The RAM signaling pathway links morphology, thermotolerance, and CO2 tolerance in the global fungal pathogen Cryptococcus neoformans

  1. Benjamin J Chadwick
  2. Tuyetnhu Pham
  3. Xiaofeng Xie
  4. Laura C Ristow
  5. Damian J Krysan
  6. Xiaorong Lin  Is a corresponding author
  1. University of Georgia, United States
  2. University of Iowa, United States

Abstract

The environmental pathogen Cryptococcus neoformans claims over 180,000 lives each year. Survival of this basidiomycete at host CO2 concentrations has only recently been considered an important virulence trait. Through screening gene knockout libraries constructed in a CO2-tolerant clinical strain, we found mutations leading to CO2 sensitivity are enriched in pathways activated by heat stress, including calcineurin, Ras1-Cdc24, cell wall integrity, and Regulator of Ace2 and Morphogenesis (RAM). Overexpression of Cbk1, the conserved terminal kinase of the RAM pathway, partially restored defects of these mutants at host CO2 or temperature levels. In ascomycetes such as Saccharomyces cerevisiae and Candida albicans, transcription factor Ace2 is an important target of Cbk1, activating genes responsible for cell separation. However, no Ace2 homolog or any downstream component of the RAM pathway has been identified in basidiomycetes. Through in vitro evolution and comparative genomics, we characterized mutations in suppressors of cbk1D in C. neoformans that partially rescued defects in CO2 tolerance, thermotolerance, and morphology. One suppressor is the RNA translation repressor Ssd1, which is highly conserved in ascomycetes and basidiomycetes. The other is a novel ribonuclease domain-containing protein, here named PSC1, which is present in basidiomycetes and humans but surprisingly absent in most ascomycetes. Loss of Ssd1 in cbk1D partially restored cryptococcal ability to survive and amplify in the inhalation and intravenous murine models of cryptococcosis. Our discoveries highlight the overlapping regulation of CO2 tolerance and thermotolerance, the essential role of the RAM pathway in cryptococcal adaptation to the host condition, and the potential importance of post-transcriptional control of virulence traits in this global pathogen.

Data availability

Sequences generated from this research has been deposited to the Sequence Read Archive (SRA) under project accession number: PRJNA791949.

The following data sets were generated

Article and author information

Author details

  1. Benjamin J Chadwick

    Department of Plant Biology, University of Georgia, Athens, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8244-6190
  2. Tuyetnhu Pham

    Department of Plant Biology, University of Georgia, Athens, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Xiaofeng Xie

    Department of Microbiology, University of Georgia, Athens, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Laura C Ristow

    Department of Pediatrics, University of Iowa, Iowa City, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Damian J Krysan

    Department of Pediatrics, University of Iowa, Iowa City, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Xiaorong Lin

    Department of Microbiology, University of Georgia, Athens, United States
    For correspondence
    Xiaorong.Lin@uga.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3390-8387

Funding

National Institutes of Health (R01AI147541)

  • Damian J Krysan
  • Xiaorong Lin

National Institutes of Health (R01AI140719)

  • Xiaorong Lin

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Arturo Casadevall, Johns Hopkins Bloomberg School of Public Health, United States

Ethics

Animal experimentation: This study was performed according to the guidelines of NIH and the University of Georgia Institutional Animal Care and Use Committee (IACUC). The animal models and procedures used have been approved by the IACUC (AUP protocol numbers: A2017 08-023 and A2020 06-015).

Version history

  1. Received: August 9, 2022
  2. Preprint posted: August 14, 2022 (view preprint)
  3. Accepted: November 22, 2022
  4. Accepted Manuscript published: November 23, 2022 (version 1)
  5. Version of Record published: November 29, 2022 (version 2)

Copyright

© 2022, Chadwick et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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  1. Benjamin J Chadwick
  2. Tuyetnhu Pham
  3. Xiaofeng Xie
  4. Laura C Ristow
  5. Damian J Krysan
  6. Xiaorong Lin
(2022)
The RAM signaling pathway links morphology, thermotolerance, and CO2 tolerance in the global fungal pathogen Cryptococcus neoformans
eLife 11:e82563.
https://doi.org/10.7554/eLife.82563

Share this article

https://doi.org/10.7554/eLife.82563

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