Low-level repressive histone marks fine-tune gene transcription in neural stem cells.

Coordinated regulation of gene activity by transcriptional and translational mechanisms poise stem cells for a timely cell-state transition during differentiation. Although important for all stemness-to-differentiation transitions, mechanistic understanding of the fine-tuning of gene transcription is lacking due to the compensatory effect of translational control. We used intermediate neural progenitor (INP) identity commitment to define the mechanisms that fine-tune stemness gene transcription in fly neural stem cells (neuroblasts). We demonstrate that the transcription factor FruitlessC (FruC) binds cis-regulatory elements of most genes uniquely transcribed in neuroblasts. Loss of fruC function alone has no effect on INP commitment but drives INP dedifferentiation when translational control is reduced. FruC negatively regulates gene expression by promoting low-level enrichment of the repressive histone mark H3K27me3 in gene cis-regulatory regions. Identical to fruC loss-of-function, reducing Polycomb Repressive Complex 2 activity increases stemness gene activity. We propose low-level H3K27me3 enrichment fine-tunes gene transcription in stem cells, a mechanism likely conserved from flies to humans.

From neurons to sperm, our bodies are formed of a range of cells tailored to perform a unique role. However, organisms also host small reservoirs of unspecialized 'stem cells' that retain the ability to become different kinds of cells. When these stem cells divide, one daughter cell remains a stem cell while the other undergoes a series of changes that allows it to mature into a specific cell type. This 'differentiation' process involves quickly switching off the stem cell programme, the set of genes that give a cell the ability to keep dividing while maintaining an unspecialized state. Failure to do so can result in the differentiating cell reverting towards its initial state and multiplying uncontrollably, which can lead to tumours and other health problems. While scientists have a good understanding of how the stem cell programme is turned off during differentiation, controlling these genes is a balancing act that starts even before division: if the program is over-active in the 'mother' stem cell, for instance, the systems that switch it off in its daughter can become overwhelmed. The mechanisms presiding over these steps are less well-understood. To address this knowledge gap, Rajan, Anhezini et al. set out to determine how stem cells present in the brains of fruit flies could control the level of activity of their own stem cell programme. RNA sequencing and other genetic analyses revealed that a protein unique to these cells, called Fruitless, was responsible for decreasing the activity of the programme. Biochemical experiments then showed that Fruitless performed this role by attaching a small amount of chemical modifications (called methyl groups) to the proteins that 'package' the DNA near genes involved in the stem cell programme. High levels of methyl groups present near a gene will switch off this sequence completely; however, the amount of methyl groups that Fruitless helped to deposit is multiple folds lower. Consequently, Fruitless 'fine-tunes' the activity of the stem cell programme instead, dampening it just enough to stop it from overpowering the 'off' mechanism that would take place later in the daughter cell. These results shed new light on how stem cells behave ­ and how our bodies stop them from proliferating uncontrollably. In the future, Rajan, Anhezini et al. hope that this work will help to understand and treat diseases caused by defective stem cell differentiation.

Sequential activation of transcriptional repressors promotes progenitor commitment by silencing stem cell identity genes.

Stem cells that indirectly generate differentiated cells through intermediate progenitors drives vertebrate brain evolution. Due to a lack of lineage information, how stem cell functionality, including the competency to generate intermediate progenitors, becomes extinguished during progenitor commitment remains unclear. Type II neuroblasts in fly larval brains divide asymmetrically to generate a neuroblast and a progeny that commits to an intermediate progenitor (INP) identity. We identified Tailless (Tll) as a master regulator of type II neuroblast functional identity, including the competency to generate INPs. Successive expression of transcriptional repressors functions through Hdac3 to silence tll during INP commitment. Reducing repressor activity allows re-activation of Notch in INPs to ectopically induce tll expression driving supernumerary neuroblast formation. Knocking-down hdac3 function prevents downregulation of tll during INP commitment. We propose that continual inactivation of stem cell identity genes allows intermediate progenitors to stably commit to generating diverse differentiated cells during indirect neurogenesis.

