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Transcriptional profile of Paracoccidioides induced by oenothein B, a potential antifungal agent from the Brazilian Cerrado plant Eugenia uniflora.

Zambuzzi-Carvalho PF, Tomazett PK, Santos SC, Ferri PH, Borges CL, Martins WS, de Almeida Soares CM, Pereira M - BMC Microbiol. (2013)

Bottom Line: Infection experiments in macrophages corroborated the in vitro results.Several other genes, such as those involved in a variety of important cellular processes (i.e., membrane maintenance, stress and virulence) were found to be up-regulated in response to OenB treatment.The exposure of Paracoccidioides to OenB resulted in a complex altered gene expression profile.

View Article: PubMed Central - HTML - PubMed

Affiliation: Departamento de Bioquímica e Biologia Molecular, Laboratório de Biologia Molecular, Instituto de Ciências Biológicas, ICBII, Campus II, Universidade Federal de Goiás, C,P, 131, 74001-970 Goiânia, GO, Brazil. maristelaufg@gmail.com.

ABSTRACT

Background: The compound oenothein B (OenB), which is isolated from the leaves of Eugenia uniflora, a Brazilian Cerrado plant, interferes with Paracoccidioides yeast cell morphology and inhibits 1,3-β-D-glucan synthase (PbFKS1) transcript accumulation, which is involved in cell wall synthesis. In this work we examined the gene expression changes in Paracoccidioides yeast cells following OenB treatment in order to investigate the adaptive cellular responses to drug stress.

Results: We constructed differential gene expression libraries using Representational Difference Analysis (RDA) of Paracoccidioides yeast cells treated with OenB for 90 and 180 min. Treatment for 90 min resulted in the identification of 463 up-regulated expressed sequences tags (ESTs) and 104 down-regulated ESTs. For the 180 min treatment 301 up-regulated ESTs and 143 down-regulated were identified. Genes involved in the cell wall biosynthesis, such as GLN1, KRE6 and FKS1, were found to be regulated by OenB. Infection experiments in macrophages corroborated the in vitro results. Fluorescence microscopy showed increased levels of chitin in cells treated with OenB. The carbohydrate polymer content of the cell wall of the fungus was also evaluated, and the results corroborated with the transcriptional data. Several other genes, such as those involved in a variety of important cellular processes (i.e., membrane maintenance, stress and virulence) were found to be up-regulated in response to OenB treatment.

Conclusions: The exposure of Paracoccidioides to OenB resulted in a complex altered gene expression profile. Some of the changes may represent specific adaptive responses to this compound in this important pathogenic fungus.

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Functional classification of differentially expressed genes in biological functional groups. (A) Down- and up-regulated genes obtained from cDNAs synthesized from RNAs of Paracoccidioides cells exposure to OenB for 90 min and (B) 180 min. Functional classification was based on Blast X homology of each EST against the GenBank non-redundant database and the nucleotide database from the Paracoccidioides structural genome at a significant homology cut-off ≤ 10-5 and a MIPS functional annotation scheme. Each functional class is represented as a segment and expressed as the total number of ESTs in each library.
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Figure 1: Functional classification of differentially expressed genes in biological functional groups. (A) Down- and up-regulated genes obtained from cDNAs synthesized from RNAs of Paracoccidioides cells exposure to OenB for 90 min and (B) 180 min. Functional classification was based on Blast X homology of each EST against the GenBank non-redundant database and the nucleotide database from the Paracoccidioides structural genome at a significant homology cut-off ≤ 10-5 and a MIPS functional annotation scheme. Each functional class is represented as a segment and expressed as the total number of ESTs in each library.

Mentions: The distribution of differentially expressed genes in the different functional categories and biological processes was evaluated (Table 1; Figure 1). The analysis demonstrated that most of the genes that were down-regulated after 90 min participate in cellular processes related to transport, followed by signal transducers and those involved in transcription and metabolism. On the other hand, the up-regulated genes were mostly found to be functionally unclassified proteins, followed by unclassified proteins, and those involved in cell cycle and DNA processing and metabolism. After 180 min treatment, most of the down-regulated genes were also involved in signal transduction, metabolism and cellular transport. The up-regulated genes were unclassified proteins or those involved in transcription and morphology determination. The percentage of occurrence of each gene in the libraries was indicated in the Additional file 2: Table S1.


