Flucytosine resistance in Cryptococcus gattii is indirectly mediated by the FCY2-FCY1-FUR1 pathway

Kiem Vu, George Richard Thompson, Chandler C. Roe, Jane E Sykes, Elizabeth M. Dreibe, Shawn R. Lockhart, Wieland Meyer, David M. Engelthaler, Angela C Gelli

Research output: Contribution to journalArticle

1 Citation (Scopus)

Abstract

Cryptococcosis is an opportunistic fungal infection caused by members of the two sibling species complexes: Cryptococcus neoformans and Cryptococcus gattii. Flucytosine (5FC) is one of the most widely used antifungals against Cryptococcus spp., yet very few studies have looked at the molecular mechanisms responsible for 5FC resistance in this pathogen. In this study, we examined 11 C. gattii clinical isolates of the major molecular type VGIII based on differential 5FC susceptibility and asked whether there were genomic changes in the key genes involved in flucytosine metabolism. Susceptibility assays and sequencing analysis revealed an association between a point mutation in the cytosine deaminase gene (FCY1) and 5FC resistance in two of the studied 5FC resistant C. gattii VGIII clinical isolates, B9322 and JS5. This mutation results in the replacement of arginine for histidine at position 29 and occurs within a variable stretch of amino acids. Heterologous expression of FCY1 and spot sensitivity assays, however, demonstrated that this point mutation did not have any effect on FCY1 activities and was not responsible for 5FC resistance. Comparative sequence analysis further showed that no changes in the amino acid sequence and no genomic alterations were observed within 1 kb of the upstream and downstream sequences of either cytosine permeases (FCY2-4) or uracil phosphoribosyltransferase (FUR1) genes in 5FC resistant and 5FC susceptible C. gattii VGIII isolates. The herein obtained results suggest that the observed 5FC resistance in the isolates B9322 and JS5 is due to changes in unknown protein(s) or pathway(s) that regulate flucytosine metabolism.

Original languageEnglish (US)
Pages (from-to)857-867
Number of pages11
JournalMedical Mycology
Volume56
Issue number7
DOIs
StatePublished - Oct 1 2018

Fingerprint

Cryptococcus gattii
Flucytosine
uracil phosphoribosyltransferase
Point Mutation
Cytosine Deaminase
Genes
Cryptococcus
Cryptococcosis
Cryptococcus neoformans
Membrane Transport Proteins
Mycoses
Cytosine
Opportunistic Infections
Histidine
Sequence Analysis
Arginine
Amino Acid Sequence
Amino Acids
Mutation
Proteins

Keywords

  • Cryptococcus gattii
  • cytosine deaminase
  • FCY1
  • flucytosine resistance

ASJC Scopus subject areas

  • Infectious Diseases

Cite this

Flucytosine resistance in Cryptococcus gattii is indirectly mediated by the FCY2-FCY1-FUR1 pathway. / Vu, Kiem; Thompson, George Richard; Roe, Chandler C.; Sykes, Jane E; Dreibe, Elizabeth M.; Lockhart, Shawn R.; Meyer, Wieland; Engelthaler, David M.; Gelli, Angela C.

In: Medical Mycology, Vol. 56, No. 7, 01.10.2018, p. 857-867.

Research output: Contribution to journalArticle

Vu, Kiem ; Thompson, George Richard ; Roe, Chandler C. ; Sykes, Jane E ; Dreibe, Elizabeth M. ; Lockhart, Shawn R. ; Meyer, Wieland ; Engelthaler, David M. ; Gelli, Angela C. / Flucytosine resistance in Cryptococcus gattii is indirectly mediated by the FCY2-FCY1-FUR1 pathway. In: Medical Mycology. 2018 ; Vol. 56, No. 7. pp. 857-867.
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abstract = "Cryptococcosis is an opportunistic fungal infection caused by members of the two sibling species complexes: Cryptococcus neoformans and Cryptococcus gattii. Flucytosine (5FC) is one of the most widely used antifungals against Cryptococcus spp., yet very few studies have looked at the molecular mechanisms responsible for 5FC resistance in this pathogen. In this study, we examined 11 C. gattii clinical isolates of the major molecular type VGIII based on differential 5FC susceptibility and asked whether there were genomic changes in the key genes involved in flucytosine metabolism. Susceptibility assays and sequencing analysis revealed an association between a point mutation in the cytosine deaminase gene (FCY1) and 5FC resistance in two of the studied 5FC resistant C. gattii VGIII clinical isolates, B9322 and JS5. This mutation results in the replacement of arginine for histidine at position 29 and occurs within a variable stretch of amino acids. Heterologous expression of FCY1 and spot sensitivity assays, however, demonstrated that this point mutation did not have any effect on FCY1 activities and was not responsible for 5FC resistance. Comparative sequence analysis further showed that no changes in the amino acid sequence and no genomic alterations were observed within 1 kb of the upstream and downstream sequences of either cytosine permeases (FCY2-4) or uracil phosphoribosyltransferase (FUR1) genes in 5FC resistant and 5FC susceptible C. gattii VGIII isolates. The herein obtained results suggest that the observed 5FC resistance in the isolates B9322 and JS5 is due to changes in unknown protein(s) or pathway(s) that regulate flucytosine metabolism.",
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T1 - Flucytosine resistance in Cryptococcus gattii is indirectly mediated by the FCY2-FCY1-FUR1 pathway