Synergism between canoe and scribble mutations causes tumor-like overgrowth via Ras activation in neural stem cells and epithelia.

Over the past decade an intriguing connection between asymmetric cell division, stem cells and tumorigenesis has emerged. Neuroblasts, which are the neural stem cells of the Drosophila central nervous system, divide asymmetrically and constitute an excellent paradigm for investigating this connection further. Here we show that the simultaneous loss of the asymmetric cell division regulators Canoe (afadin in mammals) and Scribble in neuroblast clones leads to tumor-like overgrowth through both a severe disruption of the asymmetric cell division process and canoe loss-mediated Ras-PI3K-Akt activation. Moreover, canoe loss also interacts synergistically with scribble loss to promote overgrowth in epithelial tissues, here just by activating the Ras-Raf-MAPK pathway. discs large 1 and lethal (2) giant larvae, which are functionally related to scribble, contribute to repress the Ras-MAPK signaling cascade in epithelia. Hence, our work uncovers novel cooperative interactions between all these well-conserved tumor suppressors that ensure tight regulation of the Ras signaling pathway.

The hippo pathway core cassette regulates asymmetric cell division.

Asymmetric cell division (ACD) is a crucial process during development, homeostasis, and cancer. Stem and progenitor cells divide asymmetrically, giving rise to two daughter cells, one of which retains the parent cell self-renewal capacity, while the other is committed to differentiation. Any imbalance in this process can induce overgrowth or even a cancer-like state. Here, we show that core components of the Hippo signaling pathway, an evolutionarily conserved organ growth regulator, modulate ACD in Drosophila. Hippo pathway inactivation disrupts the asymmetric localization of ACD regulators, leading to aberrant mitotic spindle orientation and defects in the generation of unequal-sized daughter cells. The Hippo pathway downstream kinase Warts, LATS1-2 in mammals, associates with the ACD modulators Inscuteable and Bazooka in vivo and phosphorylates Canoe, the ortholog of Afadin/AF-6, in vitro. Moreover, phosphosite mutant Canoe protein fails to form apical crescents in dividing neuroblasts in vivo, and the lack of Canoe phosphorylation by Warts leads to failures of Discs Large apical localization in metaphase neuroblasts. Given the relevance of ACD in stem cells during tissue homeostasis, and the well-documented role of the Hippo pathway as a tumor suppressor, these results represent a potential route for perturbations in the Hippo signaling to induce tumorigenesis via aberrant stem cell divisions.

A Serrate-Notch-Canoe complex mediates essential interactions between glia and neuroepithelial cells during Drosophila optic lobe development.

It is firmly established that interactions between neurons and glia are fundamental across species for the correct establishment of a functional brain. Here, we found that the glia of the Drosophila larval brain display an essential non-autonomous role during the development of the optic lobe. The optic lobe develops from neuroepithelial cells that proliferate by dividing symmetrically until they switch to asymmetric/differentiative divisions that generate neuroblasts. The proneural gene lethal of scute (l'sc) is transiently activated by the epidermal growth factor receptor (EGFR)-Ras signal transduction pathway at the leading edge of a proneural wave that sweeps from medial to lateral neuroepithelium, promoting this switch. This process is tightly regulated by the tissue-autonomous function within the neuroepithelium of multiple signaling pathways, including EGFR-Ras and Notch. This study shows that the Notch ligand Serrate (Ser) is expressed in the glia and it forms a complex in vivo with Notch and Canoe, which colocalize at the adherens junctions of neuroepithelial cells. This complex is crucial for interactions between glia and neuroepithelial cells during optic lobe development. Ser is tissue-autonomously required in the glia where it activates Notch to regulate its proliferation, and non-autonomously in the neuroepithelium where Ser induces Notch signaling to avoid the premature activation of the EGFR-Ras pathway and hence of L'sc. Interestingly, different Notch activity reporters showed very different expression patterns in the glia and in the neuroepithelium, suggesting the existence of tissue-specific factors that promote the expression of particular Notch target genes or/and a reporter response dependent on different thresholds of Notch signaling.