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ZFIN ID: ZDB-PUB-160702-5
Visualization of Neuregulin 1 ectodomain shedding reveals its local processing in vitro and in vivo
Kamezaki, A., Sato, F., Aoki, K., Asakawa, K., Kawakami, K., Matsuzaki, F., Sehara-Fujisawa, A.
Date: 2016
Source: Scientific Reports   6: 28873 (Journal)
Registered Authors: Asakawa, Kazuhide, Kawakami, Koichi, Sehara-Fujisawa, Atsuko
Keywords: Fluorescence imaging, Protein quality control
MeSH Terms:
  • Animals
  • Animals, Genetically Modified
  • Binding Sites/genetics
  • Cell Line
  • Embryo, Nonmammalian/metabolism
  • Green Fluorescent Proteins/genetics
  • Green Fluorescent Proteins/metabolism*
  • HEK293 Cells
  • Humans
  • Luminescent Proteins/genetics
  • Luminescent Proteins/metabolism*
  • Mice
  • Microscopy, Fluorescence
  • Neuregulin-1/genetics
  • Neuregulin-1/metabolism*
  • Phorbol Esters/pharmacology
  • Protein Processing, Post-Translational*
  • Proteolysis/drug effects
  • Time-Lapse Imaging/methods*
  • Zebrafish/embryology
  • Zebrafish/genetics
  • Zebrafish/metabolism
PubMed: 27364328 Full text @ Sci. Rep.
FIGURES
ABSTRACT
Neuregulin1 (NRG1) plays diverse developmental roles and is likely involved in several neurological disorders including schizophrenia. The transmembrane NRG1 protein is proteolytically cleaved and released as a soluble ligand for ErbB receptors. Such post-translational processing, referred to as 'ectodomain shedding', is thought to be crucial for NRG1 function. However, little is known regarding the regulatory mechanism of NRG1 cleavage in vivo. Here, we developed a fluorescent probe, NRG1 Cleavage Indicating SenSOR (N-CISSOR), by fusing mCherry and GFP to the extracellular and intracellular domains of NRG1, respectively. N-CISSOR mimicked the subcellular localization and biochemical properties of NRG1 including cleavage dynamics and ErbB phosphorylation in cultured cells. mCherry/GFP ratio imaging of phorbol-12-myristate-13-acetate-stimulated N-CISSOR-expressing HEK293T cells enabled to monitor rapid ectodomain shedding of NRG1 at the subcellular level. Utilizing N-CISSOR in zebrafish embryos revealed preferential axonal NRG1 ectodomain shedding in developing motor neurons, demonstrating that NRG1 ectodomain shedding is spatially regulated at the subcellular level. Thus, N-CISSOR will be a valuable tool for elucidating the spatiotemporal regulation of NRG1 ectodomain shedding, both in vitro and in vivo.
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