C. elegans SWAN-1 Binds to EGL-9 and regulates HIF-1-mediated resistance to the bacterial pathogen Pseudomonas aeruginosa PAO1

PLoS Pathog. 2010 Aug 26;6(8):e1001075. doi: 10.1371/journal.ppat.1001075.

Abstract

Pseudomonas aeruginosa is a nearly ubiquitous human pathogen, and infections can be lethal to patients with impaired respiratory and immune systems. Prior studies have established that strong loss-of-function mutations in the egl-9 gene protect the nematode C. elegans from P. aeruginosa PAO1 fast killing. EGL-9 inhibits the HIF-1 transcription factor via two pathways. First, EGL-9 is the enzyme that targets HIF-1 for oxygen-dependent degradation via the VHL-1 E3 ligase. Second, EGL-9 inhibits HIF-1-mediated gene expression through a VHL-1-independent mechanism. Here, we show that a loss-of-function mutation in hif-1 suppresses P. aeruginosa PAO1 resistance in egl-9 mutants. Importantly, we find stabilization of HIF-1 protein is not sufficient to protect C. elegans from P. aeruginosa PAO1 fast killing. However, mutations that inhibit both EGL-9 pathways result in higher levels of HIF-1 activity and confer resistance to the pathogen. Using forward genetic screens, we identify additional mutations that confer resistance to P. aeruginosa. In genetic backgrounds that stabilize C. elegans HIF-1 protein, loss-of-function mutations in swan-1 increase the expression of hypoxia response genes and protect C. elegans from P. aeruginosa fast killing. SWAN-1 is an evolutionarily conserved WD-repeat protein belonging to the AN11 family. Yeast two-hybrid and co-immunoprecipitation assays show that EGL-9 forms a complex with SWAN-1. Additionally, we present genetic evidence that the DYRK kinase MBK-1 acts downstream of SWAN-1 to promote HIF-1-mediated transcription and to increase resistance to P. aeruginosa. These data support a model in which SWAN-1, MBK-1 and EGL-9 regulate HIF-1 transcriptional activity and modulate resistance to P. aeruginosa PAO1 fast killing.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Caenorhabditis elegans / genetics
  • Caenorhabditis elegans / immunology*
  • Caenorhabditis elegans / metabolism
  • Caenorhabditis elegans Proteins / genetics
  • Caenorhabditis elegans Proteins / immunology*
  • Caenorhabditis elegans Proteins / metabolism
  • Gene Expression
  • Gene Expression Regulation
  • Immunoblotting
  • Immunoprecipitation
  • Mutation
  • Pseudomonas Infections / immunology*
  • Pseudomonas aeruginosa / immunology
  • Reverse Transcriptase Polymerase Chain Reaction
  • Transcription Factors / genetics
  • Transcription Factors / immunology
  • Transcription Factors / metabolism*
  • Two-Hybrid System Techniques

Substances

  • Caenorhabditis elegans Proteins
  • Egl-9 protein, C elegans
  • HIF-1 protein, C elegans
  • Transcription Factors