Programmed cell death 4 in bacterially-challenged Apostichopus japonicus: Molecular cloning, expression analysis and

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Preparation of Cell-lines for Conditional Knockdown of Gene Expression and Measurement of the Knockdown Effects on E4orf4-Induced Cell Death
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Preparation of Cell-lines for Conditional Knockdown of Gene Expression and Measurement of the Knockdown Effects on E4orf4-Induced Cell Death

Published on: October 21, 2012

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Zhimeng Lv1, Chenghua Li1, Yina Shao1

  • 1School of Marine Sciences, Ningbo University, Ningbo, Zhejiang 315211, PR China.

Molecular Immunology
|June 6, 2016
PubMed

Insights

A novel sea cucumber gene, AjPDCD4, is involved in cellular defense. Knocking down AjPDCD4 reduces UV-induced DNA damage and reactive oxygen species (ROS) production in sea cucumber immune cells.

Area of Science:

  • Marine biology
  • Molecular biology
  • Immunology

Background:

  • Programmed cell death 4 (PDCD4) is a key regulator of cellular signaling and immune responses, often involving TOLL pathways.
  • Understanding PDCD4 homologs in invertebrates can provide insights into conserved cellular defense mechanisms.

Purpose of the Study:

  • To clone and characterize a novel PDCD4 homologue gene (AjPDCD4) from the sea cucumber Apostichopus japonicus.
  • To investigate the role of AjPDCD4 in cellular responses to stress, specifically UV-induced damage and reactive oxygen species (ROS) production.

Main Methods:

  • Rapid amplification of cDNA ends (RACE) technique was used for gene cloning.
  • Phylogenetic analysis was performed to determine evolutionary relationships.
  • Spatial expression analysis and quantitative real-time PCR were employed to assess gene expression levels.
  • RNA interference (RNAi) was used to knockdown AjPDCD4 expression in coelomocytes.

Main Results:

  • A full-length AjPDCD4 cDNA was successfully cloned, encoding a 450-amino acid protein with two MA3 domains.
  • AjPDCD4 mRNA was highly expressed in tentacles and lowly in muscle tissue, with lower expression in coelomocytes.
  • Expression of AjPDCD4 was significantly downregulated by Vibrio splendidus challenge and LPS exposure.
  • Knockdown of AjPDCD4 repressed UV-induced ROS production and DNA damage, accompanied by upregulation of NF-kappa B homologue p105.

Conclusions:

  • AjPDCD4 is a functional PDCD4 homologue in Apostichopus japonicus.
  • AjPDCD4 plays a role in modulating DNA damage and ROS production in sea cucumber coelomocytes.
  • The findings suggest AjPDCD4 may influence the TLR pathway in response to cellular stress.
Keywords:
Apostichopus japonicusDNA damageProgrammed cell death 4ROS productionp105

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