Querkopf is a key marker of self-renewal and multipotency of adult neural stem cells Bilal N. Sheikh 1,2 , Mathew P. Dixon 1 , Tim Thomas 1,2, * ,` and Anne K. Voss 1,2, * ,` 1 The Walter and Eliza Hall Institute of Medical Research, Parkville 3050, Victoria, Australia 2 Department of Medical Biology, University of Melbourne, Parkville, Victoria, Australia *These authors co-supervised this project and contributed equally to this work ` Authors for correspondence (avoss@wehi.edu.au; tthomas@wehi.edu.au) Accepted 22 July 2011 Journal of Cell Science 125, 1–15 ß 2011. Published by The Company of Biologists Ltd doi: 10.1242/jcs.077271 Summary Adult neural stem cells (NSCs) reside in the subventricular zone (SVZ) and produce neurons throughout life. Although their regenerative potential has kindled much interest, few factors regulating NSCs in vivo are known. Among these is the histone acetyltransferase querkopf (QKF, also known as MYST4, MORF, KAT6B), which is strongly expressed in a small subset of cells in the neurogenic subventricular zone. However, the relationship between Qkf gene expression and the hierarchical levels within the neurogenic lineage is currently unknown. We show here that the 10% of SVZ cells with the highest Qkf expression possess the defining NSC characteristics of multipotency and self-renewal and express markers previously shown to enrich for NSCs. A fraction of cells expressing Qkf at medium to high levels is enriched for multipotent progenitor cells with limited self-renewal, followed by a population containing migrating neuroblasts. Cells low in Qkf promoter activity are predominantly ependymal cells. In addition, we show that mice deficient for Bmi1,a central regulator of NSC self-renewal, show an age-dependent decrease in the strongest Qkf-expressing cell population in the SVZ. Our results show a strong relationship between Qkf promoter activity and stem cell characteristics, and a progressive decrease in Qkf gene activity as lineage commitment and differentiation proceed in vivo. Key words: MYST, Epigenetics, Neurogenesis, Neural stem cells, HAT, Histone acetyltransferase, Fluorescence-activated cell sorting, Polycomb Introduction Adult neural stem cells (NSCs) are present in the subventricular zone (SVZ) and the subgranular layer of the hippocampus and are a source of new neurons throughout life (Zhao et al., 2008). SVZ NSCs give rise to transit amplifying cells, which in turn produce neuroblasts that migrate to the olfactory bulbs and differentiate into olfactory interneurons (Lois et al., 1996). SVZ NSCs have been induced to differentiate into an array of different neural cell types, both in vivo and in vitro (Curtis et al., 2003; Lois and Alvarez-Buylla, 1993). NSCs have therefore aroused great interest, as they hold much promise for the generation of new therapies for neurodegenerative disorders and neural injury. However, two major issues have hindered progress in the field. First, it has been difficult to isolate a pure population of NSCs in their most primitive state. Although some markers are available for the enrichment of NSCs, a definitive marker for NSCs remains to be defined. Second, in order to direct differentiation of NSCs into specific neuronal cell types, which would ideally include cell types not normally formed by NSCs, a greater understanding of factors underlying lineage commitment and hierarchy is required. NSCs have been shown to express a number of genes including intracellular proteins such as glial fibrillary acidic protein (GFAP), nestin and inhibitor of DNA binding 1 (Id1) (Doetsch et al., 1999; Mignone et al., 2004; Nam and Benezra, 2009), cell surface proteins such as SSEA-1 (also known as FUT4 and LeX) and CD133 (Capela and Temple, 2002; Coskun et al., 2008; Beckervordersandforth et al., 2010), and low levels of heat stable antigen (HSA; HSA is also known as CD24). NSCs also show little affinity for the lectin peanut agglutinin (PNA) (Rietze et al., 2001). Although some of these characteristics have been used to isolate cell fractions enriched for NSCs, most of the markers are expressed strongly in other cells in the brain, whereas others only mark a subset of NSCs. For example, GFAP is expressed by both NSCs and differentiated astrocytes. Of note, none of the markers currently in use to enrich for or to identify NSCs play a non- redundant role in adult NSC biology. We show here that high levels of querkopf (Qkf; also known as Kat6b and Myst4) expression identifies multipotent, self-renewing NSCs. We have previously shown that a lack of the MYST-family histone acetyltransferase, QKF, leads to defects in the establishment and self-renewal of adult NSCs and consequently to a progressive defect in adult neurogenesis (Merson et al., 2006; Rietze et al., 2001). Remarkably, in humans loss of only one allele of MYST4 (QKF) leads to intellectual disability (Kraft et al., 2011; Clayton-Smith et al., 2011). Unlike other histone acetyltransferases, the expression of Qkf is both spatially and temporally regulated, and strong Qkf expression is localized to neurogenic regions both during development and in the adult (Merson et al., 2006; Thomas et al., 2000). We observed strong expression of Qkf gene in a small subset of neurogenic SVZ cells, raising the question of whether there is a relationship between Qkf gene expression levels and cell identity. Using Qkf–GFP transgenic mice to trace levels of Qkf gene activity, we show that the strongest Qkf–GFP-expressing cells possess all NSC characteristics, namely multipotency and self-renewal. Research Article 1 Journal of Cell Science JCS ePress online publication date 13 February 2012