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Deciphering the Role of EGL-3 for Neuropeptides Processing in Caenorhabditis elegans Using High-Resolution Quadrupole–Orbitrap Mass Spectrometry

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Abstract

Neuropeptides are derived from large and inactive proteins which require endoproteolytic processing for the biosynthesis of the bioactive peptides. The maturation of pro-neuropeptide to neuropeptide is believed to be performed by ortholog pro-protein convertase EGL-3 in Caenorhabditis elegans (C. elegans). Furthermore, ortholog of Cathepsin L, CPL-1 are found in C. elegans and can potentially cleave paired basic amino acids at the N-terminal suggesting the presence of both pathways. The objective of this study was to decipher the role of EGL-3 in the proteolysis of FMRF amide-related peptides (FLPs) or neuropeptide-like proteins (NLPs) using synthetic surrogate peptides based on a universal enzymatic cleavage pattern published by Schechter and Berger and used widely in enzymology. The results show evidence that proteolysis controls FLP-21 and NLP-8 related neuropeptide levels in C. elegans. Surrogate peptides were degraded rapidly when exposed to C. elegans S9 fractions leading to the formation of specific peptide fragments related to EGL-3 and CPL-1 pathway. The results suggest that CPL-1 pathway does not compensate for the loss of the EGL-3 pathway. Proteolysis of pro-neuropeptides associated to FLP-21 and NLP-8 in elg-3 mutants are severely hampered leading to a lack of mature bioactive neuropeptides.

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Fig. 1

(Reproduced with permission from Hook et al. [6]). b FLP-21 processing by proprotein convertases EGL-3 leading to active FLP-21 neuropeptide. c NLP-8 processing by proprotein convertases EGL-3 and carboxypeptidase E EGL-21 leading to several actives neuropeptides. Identification of surrogate peptides used for further in vitro exploration was based on subsite nomenclature adopted from a scheme developed by Schechter and Berger [26, 27] and used to describe of enzyme specificities

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Acknowledgements

This project was funded by the National Sciences and Engineering Research Council of Canada (F. Beaudry discovery Grant No. RGPIN-2015-05071). The mass spectrometry analyses were performed using an infrastructure funded by the Canadian Foundation for Innovation (CFI) and the Fonds de Recherche du Québec (FRQ), Government of Quebec (F. Beaudry CFI John R. Evans Leaders Grant No. 36706). A PhD scholarship was awarded to J. Ben Salam with a grant obtained from Fondation de France (DN Arvanitis Grant No. RAF18002BBA).

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  1. Groupe de Recherche en Pharmacologie Animal du Québec (GREPAQ), Département de Biomédecine Vétérinaire, Faculté de Médecine Vétérinaire, Université de Montréal, 3200 Sicotte, Saint-Hyacinthe, QC, J2S 2M2, Canada

    Jennifer Ben Salem, Bruno Nkambeu & Francis Beaudry

  2. Institut des Maladies Métaboliques et Cardiovasculaires, INSERM UMR1048, Université de Toulouse, 118 route de Narbonne, 31062, Toulouse Cedex 9, France

    Jennifer Ben Salem & Dina N. Arvanitis

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  1. Jennifer Ben Salem
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  2. Bruno Nkambeu
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  4. Francis Beaudry
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Correspondence to Francis Beaudry.

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Salem, J.B., Nkambeu, B., Arvanitis, D.N. et al. Deciphering the Role of EGL-3 for Neuropeptides Processing in Caenorhabditis elegans Using High-Resolution Quadrupole–Orbitrap Mass Spectrometry. Neurochem Res 43, 2121–2131 (2018). https://doi.org/10.1007/s11064-018-2636-2

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  • DOI: https://doi.org/10.1007/s11064-018-2636-2

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