Abstract
IgLON proteins are GPI anchored adhesion molecules that control neurite outgrowth. In particular, Negr1 down-regulation negatively influences neuronal arborization in vitro and in vivo. In the present study, we found that the metalloprotease ADAM10 releases Negr1 from neuronal membrane. Ectodomain shedding influences several neuronal mechanisms, including survival, synaptogenesis, and the formation of neurite trees. By combining morphological analysis and virus-mediated selective protein silencing in primary murine cortical neurons, we found that pharmacologically inhibition of ADAM10 results in an impairment of neurite tree maturation that can be rescued upon treatment with soluble Negr1. Furthermore, we report that released Negr1 influences neurite outgrowth in a P-ERK1/2 and FGFR2 dependent manner. Together our findings suggest a role for Negr1 in regulating neurite outgrowth through the modulation of FGFR2 signaling pathway. Given the physiological and pathological role of ADAM10, Negr1, and FGFR2, the regulation of Negr1 shedding may play a crucial role in sustaining brain function and development.
Highlights
Neurite outgrowth is a fundamental process that allows the establishment of functional wiring in the developing brain
To investigate these two scenarios, we infected cortical neurons at DIV4 with viruses expressing GFP marker together with either Negr1 specific siRNA or scramble control following two experimental paradigms: (1) cultures infected with a high viral titer (MOI = 3); (2) cultures infected with a low viral titer (MOI = 0.3)
To confirm the biological relevance of Negr1 as soluble factor, we investigated in detail the presence of Negr1, NCAM, and S6 ribosomial protein, a well-established cytoplasmic marker, in samples prepared from neuronal culture at DIV18 or from the relative conditioned media
Summary
Neurite outgrowth is a fundamental process that allows the establishment of functional wiring in the developing brain. This process can be summarized as a mechanism where extracellular cues bind to transmembrane receptors and trigger signaling cascades which eventually reorganize neuronal structure (Raper and Mason, 2010). Accumulating evidence indicates that IgLON are tightly implicated in neuronal functions such as synaptic formation, plasticity, and neurite tree development (Gil et al, 1998; Schäfer et al, 2005; Hashimoto et al, 2008, 2009; Pischedda et al, 2014; Sanz et al, 2015). IgLON proteins may form homophilic and heterophilic complexes at the cell surface or with juxtaposed cells to modulate adhesion and neurite outgrowth
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