Legionella pneumophila Cas2 Promotes the Expression of Small Heat Shock Protein C2 That Is Required for Thermal Tolerance and Optimal Intracellular Infection

Jackson A. Campbell, Nicholas P. Cianciotto*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

3 Scopus citations

Abstract

Previously, we demonstrated that Cas2 encoded within the CRISPR-Cas locus of Legionella pneumophila strain 130b promotes the ability of the Legionella pathogen to infect amoebal hosts. Given that L. pneumophila Cas2 has RNase activity, we posited that the cytoplasmic protein is regulating the expression of another Legionella gene(s) that fosters intracellular infection. Proteomics revealed 10 proteins at diminished levels in the cas2 mutant, and reverse transcription-quantitative (qRT-PCR) confirmed the reduced expression of a gene encoding putative small heat shock protein C2 (HspC2), among several others. As predicted, the gene was expressed more highly at 37°C to 50°C than that at 30°C, and an hspC2 mutant, but not its complemented derivative, displayed ;100-fold reduced CFU following heat shock at 55°C. Compatible with the effect of Cas2 on hspC2 expression, strains lacking Cas2 also had impaired thermal tolerance. The hspC2 mutant, like the cas2 mutant before it, was greatly impaired for infection of Acanthamoeba castellanii, a frequent host for legionellae in waters. HspC2 and Cas2 were not required for entry into these host cells but promoted the replicative phase of intracellular infection. Finally, the hspC2 mutant exhibited an additional defect during the infection of macrophages, which are the primary host for legionellae during lung infection. In summary, hspC2 is upregulated by the presence of Cas2, and HspC2 uniquely promotes both L. pneumophila extracellular survival at high temperatures and infection of amoebal and human host cells. To our knowledge, these findings also represent the first genetic proof linking Cas2 to thermotolerance, expanding the repertoire of noncanonical functions associated with CRISPR-Cas proteins.

Original languageEnglish (US)
JournalInfection and immunity
Volume90
Issue number10
DOIs
StatePublished - Oct 2022

Keywords

  • CRISPR-Cas
  • Cas2
  • Legionella pneumophila
  • intracellular infection
  • small heat shock protein
  • thermal tolerance

ASJC Scopus subject areas

  • Infectious Diseases
  • Parasitology
  • Microbiology
  • Immunology

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