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Semi-Quantitative Analysis of Peptidoglycan by Liquid Chromatography Mass Spectrometry and Bioinformatics
09:09

Semi-Quantitative Analysis of Peptidoglycan by Liquid Chromatography Mass Spectrometry and Bioinformatics

Published on: October 13, 2020

Peptidoglycan activation of the proPO-system without a peptidoglycan receptor protein (PGRP)?

Haipeng Liu1, Chenglin Wu, Yasuyuki Matsuda

  • 1Department of Comparative Physiology, Uppsala University, Norbyvägen 18A, SE-752 36 Uppsala, Sweden.

Developmental and Comparative Immunology
|August 18, 2010
PubMed

Insights

Lysine-type peptidoglycan (PGN) activates the prophenoloxidase (proPO) system in crayfish, triggering immune responses. Two serine proteinase homologues (Pl-SPHs) and LGBP are involved in this PGN-induced proPO activation, notably without pattern recognition receptors (PGRPs).

Area of Science:

  • Innate immunity
  • Crustacean physiology
  • Biochemistry

Background:

  • Pattern recognition receptors (PGRPs) initiate the prophenoloxidase (proPO) cascade upon microbial polysaccharide recognition, leading to melanin synthesis.
  • Serine proteinase homologues (SPHs) and masquerade-like proteins are implicated in insect proPO activation.
  • The role of peptidoglycan (PGN) and its recognition in crustacean proPO activation remains largely uncharacterized.

Purpose of the Study:

  • To identify novel proteins involved in the prophenoloxidase (proPO) system activation in the freshwater crayfish, Pacifastacus leniusculus.
  • To investigate the role of Lysine-type peptidoglycan (PGN) as a trigger for proPO activation in crustaceans.
  • To elucidate the involvement of serine proteinase homologues (SPHs) and lipopolysaccharide- and β-1,3-glucan-binding protein (LGBP) in PGN-mediated immune responses.

Main Methods:

  • Isolation and characterization of proteins from crayfish hemocytes based on binding to Lys-type peptidoglycan (PGN).
  • Utilizing various PGN-binding assays to identify interacting proteins, including Pl-SPH2, Pl-SPH1, and LGBP.
  • Employing RNA interference (RNAi) in crayfish hematopoietic cell cultures to assess the functional role of identified proteins in proPO activity.

Main Results:

  • A novel serine proteinase homologue, Pl-SPH2, was identified and isolated from crayfish hemocytes.
  • Lysine-type peptidoglycan (PGN) was confirmed as a potent activator of the proPO system in crayfish.
  • RNA interference of Pl-SPH2, Pl-SPH1, or LGBP led to reduced proPO activity upon PGN stimulation, indicating their crucial roles.
  • No PGRP homologues were detected in the crayfish, suggesting a PGRP-independent activation pathway.

Conclusions:

  • Lysine-type peptidoglycan (PGN) is a novel trigger for the prophenoloxidase (proPO) system in crustaceans.
  • Two serine proteinase homologues (Pl-SPHs) and LGBP are essential components in the PGN-induced proPO activation pathway.
  • The proPO system activation by Lys-type PGN in crayfish operates independently of pattern recognition receptors (PGRPs).

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