Discovery of a functional protein complex of netrin-4, laminin γ1 chain, and integrin α6β1 in mouse neural stem cells

Abstract

Molecular and cellular interactions coordinating the origin and fate of neural stem cells (NSCs) in the adult brain are far from being understood. We present a protein complex that controls proliferation and migration of adult NSCs destined for the mouse olfactory bulb (OB). Combinatorial selection based on phage display technology revealed a previously unrecognized complex between the soluble protein netrin-4 and laminin ␥1 subunit that in turn activates an ␣6␤1 integrin-mediated signaling pathway in NSCs. Differentiation of NSCs is accompanied by a decrease in netrin-4 receptors, indicating that netrin-4 participates in the continual propagation of this stem cell population. Notably, the stem cells themselves do not synthesize netrin-4. Further, we show that netrin-4 is produced by selected GFAP-positive astrocytes positioned close to newborn neurons migrating in the anterior part of the rostral migratory stream (RMS) and within the OB. Our findings present a unique molecular mechanism mediating astrocytic/neuronal crosstalk that regulates ongoing neurogenesis in the adult olfactory system. olfactory system ͉ phage display ͉ merogenesis ͉ rostral migratory stream ͉ astrocytes N etrins are a family of secreted proteins initially described as developmental axon attractants (1). Yet, with critical roles in tissue morphogenesis (2-4), vascular patterning (5), and angiogenesis (6-8), netrins have become recognized to mediate functions far beyond axonal guidance. Netrin-4 is expressed in both neural and nonneural tissues (8-11). In the nervous system, it promotes neurite extension from olfactory bulb (OB) explants but the molecular mechanisms underlying this phenomenon remain unknown. In the vasculature of the kidney, heart, and ovary, the protein is located in the basement membrane. Integrins and netrins, along with the established netrin-1 and netrin-4 receptors DCC and UNC5, appear to cooperate to regulate multiple aspects of development (3,12,13). However, certain netrin-4 functions are mediated by neogenin (8) and likely by other, as yet unidentified receptors (14, 15).Here, we provide evidence for a direct relationship between netrin-4, laminin ␥1 chain, and ␣6␤1 integrin in the context of neural stem cells (NSCs). By exploiting the power of the unbiased phage display combinatorial approach (16-18), followed by peptide affinity chromatography and functional validation, we found that a netrin-4/laminin ␥1 chain complex binds to ␣6␤1 integrin, with subsequent activation of the mitogenactivated protein (MAP) kinase signaling pathway, resulting in NSC migration and proliferation. We also show that in the adult mouse brain, netrin-4 is produced by a subset of astrocytes on the edge of the rostral migratory system (RMS) close to the site of neuron entrance into the OB. Our findings support the conclusion that netrin-4 is a regulatory factor in NSC biology and uncover a previously unrecognized mechanism in which a multimeric complex functions as an integrated supramolecular switch that regulates NSC fate. Results and...

Kruppel-like Factor-9 (KLF9) Inhibits Glioblastoma Stemness through Global Transcription Repression and Integrin α6 Inhibition

“…Integrins, a family of cell adhesion receptors, are up-regulated in stem cells and are believed to support cell stemness by mediating adhesive interactions between stem cells and their niches (60). Integrin ␣6, a subunit for two laminin receptors, is also highly expressed in several types of stem cells, including subventricular neural stem cells (61,62), and facilitates neural stem cell adherence to endothelial cells in the perivascular niche (62). Integrin ␣6 has also been identified as a marker for cancer stem cells, including GSCs (48,53).…”

Laminin/β1 integrin signal triggers axon formation by promoting microtubule assembly and stabilization

“…Some of these factors interact with integrin signaling during cell proliferation [64], adhesion and migration [65][66][67], neurite outgrowth, and growth cone guidance [68][69][70]. It is thus possible that other extracellular cues execute their roles through modulating integrin signaling directly or indirectly.…”

Adhesion in the stem cell niche: biological roles and regulation

“…The lamininintegrin-mediated interaction also contributes to the development of the NSC niche and the maintenance of its integrity (Loulier et al, 2009). In addition, netrin 4 secreted by glial fibrillary acidic protein (GFAP)-positive astrocytes and laminin secreted by NSCs themselves can activate the 61 integrin-mediated signaling pathway to control NSC proliferation (Staquicini et al, 2009). Integrins can also promote NSC self-renewal by facilitating Notch and EGFR signaling (Campos et al, 2006), and integrins have recently been shown to regulate thrombopoietin-mediated HSC maintenance (Umemoto et al, 2012).…”