Orphan nuclear receptor TLX activates Wnt/β-catenin signalling to stimulate neural stem cell proliferation and self-renewal

Abstract

The nuclear receptor TLX (also known as NR2E1) is essential for adult neural stem cell self-renewal; however, the molecular mechanisms involved remain elusive. Here we show that TLX activates the canonical Wnt/β-catenin pathway in adult mouse neural stem cells. Furthermore, we demonstrate that Wnt/β-catenin signalling is important in the proliferation and self-renewal of adult neural stem cells in the presence of epidermal growth factor and fibroblast growth factor. Wnt7a and active β-catenin promote neural stem cell self-renewal, whereas the deletion of Wnt7a or the lentiviral transduction of axin, a β-catenin inhibitor, led to decreased cell proliferation in adult neurogenic areas. Lentiviral transduction of active β-catenin led to increased numbers of type B neural stem cells in the subventricular zone of adult brains, whereas deletion of Wnt7a or TLX resulted in decreased numbers of neural stem cells retaining bromodeoxyuridine label in the adult brain. Both Wnt7a and active β-catenin significantly rescued a TLX (also known as Nr2e1) short interfering RNA-induced deficiency in neural stem cell proliferation. Lentiviral transduction of an active β-catenin increased cell proliferation in neurogenic areas of TLX-null adult brains markedly. These results strongly support the hypothesis that TLX acts through the Wnt/β-catenin pathway to regulate neural stem cell proliferation and self-renewal. Moreover, this study suggests that neural stem cells can promote their own self-renewal by secreting signalling molecules that act in an autocrine/paracrine mode. 8 Correspondence should be addressed to Y.S. (yshi@coh.org). 6,7 These authors contributed equally to this work.Note: Supplementary Information is available on the Nature Cell Biology website. AUTHOR CONTRIBUTIONSY.S. conceived and designed the study. Y.S., Q.Q., G.S., W.L., S.Y., P.Y. and C.Z. performed the experiments and analysed the data. Y.S., Q.Q., G.S., F.H.G. and R.M.E. interpreted the data. Y.S. wrote the paper with comments from Q.Q., G.S., R.T.Y., F.H.G. and R.M.E. COMPETING FINANCIAL INTERESTSThe authors declare no competing financial interests. The finding of neurogenesis in the adult brain led to the discovery of adult neural stem cells. Neural stem cells are defined as a subset of undifferentiated precursors that retain the ability to proliferate and self-renew and have the capacity to give rise to both neuronal and glial lineages [1][2][3][4] . Under normal conditions, neurogenesis in the adult mammalian brain is restricted to two discrete germinal centres: the subgranular layer of the hippocampal dentate gyrus 3 and the subventricular zones of the lateral ventricles 5,6 . A complete understanding of adult neural stem cells requires the identification of molecules that determine the self-renewal and multipotent characteristics of these cells.TLX is an orphan nuclear receptor that is expressed in vertebrate forebrains 7,8 . We showed previously that TLX is an important regulator of neural stem cell maintenance and self-renewal in both embryo...

The Orphan Nuclear Receptor TLX/NR2E1 in Neural Stem Cells and Diseases

“…This study confirmed that the TLX LBD structure resembles other NR LBD conformations as a canonical helical sandwich, and also revealed notable differences, in that the TLX LBD lacks the first helices a1 and a2 and contains a unique 5-amino-acid insertion between helices a8 and a9. In contrast to the previous description of the full-length TLXforming monomer [21], Benod et al suggested that the recombinant TLX LBD can form a homodimer in solution [18]. More interestingly, three compounds were shown to be able to bind the recombinant TLX LBD, indicating that TLX functions can be manipulated using selected compounds [18].…”

“…Prospero, which encodes a homeodomain transcription factor associated with the differentiation of ganglion mother cells after asymmetric stem cell division of neuroblasts in the central brain [42,43], is the only target gene that has been found to be directly regulated by Tll in postembryonic stages [44]. In the mouse, multiple genes involved in the development of the CNS and visual system have been identified as Tlx targets, such as Gsh2, Pax2, Pten, p21, p57, S100b, Aqp4, Plce1, Wnt7a, microRNA-9, Bmp4, and GFAP [21,[32][33][34][35][36][37][45][46][47][48][49][50] (Table 1). Interestingly, Tlx may act as a transcriptional activator for the expression of sirt1, Wnt7a, Mash1, and Oct-3/4 based on in vitro luciferase assays and chromatin immunoprecipitation assays [16,21,[51][52][53].…”

“…In the mouse, multiple genes involved in the development of the CNS and visual system have been identified as Tlx targets, such as Gsh2, Pax2, Pten, p21, p57, S100b, Aqp4, Plce1, Wnt7a, microRNA-9, Bmp4, and GFAP [21,[32][33][34][35][36][37][45][46][47][48][49][50] (Table 1). Interestingly, Tlx may act as a transcriptional activator for the expression of sirt1, Wnt7a, Mash1, and Oct-3/4 based on in vitro luciferase assays and chromatin immunoprecipitation assays [16,21,[51][52][53]. A consensus Nr2e1 binding sequence within the promoter region of its target gene has been proposed to contain AAGTCA, which has been identified in the promoters of Wnt7a, GFAP, S100b, Oct3/ 4, p21, Pax2, Plce1, and Pten [21,32,34,45,46,53,54].…”

“…Interestingly, Tlx may act as a transcriptional activator for the expression of sirt1, Wnt7a, Mash1, and Oct-3/4 based on in vitro luciferase assays and chromatin immunoprecipitation assays [16,21,[51][52][53]. A consensus Nr2e1 binding sequence within the promoter region of its target gene has been proposed to contain AAGTCA, which has been identified in the promoters of Wnt7a, GFAP, S100b, Oct3/ 4, p21, Pax2, Plce1, and Pten [21,32,34,45,46,53,54]. Given the functional equivalence and the highly-conserved DBDs between human TLX and mouse Tlx [13,14], TLX may have the capacity to bind a similar consensus promoter sequence and suppress or activate the expression of multiple human counterparts of the identified mouse Tlx targets.…”

“…Tlx has been reported to interact with BMP7 and/or Sonic Hedgehog (SHH) and relieve the repression of Pax2 expression in mouse retinal astrocytes [50]. Mouse Tlx also regulates the Wnt-Frizzled signaling pathway by directly activating Wnt7a expression [21]. Niu et al showed that mouse Tlx is involved in regulating DNA replication and cell-cycle signaling in postnatal NSCs [49].…”