Low-Density Lipoprotein Receptor Related protein-1 (LRP1)-Dependent Cell Signaling Promotes Neurotrophic Activity in Embryonic Sensory Neurons

Kazuyo Yamauchi,Elisabetta Mantuano,Kazuhisa Takahashi,Kenichi Murakami,Tomonori Yamauchi,Wendy Marie Campana,Kenneth Henry,Subhabrata Sanyal

doi:10.1371/journal.pone.0075497

Abstract

Developing sensory neurons require neurotrophic support for survival, neurite outgrowth and myelination. The low-density lipoprotein receptor-related protein-1 (LRP1) transactivates Trk receptors and thereby functions as a putative neurotrophin. Herein, we show that LRP1 is abundantly expressed in developing dorsal root ganglia (DRG) and that LRP1-dependent cell signaling supports survival, neurite extension and receptivity to Schwann cells even in the absence of neurotrophins. Cultured embryonic DRG neurons (E15) were treated with previously characterized LRP1 ligands, LRP1-receptor binding domain of α2-macroglobulin (RBD), hemopexin domain of MMP-9 (PEX) or controls (GST) for two weeks. These structurally diverse LRP1 ligands significantly activated and sustained extracellular signal-regulated kinases (ERK1/2) 5-fold (p<0.05), increased expression of growth-associated protein-43(GAP43) 15-fold (P<0.01), and increased neurite outgrowth 20-fold (P<0.01). Primary sensory neurons treated with LRP1 ligands survived > 2 weeks in vitro, to an extent equaling NGF, a finding associated with canonical signaling mechanisms and blockade of caspase-3 cleavage. LRP1 ligand-induced survival and sprouting were blocked by co-incubation with the LRP1 antagonist, receptor associated protein (RAP), whereas RAP had no effect on NGF-induced activity. Site directed mutagenesis of the LRP1 ligand, RBD, in which Lys1370 and Lys1374 are converted to alanine to preclude LRP1 binding, were ineffective in promoting cell signaling, survival or inducing neurite extension in primary sensory neurons, confirming LRP1 specificity. Furthermore, LRP1-induced neurite sprouting was mediated by Src-family kinase (SFK) activation, suggesting transactivation of Trk receptors. Co-cultures of primary embryonic neurons and Schwann cells showed that LRP1 agonists promoted axonal receptivity to myelination to Schwann cells. Collectively, these findings identify LRP1 as a novel and perhaps essential trophic molecule for sensory neuronal survival and development.

Highlights

Embryonic sensory neurons undergo substantial programmed cell death during development [1] to ensure balance between neuronal numbers and the sizes of their target areas
We showed that the receptor binding domain of α2-macroglobulin (RBD) and other lipoprotein receptor-related protein-1 (LRP1) ligands rapidly induced activation of ERK1/2 [11,19]
We implicate LRP1 as a novel neurotrophic receptor for embryonic sensory neurons. These results provide the first evidence that a new class of receptors, the lowdensity lipoprotein (LDL) receptor gene family, regulate survival and growth of developing sensory neurons

Summary

Introduction

Embryonic sensory neurons undergo substantial programmed cell death during development [1] to ensure balance between neuronal numbers and the sizes of their target areas. Developing sensory neurons are differentially regulated by various neurotrophins to survive and innervate target tissues. More recent reports have suggested a potential role for a member of the LDL gene receptor family member, the lowdensity lipoprotein (LDL) receptor-related protein-1 (LRP1), in neural development and regeneration. The processed form of the receptor includes an 85 kDa fragment that contains the intracellular and transmembrane domain and a 515 kDa extracellular domain that is capable of binding multiple LRP1 ligands [6]. Diverse ligands including tissue-type plasminogen activator (tPA), matrix metalloproteinase-9 (MMP-9), and activated α2macroglobulin have been shown to activate LRP1-dependent cell signaling [5,8,9,10,11]. LRP1 ligands were shown to transactivate Trk receptors in a Src-family kinase dependent manner [12]

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