Limits...
Fungal Morphology, Iron Homeostasis, and Lipid Metabolism Regulated by a GATA Transcription Factor in Blastomyces dermatitidis.

Marty AJ, Broman AT, Zarnowski R, Dwyer TG, Bond LM, Lounes-Hadj Sahraoui A, Fontaine J, Ntambi JM, Keleş S, Kendziorski C, Gauthier GM - PLoS Pathog. (2015)

Bottom Line: This included genes involved with siderophore biosynthesis and uptake, iron homeostasis, and genes unrelated to iron assimilation.Chromatin immunoprecipitation, RNA interference, and overexpression analyses suggested that SREB was in a negative regulatory circuit with the bZIP transcription factor encoded by HAPX.Both SREB and HAPX affected morphogenesis at 22°C; however, large changes in transcript abundance by gene deletion for SREB or strong overexpression for HAPX were required to alter the phase transition.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, University of Wisconsin, Madison, Madison, Wisconsin, United States of America.

ABSTRACT
In response to temperature, Blastomyces dermatitidis converts between yeast and mold forms. Knowledge of the mechanism(s) underlying this response to temperature remains limited. In B. dermatitidis, we identified a GATA transcription factor, SREB, important for the transition to mold. Null mutants (SREBΔ) fail to fully complete the conversion to mold and cannot properly regulate siderophore biosynthesis. To capture the transcriptional response regulated by SREB early in the phase transition (0-48 hours), gene expression microarrays were used to compare SREB∆ to an isogenic wild type isolate. Analysis of the time course microarray data demonstrated SREB functioned as a transcriptional regulator at 37°C and 22°C. Bioinformatic and biochemical analyses indicated SREB was involved in diverse biological processes including iron homeostasis, biosynthesis of triacylglycerol and ergosterol, and lipid droplet formation. Integration of microarray data, bioinformatics, and chromatin immunoprecipitation identified a subset of genes directly bound and regulated by SREB in vivo in yeast (37°C) and during the phase transition to mold (22°C). This included genes involved with siderophore biosynthesis and uptake, iron homeostasis, and genes unrelated to iron assimilation. Functional analysis suggested that lipid droplets were actively metabolized during the phase transition and lipid metabolism may contribute to filamentous growth at 22°C. Chromatin immunoprecipitation, RNA interference, and overexpression analyses suggested that SREB was in a negative regulatory circuit with the bZIP transcription factor encoded by HAPX. Both SREB and HAPX affected morphogenesis at 22°C; however, large changes in transcript abundance by gene deletion for SREB or strong overexpression for HAPX were required to alter the phase transition.

No MeSH data available.


Related in: MedlinePlus

Deletion of SREB affects the transcription of genes in the glycerolipid biosynthetic pathway.(A) Schematic of the glycerolipid biosynthetic pathway in B. dermatitidis. Putative lipases and esterases involved with sterol, diacylglycerol, and triacylglycerol breakdown are also shown. Differentially expressed (DE) genes are labeled in green. (B) Heat map of fluorescent intensity values (log2) for genes in glycerolipid biosynthetic pathway in wild-type (WT) and SREB∆ isolates. Genes are arranged in the same order as in (A). The time point at which gene transcription was DE in SREB∆ versus WT is adjacent to the heat map. DE genes are labeled in green. (C) Quantitative RT-PCR analysis of a subset of DE glycerolipid biosynthetic genes at 48-hrs 22°C. Log2 fold change values (SREB∆ versus wild-type) for qRT-PCR were compared to gene expression microarray data for BDBG_06687 (glycerol kinase), BDBG_03027 (acetyltransferase), BDBG_07704 (MBOAT family protein), BDBG_02489 (Phospholipid:diacylglycerol acetyltansferase), BDBG_07403 (Sterol-O-acyltransferase). The qRT-PCR results were averaged from 2 experiments.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4482641&req=5

