produce gametes and, therefore, impact the next generation. Mechanisms of GSC competition have been elucidated by studies in Drosophila gonads, tunicates, and the mammalian testis. Competition between ASCs is thought to underlie various forms of cancer, including spermatocytic tumors in the human testis. Paternal age effect (PAE) disorders are caused by de novo mutations in human GSCs that increase their competitive ability and make them more likely to be inherited, leading to skeletal and craniofacial abnormalities in offspring. Given its widespread effects on human health, it is important to study GSC competition to elucidate how cells can become winners or losers.

Stem cell pools are dynamic and capable of reacting to insults like injury and starvation. Recent work has highlighted the key role of dedifferentiation as a conserved mechanism for replenishing stem cell pools after their loss, thereby maintaining tissue homeostasis. The testis of the fruit fly Drosophila melanogaster offers a simple but powerful system to study dedifferentiation, the process by which differentiating spermatogonia can revert their fate to become fully functional germline stem cells (GSCs). Dedifferentiated GSCs show interesting characteristics, such as being more proliferative than their wild-type sibling GSCs. To facilitate the study of the cellular and molecular mechanisms underlying the process of germline dedifferentiation in the Drosophila testis, here we describe techniques for inducing high rates of dedifferentiation and for unambiguously labeling dedifferentiated GSCs. Key words Germline Dedifferentiation Stem cells Drosophila melanogaster Testis