Swedish Nerve Growth Factor Mutation (NGFR100W) Defines a Role for TrkA and p75NTR in Nociception
Authors: Sung Kijung , Ferrari Luiz F. , Yang Wanlin , Chung ChiHye , Zhao Xiaobei , Gu Yingli , Lin Suzhen , Zhang Kai , Cui Bianxiao , Pearn Matthew L. , Maloney Michael T. , Mobley William C. , Levine Jon D. , Wu Chengbiao 
Published: February 2018

Nerve growth factor (NGF) exerts multiple functions on target neurons throughout development. The recent discovery of a point mutation leading to a change from arginine to tryptophan at residue 100 in the mature NGFβ sequence (NGFR100W) in patients with hereditary sensory and autonomic neuropathy, type V (HSAN V), made it possible to distinguish the signaling mechanisms that lead to two functionally different outcomes of NGF: trophic versus nociceptive. We performed extensive biochemical, cellular and live imaging experiments to examine the binding and signaling properties of NGFR100W. Our results show that, similar to the wildtype NGF (wtNGF), the naturally occurring NGFR100Wmutant was capable of binding to and activating the TrkA receptor and its downstream signaling pathways to support neuronal survival and differentiation. However, NGFR100Wfailed to bind and stimulate the 75kD neurotrophic factor receptor (p75NTR)-mediated signaling cascades (i.e. the RhoA-Cofilin pathway). Intraplantar injection of NGFR100Winto adult rats induced neither TrkA-mediated thermal nor mechanical acute hyperalgesia, but retained the ability to induce chronic hyperalgesia based on agonism for TrkA signaling. Taken together, our studies provide evidence that NGFR100Wretains trophic support capability through TrkA and one aspect of its nociceptive signaling, but fails to engage p75NTRsignaling pathways. Our findings suggest that wtNGF acts through TrkA to regulate the delayed priming of nociceptive responses. The integration of both TrkA and p75NTRsignaling thus appears to regulate neuroplastic effects of NGF in peripheral nociception.

Significance Statement

In the present study, we characterized the naturally occurring NGFR100Wmutant that is associated with hereditary sensory and autonomic neuropathy, type V. We have demonstrated for the first time that NGFR100Wretains trophic support capability through TrkA but fails to engage p75NTRsignaling pathways. Furthermore, following Intraplantar injection into adult rats, NGFR100Winduced neither thermal nor mechanical acute hyperalgesia, but retained the ability to induce chronic hyperalgesia. We have also provided evidence that the integration of both TrkA-and p75NTR-mediated signaling thus appears to regulate neuroplastic effects of NGF in peripheral nociception. Our study with NGFR100Wsuggests that it is possible to uncouple trophic effect from nociceptive function, both induced by wildtype NGF.

Abbreviations

NGF

nerve growth factor;

NGFR100W

NGF mutation with a change from tryptophan (W) to arginine (R) at the 100 residue.

TrkA

Tropomyosin receptor kinase A;

p75NTR

the 75kD neurotrophic factor receptor;

HSAN V

hereditary sensory and autonomic neuropathy, type V;

BFCN

basal forebrain cholinergic neurons;

PNS

peripheral nervous system;

CNS

central nervous system;

BDNF

brain-derived neurotrophic factor;

NT

neurotrophin;

TrkB

Tropomyosin receptor kinase B;

TrkC

Tropomyosin receptor kinase C;

RhoA

Ras homolog gene family, member A;

NF-κB

nuclear factor kappa B;

Akt

Protein kinase B;

JNK

c-Jun N-terminal kinases;

AD

Alzheimer’s disease;

CSF

cerebrospinal fluid;

HIV

human immunodeficiency virus

CIPA

congenital insensitivity to pain with anhidrosis;

ERK

extracellular signal-related kinase;

PI3K

phosphatidylinositol 3-kinase;

PLCγ

and phospholipase Cγ;

PGE2

Prostaglandin E2;

PCR

polymerase chain reaction;

GFP

green fluorescent protein;

QD

quantum dots;

HEK293FT

Human embryonic kidney 293FT cell line;

KKE

NGF mutant protein with mutation at:K32A/K34A/E35A in the mature sequence;

Δ9/13

NGF mutant protein with deletion of N-terminal 9-13 residues in the mature sequence;

DMEM

Dulbecco’s Modified Eagle’s Medium;

PMSF

phenylmethylsulfonyl fluoride;

SDS-PAGE

sodium dodecyl sulfate polyacrylamide gel electrophoresis;

DMSO

dimethyl sulfoxide;

AS

anti-sense oligos;

MM

mis-matched oligos;

E15.5 DRG

embryonic day 15.5 dorsal root ganglion;

ECD

extracellular domain;

MEM

Minimum Essential Medium;

TIRF

Total Internal Reflection Fluorescence;

EMCCD

electron multiplying charge-coupled device;

PC12

a cell line derived from a pheochromocytoma of the rat adrenal medulla;

BSA

Bovine Serum Albumin;

PBS

phosphate buffered saline;

IgG

Immunoglobulin G;

DIC

Days In Culture;