Transcriptional profile of Paracoccidioides induced by oenothein B, a potential antifungal agent from the Brazilian Cerrado plant Eugenia uniflora.

Zambuzzi-Carvalho PF, Tomazett PK, Santos SC, Ferri PH, Borges CL, Martins WS, de Almeida Soares CM, Pereira M - BMC Microbiol. (2013)

Functional classification of differentially expressed genes in biological functional groups. (A) Down- and up-regulated genes obtained from cDNAs synthesized from RNAs of Paracoccidioides cells exposure to OenB for 90 min and (B) 180 min. Functional classification was based on Blast X homology of each EST against the GenBank non-redundant database and the nucleotide database from the Paracoccidioides structural genome at a significant homology cut-off ≤ 10-5 and a MIPS functional annotation scheme. Each functional class is represented as a segment and expressed as the total number of ESTs in each library.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3852496&req=5

Figure 1: Functional classification of differentially expressed genes in biological functional groups. (A) Down- and up-regulated genes obtained from cDNAs synthesized from RNAs of Paracoccidioides cells exposure to OenB for 90 min and (B) 180 min. Functional classification was based on Blast X homology of each EST against the GenBank non-redundant database and the nucleotide database from the Paracoccidioides structural genome at a significant homology cut-off ≤ 10-5 and a MIPS functional annotation scheme. Each functional class is represented as a segment and expressed as the total number of ESTs in each library.
Mentions: The distribution of differentially expressed genes in the different functional categories and biological processes was evaluated (Table 1; Figure 1). The analysis demonstrated that most of the genes that were down-regulated after 90 min participate in cellular processes related to transport, followed by signal transducers and those involved in transcription and metabolism. On the other hand, the up-regulated genes were mostly found to be functionally unclassified proteins, followed by unclassified proteins, and those involved in cell cycle and DNA processing and metabolism. After 180 min treatment, most of the down-regulated genes were also involved in signal transduction, metabolism and cellular transport. The up-regulated genes were unclassified proteins or those involved in transcription and morphology determination. The percentage of occurrence of each gene in the libraries was indicated in the Additional file 2: Table S1.

Bottom Line: Infection experiments in macrophages corroborated the in vitro results.Several other genes, such as those involved in a variety of important cellular processes (i.e., membrane maintenance, stress and virulence) were found to be up-regulated in response to OenB treatment.The exposure of Paracoccidioides to OenB resulted in a complex altered gene expression profile.

View Article: PubMed Central - HTML - PubMed

Affiliation: Departamento de Bioquímica e Biologia Molecular, Laboratório de Biologia Molecular, Instituto de Ciências Biológicas, ICBII, Campus II, Universidade Federal de Goiás, C,P, 131, 74001-970 Goiânia, GO, Brazil. maristelaufg@gmail.com.

ABSTRACT

Background: The compound oenothein B (OenB), which is isolated from the leaves of Eugenia uniflora, a Brazilian Cerrado plant, interferes with Paracoccidioides yeast cell morphology and inhibits 1,3-β-D-glucan synthase (PbFKS1) transcript accumulation, which is involved in cell wall synthesis. In this work we examined the gene expression changes in Paracoccidioides yeast cells following OenB treatment in order to investigate the adaptive cellular responses to drug stress.

Results: We constructed differential gene expression libraries using Representational Difference Analysis (RDA) of Paracoccidioides yeast cells treated with OenB for 90 and 180 min. Treatment for 90 min resulted in the identification of 463 up-regulated expressed sequences tags (ESTs) and 104 down-regulated ESTs. For the 180 min treatment 301 up-regulated ESTs and 143 down-regulated were identified. Genes involved in the cell wall biosynthesis, such as GLN1, KRE6 and FKS1, were found to be regulated by OenB. Infection experiments in macrophages corroborated the in vitro results. Fluorescence microscopy showed increased levels of chitin in cells treated with OenB. The carbohydrate polymer content of the cell wall of the fungus was also evaluated, and the results corroborated with the transcriptional data. Several other genes, such as those involved in a variety of important cellular processes (i.e., membrane maintenance, stress and virulence) were found to be up-regulated in response to OenB treatment.

Conclusions: The exposure of Paracoccidioides to OenB resulted in a complex altered gene expression profile. Some of the changes may represent specific adaptive responses to this compound in this important pathogenic fungus.

Show MeSH
Related in: MedlinePlus