Brain-derived neurotrophic factor receptor TrkB exists as a preformed dimer in living cells

Abstract

BackgroundNeurotrophins (NTs) and their receptors play crucial roles in the development, functions and maintenance of nervous systems. It is widely believed that NT-induced dimerization of the receptors initiates the transmembrane signaling. However, it is still controversial whether the receptor molecule has a monomeric or dimeric structure on the cell surface before its ligand binding.FindingsUsing chemical cross-linking, bimolecular fluorescence complementation (BiFC) and luciferase fragment complementation (LFC) assays, in this study, we show the brain-derived neurotrophic factor (BDNF) receptor TrkB exists as a homodimer before ligand binding. We have also found by using BiFC and LFC that the dimer forms in the endoplasmic reticulum (ER), and that the receptor lacking its intracellular domain cannot form the dimeric structure.ConclusionsMost, if not all, of the TrkB receptor has a preformed, yet inactive, homodimeric structure before BDNF binding. The intracellular domain of TrkB plays a crucial role in the spontaneous dimerization of the newly synthesized receptors, which occurs in ER. These findings provide new insight into an understanding of a molecular mechanism underlying transmembrane signaling mediated by NT receptors.

Mechanisms of Activation of Receptor Tyrosine Kinases: Monomers or Dimers

“…However, an increasing number of studies demonstrate that RTKs exist as pre-formed, yet inactive, dimers, even in the absence of activating ligand [15,16,17,18,19,20,21,22,23,24,25,26,27,28,29]. IR and IGF-IR are also expressed as disulfide-linked (αβ) 2 homodimers at the cell surface [30].…”

Reconstitution of Torso signaling in cultured cells suggests a role for both Trunk and Torso-like in receptor activation

“…Indeed, when Torso was expressed alone we saw an almost twofold increase in reporter activity relative to vector control. Although some receptor tyrosine kinases remain in a monomeric state until binding of ligand leads to dimerization (Ullrich and Schlessinger, 1990;Lemmon and Schlessinger, 2010), others are present in an inactive dimeric state with a conformation in which the intracellular domains of the two proteins are unable to interact productively in the absence of ligand (Ward et al, 2007;Tao and Maruyama, 2008;Shen and Maruyama, 2012). Recently, it has been reported that B. mori Torso expressed in Drosophila S2 cells undergoes ligand-independent dimerization via disulfide linkages formed by cysteine residues present in the transmembrane domain (Konogami et al, 2016).…”

“…To examine whether dimerization of Torso in response to Trk or Tsl could be detected in our system, we used bimolecular fluorescence complementation (BiFC) (Hu et al, 2002;Kodama and Hu, 2010;Tao and Maruyama, 2008;Shen and Maruyama, 2012), a method for visualizing interactions between proteins via dimerization-mediated complementation of split fluorescent proteins. We generated constructs to express wild-type and Torso (Fig.…”

“…The ligand that activates Torso to regulate these two behaviors is prothoracicotropic hormone (PTTH) (Kawakami et al, 1990;Rewitz et al, 2009), a secreted cystine knot-containing peptide that is expressed in a bilateral pair of neurons in the brain. PTTH expressed in these neurons activates Torso in cells of the prothoracic gland (Siegmund and Korge, 2001;Rybczynski, 2005;McBrayer et al, 2007), in photoreceptive dendritic arborization neurons in the body wall of the larva (Xiang et al, 2010) and in cells of Bolwig's organ (Hassan et al, 2005;Mazzoni et al, 2005). Although the extent to which Tsl participates in the activation of Torso in the larval prothoracic gland remains controversial (Grillo et al, 2012;Johnson et al, 2013), transgenic expression of PTTH in the embryo activates Torso in a Tsl-independent manner, leading to a terminalizing phenotype similar to that produced by Torso gainof-function mutations.…”

TrkB interacts with ErbB4 and regulates NRG1-induced NR2B phosphorylation in cortical neurons before synaptogenesis

“…pBiFC-TrkB-FL and pBiFC-TrkB-Δ-ICD constructs were kindly provided by Dr Maruyama, Okinawa Institute of Science and Technology, Japan [39]. These constructs were transfected into primary cortical neurons using Effectene Transfection Reagent (Qiagen) 48 h before NRG1 treatment.…”