ppat.1004959.g005: Deletion of SREB affects the transcription of genes in the glycerolipid biosynthetic pathway.(A) Schematic of the glycerolipid biosynthetic pathway in B. dermatitidis. Putative lipases and esterases involved with sterol, diacylglycerol, and triacylglycerol breakdown are also shown. Differentially expressed (DE) genes are labeled in green. (B) Heat map of fluorescent intensity values (log2) for genes in glycerolipid biosynthetic pathway in wild-type (WT) and SREB∆ isolates. Genes are arranged in the same order as in (A). The time point at which gene transcription was DE in SREB∆ versus WT is adjacent to the heat map. DE genes are labeled in green. (C) Quantitative RT-PCR analysis of a subset of DE glycerolipid biosynthetic genes at 48-hrs 22°C. Log2 fold change values (SREB∆ versus wild-type) for qRT-PCR were compared to gene expression microarray data for BDBG_06687 (glycerol kinase), BDBG_03027 (acetyltransferase), BDBG_07704 (MBOAT family protein), BDBG_02489 (Phospholipid:diacylglycerol acetyltansferase), BDBG_07403 (Sterol-O-acyltransferase). The qRT-PCR results were averaged from 2 experiments.

Mentions: GO enrichment and WGCNA analyses suggested that deletion of SREB affected lipid metabolism at 37°C and 22°C. To investigate for defects in lipid biosynthesis, glycerolipids (triacylglycerol (TAG) and diacylglycerol (DAG)), phospholipids, free fatty acids, and ergosterol were extracted from WT and SREB∆ at 37°C and 6, 24, and 48-hrs at 22°C. Gene expression microarray analysis indicated 8 of 14 genes in the glycerolipid pathway involved with TAG biosynthesis were DE in SREB∆ versus WT (Fig 5A and 5B). Seven DE genes had decreased transcript abundance at 22°C, whereas a homolog of S. cerevisiae AYR1 (BDBG_09292) had increased transcription (Fig 5A and 5B). Quantitative RT-PCR analysis validated a subset of these DE genes; log2 fold changes were similar for qRT-PCR and microarray analyses at 48-hrs 22°C (Fig 5C). Gas chromatographic analysis of extracted lipids from SREB∆ suggested decreased TAG concentrations at 37°C and 22°C, and similar levels of DAG compared to WT (Fig 6A and 6B).


Fungal Morphology, Iron Homeostasis, and Lipid Metabolism Regulated by a GATA Transcription Factor in Blastomyces dermatitidis.

Marty AJ, Broman AT, Zarnowski R, Dwyer TG, Bond LM, Lounes-Hadj Sahraoui A, Fontaine J, Ntambi JM, Keleş S, Kendziorski C, Gauthier GM - PLoS Pathog. (2015)

Deletion of SREB affects the transcription of genes in the glycerolipid biosynthetic pathway.(A) Schematic of the glycerolipid biosynthetic pathway in B. dermatitidis. Putative lipases and esterases involved with sterol, diacylglycerol, and triacylglycerol breakdown are also shown. Differentially expressed (DE) genes are labeled in green. (B) Heat map of fluorescent intensity values (log2) for genes in glycerolipid biosynthetic pathway in wild-type (WT) and SREB∆ isolates. Genes are arranged in the same order as in (A). The time point at which gene transcription was DE in SREB∆ versus WT is adjacent to the heat map. DE genes are labeled in green. (C) Quantitative RT-PCR analysis of a subset of DE glycerolipid biosynthetic genes at 48-hrs 22°C. Log2 fold change values (SREB∆ versus wild-type) for qRT-PCR were compared to gene expression microarray data for BDBG_06687 (glycerol kinase), BDBG_03027 (acetyltransferase), BDBG_07704 (MBOAT family protein), BDBG_02489 (Phospholipid:diacylglycerol acetyltansferase), BDBG_07403 (Sterol-O-acyltransferase). The qRT-PCR results were averaged from 2 experiments.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4482641&req=5

