Growth cone macropinocytosis of neurotrophin receptor and neuritogenesis are regulated by neuron navigator 1

Abstract

Neuron Navigator 1 (Nav1) is a cytoskeleton-associated protein expressed during brain development that is necessary for proper neuritogenesis, but the underlying mechanisms are poorly understood. Here we show that Nav1 is present in elongating axon tracts during mouse brain embryogenesis. We found that depletion of Nav1 in cultured neurons disrupts growth cone morphology and neurotrophin-stimulated neuritogenesis. In addition to regulating both F-actin and microtubule properties, Nav1 promotes actin-rich membrane ruffles in the growth cone, and promotes macropinocytosis at those membrane ruffles, including internalization of the TrkB receptor for the neurotrophin BDNF (brain-derived neurotropic factor). Growth cone macropinocytosis is important for downstream signaling, neurite targeting, and membrane recycling, implicating Nav1 in one or more of these processes. Depletion of Nav1 also induces transient membrane blebbing via disruption of signaling in the Rho GTPase signaling pathway, supporting the novel role of Nav1 in dynamic actin-based membrane regulation at the cell periphery. These data demonstrate that Nav1 works at the interface of microtubules, actin, and plasma membrane to organize the cell periphery and promote uptake of growth and guidance cues to facilitate neural morphogenesis during development.

The Neuron Navigators: Structure, function, and evolutionary history

“…In contrast, Nav1 mRNA levels were moderately reduced in Tbr1 null neocortex, suggesting that Tbr1 activates Nav1 expression for cortical neuron development ( Bedogni et al, 2010 ). Consistent with this interpretation, Nav1 protein is highly expressed in the axons of cortical neurons in vivo ( Powers et al, 2022 ). Although Nav3 is also expressed in developing cerebral cortex ( Coy et al, 2002 ), this gene does not appear to be significantly regulated by Pax6, Tbr2, or Tbr1.…”

“…Interestingly, one region of the developing brain where specific transcription factors have been found to regulate Navigator gene expression is the embryonic mouse cerebral neocortex. Within this region, all three Navigators are expressed in the intermediate zone and cortical plate ( Coy et al, 2002 ; Peeters et al, 2004 ; Martinez-Lopez et al, 2005 ; Pook et al, 2020 ; Powers et al, 2022 ), where newly generated cortical neurons migrate, differentiate, and grow axons and dendrites. Transcription factors Pax6, Tbr1, and Tbr2 (also known as Eomes) are important regulators of cortical neuron differentiation that also directly regulate Navigator gene expression ( Englund et al, 2005 ; Mihalas and Hevner, 2017 ; Elsen et al, 2018 ).…”

“…(K–N ) Nav1 protein expression (red) with DAPI (blue) to label nuclei and SMI-131 (green) to label axons. Labeling demonstrates Nav1 expression in neurons and axons in the upper layers of developing cortex (Reproduced with permission from Powers et al, 2022 ). sc, spinal cord; cx, cortex; dt, dorsal thalamus; mid, midbrain; cb, cerebellum; cp, cortical plate; iz, intermediate zone; svz, subventricular zone; vz, ventricular zone; Cx, cortex; LV, lateral ventricle; rls, rostral rhombic lip migration stream; Th, thalamus.…”

“…The only Navigator protein studied by immunohistochemistry in vivo so far is NAV1, which is expressed in growing axon tracts, soma, and dendrites within developing neocortex ( Powers et al, 2022 ; Figure 6 ). These observations are consistent with the conclusion that Navigators are important for axon guidance and neurite development, as indicated by studies of mice and humans with Navigator mutations.…”

“…The subcellular localization of Navigators has been best characterized using vertebrate neurons in primary cultures and in neuroblastoma and other cancer-derived cell lines. All three mammalian Navigators partially localize to microtubule plus ends ( van Haren et al, 2009 , 2014 ; Sanchez-Huertas et al, 2020 ; Powers et al, 2022 ), and Nav1 and Nav2 were shown to use SxIP motifs to mediate this localization. Because Nav3 also contains SxIP motifs ( Figure 1 ), it is assumed Nav3 plus tip localization is also mediated via SxIP domains.…”