AU - Vu, Kiem

AU - Thompson, George Richard

AU - Roe, Chandler C.

AU - Sykes, Jane E

AU - Dreibe, Elizabeth M.

AU - Lockhart, Shawn R.

AU - Meyer, Wieland

AU - Engelthaler, David M.

AU - Gelli, Angela C

PY - 2018/10/1

Y1 - 2018/10/1

N2 - Cryptococcosis is an opportunistic fungal infection caused by members of the two sibling species complexes: Cryptococcus neoformans and Cryptococcus gattii. Flucytosine (5FC) is one of the most widely used antifungals against Cryptococcus spp., yet very few studies have looked at the molecular mechanisms responsible for 5FC resistance in this pathogen. In this study, we examined 11 C. gattii clinical isolates of the major molecular type VGIII based on differential 5FC susceptibility and asked whether there were genomic changes in the key genes involved in flucytosine metabolism. Susceptibility assays and sequencing analysis revealed an association between a point mutation in the cytosine deaminase gene (FCY1) and 5FC resistance in two of the studied 5FC resistant C. gattii VGIII clinical isolates, B9322 and JS5. This mutation results in the replacement of arginine for histidine at position 29 and occurs within a variable stretch of amino acids. Heterologous expression of FCY1 and spot sensitivity assays, however, demonstrated that this point mutation did not have any effect on FCY1 activities and was not responsible for 5FC resistance. Comparative sequence analysis further showed that no changes in the amino acid sequence and no genomic alterations were observed within 1 kb of the upstream and downstream sequences of either cytosine permeases (FCY2-4) or uracil phosphoribosyltransferase (FUR1) genes in 5FC resistant and 5FC susceptible C. gattii VGIII isolates. The herein obtained results suggest that the observed 5FC resistance in the isolates B9322 and JS5 is due to changes in unknown protein(s) or pathway(s) that regulate flucytosine metabolism.

AB - Cryptococcosis is an opportunistic fungal infection caused by members of the two sibling species complexes: Cryptococcus neoformans and Cryptococcus gattii. Flucytosine (5FC) is one of the most widely used antifungals against Cryptococcus spp., yet very few studies have looked at the molecular mechanisms responsible for 5FC resistance in this pathogen. In this study, we examined 11 C. gattii clinical isolates of the major molecular type VGIII based on differential 5FC susceptibility and asked whether there were genomic changes in the key genes involved in flucytosine metabolism. Susceptibility assays and sequencing analysis revealed an association between a point mutation in the cytosine deaminase gene (FCY1) and 5FC resistance in two of the studied 5FC resistant C. gattii VGIII clinical isolates, B9322 and JS5. This mutation results in the replacement of arginine for histidine at position 29 and occurs within a variable stretch of amino acids. Heterologous expression of FCY1 and spot sensitivity assays, however, demonstrated that this point mutation did not have any effect on FCY1 activities and was not responsible for 5FC resistance. Comparative sequence analysis further showed that no changes in the amino acid sequence and no genomic alterations were observed within 1 kb of the upstream and downstream sequences of either cytosine permeases (FCY2-4) or uracil phosphoribosyltransferase (FUR1) genes in 5FC resistant and 5FC susceptible C. gattii VGIII isolates. The herein obtained results suggest that the observed 5FC resistance in the isolates B9322 and JS5 is due to changes in unknown protein(s) or pathway(s) that regulate flucytosine metabolism.

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KW - cytosine deaminase

KW - FCY1

KW - flucytosine resistance

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