Distinct Roles for Wnt-4 and Wnt-11 During Retinoic Acid-Induced Neuronal Differentiation

Abstract

Retinoic acid and Wnt/β‐catenin signals play important roles during neuronal differentiation but less is known about noncanonical Wnt signals in this context. We examined retinoic acid and Wnt signaling in two human embryonal carcinoma cell lines, NTERA‐2 (clone D1), which undergoes neuronal differentiation in response to retinoic acid, and 2102Ep, which does not. Retinoic acid treatment inhibited β‐catenin/Tcf activity in NTERA‐2 cells but not in 2102Ep cells. Inhibition occurred downstream of β‐catenin but did not involve competition between retinoic acid receptors and β‐catenin for binding to p300 or Tcf‐4. Ectopic expression of FZD1 partially restored inhibition in 2102Ep cells, suggesting the involvement of Wnt ligands. Retinoic acid treatment of NTERA‐2 cells induced the expression of Wnt‐4 and Wnt‐11, both of which were able to inhibit β‐catenin/Tcf activity. Wnt‐4 and Wnt‐11 were found at cell borders in islands of cells that expressed OCT4 and GFAP and were predominantly negative for Nestin, PAX6, and GATA6. Gene silencing of Wnt‐4, but not Wnt‐11, reduced retinoic acid downregulation of OCT4 and Nanog and upregulation of PAX6, ASCL1, HOXC5, and NEUROD1, suggesting that Wnt‐4 promotes early neuronal differentiation. Gene expression analysis of NTERA‐2 cells stably overexpressing Wnt‐11 suggested that Wnt‐11 potentiates retinoic acid induction of early neurogenesis. Consistent with this, overexpression of Wnt‐11 maintained a population of proliferating progenitor cells in cultures treated with retinoic acid for several weeks. These observations highlight the distinct roles of two noncanonical Wnts during the early stages of retinoic acid‐induced neuronal differentiation. STEM CELLS 2011;29:141–153

VEGF-B selectively regenerates injured peripheral neurons and restores sensory and trophic functions

“…It has been demonstrated that activation of Notch signaling results in increased microtubule stability and changes in axonal morphology and branching (21). Similarly, Wnt2 promotes cortical dendrite growth and dendritic spine formation and Wnt11 early neurogenesis (22,23). Because VEGF-induced signaling is complex and intersects with multiple pathways, we tested whether inhibition of PI3K or Notch signaling resulted in functional changes of VEGF-induced neurite growth.…”

Canonical and noncanonical Wnt signaling in neural stem/progenitor cells

“…Previous studies highlight the critical role Wnt signaling plays in NSCs, but the nature of the Wnt signals involved remains unclear, with reports of increased and decreased Wnt signaling taking place during differentiation [68,175] and disease [176,177]. Neurogenesis in the hippocampus, where Wnt signaling plays important roles, is gradually lost as we age [178], and this loss is implicated in neurodegenerative diseases [123].…”

“…Induced proliferation of hESCs/mNSCs [65,66] Recombinant Wnt-3a Induced proliferation and differentiation of hESCs [67] Wnt-4 silencing Impaired early differentiation in hECCs [68] Wnt-5a KO Impaired neurite development in the olfactory bulb (OB) [44] Wnt-1 and Wnt-5a DKO Impaired neurogenesis of midbrain dopaminergic neurons [52] Wnt-5a KO Impaired axon growth and guidance of dopaminergic neurons [45] Wnt-5a CM Increased synaptogenesis and maturation of hippocampal progenitors [69,70] Wnt-5a overexpression Induced axonal differentiation in hippocampal cultures [71] Wnt-7a KO Delayed morphological maturation of glomerular rosettes and synapsin I accumulation [46] Wnt-7a KO Impaired ventral midbrain neurogenesis [47] Wnt-7a and Dvl DKO Defective spine morphogenesis and mossy fiber-CA3 synaptic transmission [48] Wnt-7a Proposed as a key element in the regulation of NSC self-renewal/differentiation; altered spindle- Canonical Wnt receptors are also important for correct neural development (Table 2): FZD3 KO mice show impaired axonal guidance [73] while LRP6 KO mice present cortical defects [74]. Also, FZD1 has been shown to be the receptor for canonical Wnt-1 in mouse tyrosine hydroxylase positive neurons, which activates β-catenin-dependent signaling promoting neuroprotection in dopaminergic neurons [75].…”

“…For example, it induces hES cell exit from the pluripotent state, mesodermal/hematopoietic cell fate [161] and cardiac differentiation of mES cells [162]. In human embryonal carcinoma cells, Wnt-11 maintains neural progenitor cell proliferation but prevents further differentiation, which instead is driven by another noncanonical Wnt, Wnt-4 [68]. We recently found that noncanonical Wnt -1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 3a signaling stimulates differentiation of hES cells and iPS cells, something that could account for some of the controversy found in the literature regarding the role of Wnt-3a [163].…”

Wnt4 antagonises Wnt3a mediated increases in growth and glucose stimulated insulin secretion in the pancreatic beta-cell line, INS-1

“…In the nucleus β-catenin can act as a transcription factor (along with its cofactors TCF/LEF) to promote gene transcription of effectors of Wnt signalling, such as c-myc and cyclin D2. In contrast, the non-canonical pathways, such as those stimulated by Wnt4 are β-catenin independent and have been shown to antagonise the canonical Wnt pathway in several cell types [13,14,[19][20][21].…”