ppat.1004959.g005: Deletion of SREB affects the transcription of genes in the glycerolipid biosynthetic pathway.(A) Schematic of the glycerolipid biosynthetic pathway in B. dermatitidis. Putative lipases and esterases involved with sterol, diacylglycerol, and triacylglycerol breakdown are also shown. Differentially expressed (DE) genes are labeled in green. (B) Heat map of fluorescent intensity values (log2) for genes in glycerolipid biosynthetic pathway in wild-type (WT) and SREB∆ isolates. Genes are arranged in the same order as in (A). The time point at which gene transcription was DE in SREB∆ versus WT is adjacent to the heat map. DE genes are labeled in green. (C) Quantitative RT-PCR analysis of a subset of DE glycerolipid biosynthetic genes at 48-hrs 22°C. Log2 fold change values (SREB∆ versus wild-type) for qRT-PCR were compared to gene expression microarray data for BDBG_06687 (glycerol kinase), BDBG_03027 (acetyltransferase), BDBG_07704 (MBOAT family protein), BDBG_02489 (Phospholipid:diacylglycerol acetyltansferase), BDBG_07403 (Sterol-O-acyltransferase). The qRT-PCR results were averaged from 2 experiments.
Mentions: GO enrichment and WGCNA analyses suggested that deletion of SREB affected lipid metabolism at 37°C and 22°C. To investigate for defects in lipid biosynthesis, glycerolipids (triacylglycerol (TAG) and diacylglycerol (DAG)), phospholipids, free fatty acids, and ergosterol were extracted from WT and SREB∆ at 37°C and 6, 24, and 48-hrs at 22°C. Gene expression microarray analysis indicated 8 of 14 genes in the glycerolipid pathway involved with TAG biosynthesis were DE in SREB∆ versus WT (Fig 5A and 5B). Seven DE genes had decreased transcript abundance at 22°C, whereas a homolog of S. cerevisiae AYR1 (BDBG_09292) had increased transcription (Fig 5A and 5B). Quantitative RT-PCR analysis validated a subset of these DE genes; log2 fold changes were similar for qRT-PCR and microarray analyses at 48-hrs 22°C (Fig 5C). Gas chromatographic analysis of extracted lipids from SREB∆ suggested decreased TAG concentrations at 37°C and 22°C, and similar levels of DAG compared to WT (Fig 6A and 6B).

Bottom Line: This included genes involved with siderophore biosynthesis and uptake, iron homeostasis, and genes unrelated to iron assimilation.Chromatin immunoprecipitation, RNA interference, and overexpression analyses suggested that SREB was in a negative regulatory circuit with the bZIP transcription factor encoded by HAPX.Both SREB and HAPX affected morphogenesis at 22°C; however, large changes in transcript abundance by gene deletion for SREB or strong overexpression for HAPX were required to alter the phase transition.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, University of Wisconsin, Madison, Madison, Wisconsin, United States of America.

ABSTRACT
In response to temperature, Blastomyces dermatitidis converts between yeast and mold forms. Knowledge of the mechanism(s) underlying this response to temperature remains limited. In B. dermatitidis, we identified a GATA transcription factor, SREB, important for the transition to mold. Null mutants (SREBΔ) fail to fully complete the conversion to mold and cannot properly regulate siderophore biosynthesis. To capture the transcriptional response regulated by SREB early in the phase transition (0-48 hours), gene expression microarrays were used to compare SREB∆ to an isogenic wild type isolate. Analysis of the time course microarray data demonstrated SREB functioned as a transcriptional regulator at 37°C and 22°C. Bioinformatic and biochemical analyses indicated SREB was involved in diverse biological processes including iron homeostasis, biosynthesis of triacylglycerol and ergosterol, and lipid droplet formation. Integration of microarray data, bioinformatics, and chromatin immunoprecipitation identified a subset of genes directly bound and regulated by SREB in vivo in yeast (37°C) and during the phase transition to mold (22°C). This included genes involved with siderophore biosynthesis and uptake, iron homeostasis, and genes unrelated to iron assimilation. Functional analysis suggested that lipid droplets were actively metabolized during the phase transition and lipid metabolism may contribute to filamentous growth at 22°C. Chromatin immunoprecipitation, RNA interference, and overexpression analyses suggested that SREB was in a negative regulatory circuit with the bZIP transcription factor encoded by HAPX. Both SREB and HAPX affected morphogenesis at 22°C; however, large changes in transcript abundance by gene deletion for SREB or strong overexpression for HAPX were required to alter the phase transition.

No MeSH data available.


Related in: MedlinePlus