Publicação
The impact of carbon source in Candida albicans virulence: participation of RLM1 in pathogen host interaction
| Resumo: | The survival of all microbes depends upon their ability to respond to environmental challenges. To establish infection, pathogens such as Candida albicans must support effective stress responses to counter host defenses while adapting to dynamic changes in nutrient status within host niches. Studies of C. albicans stress adaptation have generally been performed on glucose-grown cells, leaving the effects of alternative carbon sources upon stress resistance largely unexplored as well alterations in its virulence. In a previous work both copies of the RLM1 gene of C. albicans were deleted and phenotypic analysis of Δrlm1/Δrlm1 mutants showed typical cell wall weakening phenotypes, such as hypersensitivity to Congo Red, Calcofluor White and caspofungin, showing its involvement in the cell wall remodelling. To understand the role of RLM1 under the influence of different carbon source, phenotypic characterization against cell wall damaging stress agents, modulation of virulence factors as well as their involvement in host-pathogen interaction was preformed in the present work. Candida albicans Δrlm1/Δrlm1 mutant displayed phenotypes associated to cell wall deficiency such as, hypersensitivity to Congo red, caspofungin in glucose- and lactate- grown cells. However the RLM1 mutants were slight more resistant to Congo Red when grown in lactate. On the other hand lactate-grown cells were not able to growth al all in the presence of SDS and presented sensitivity to caffeine, in comparison with glucose-grown cells. The increased transcription of genes already reported to be involved in cell adhesion correlated well with adhesion and biofilm assays, in which RLM1 mutant presented greater biofilm formation than WT in both growth condition. However cell adapted to lactate adhered more and biofilm formation was more pronounced. The host-pathogen interaction was accessed by co-incubation with murine macrophages-like cell line (J774). In general, lactate-grown cells were less efficiently killed in comparison to glucose-grown cells. However the mutant presented distinct behaviors, they were more resistant when adapted to glucose than to lactate. The TNF-α and IL-10 were lower in response to Δrlm1/Δrlm1 mutant and the cellular toxicity, measured as extracellular lactate dehydrogenase activity, was significantly lower in comparison with the WT and complemented strains in glucose-grown cells. The effect observed before was reverted when C. albicans cells were grown on lactate. In conclusion, C. albicans cells adapted to different carbon sources behave differently, particularly in the interaction with macrophages, in which the RLM1 mutation plays a decisive effect since it affects the cell wall integrity. |
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| Autores principais: | Pacheco, João Miguel Oliveira |
| Assunto: | Ciências Naturais::Ciências Biológicas |
| Ano: | 2015 |
| País: | Portugal |
| Tipo de documento: | dissertação de mestrado |
| Tipo de acesso: | acesso aberto |
| Instituição associada: | Universidade do Minho |
| Idioma: | inglês |
| Origem: | RepositóriUM - Universidade do Minho |
| Resumo: | The survival of all microbes depends upon their ability to respond to environmental challenges. To establish infection, pathogens such as Candida albicans must support effective stress responses to counter host defenses while adapting to dynamic changes in nutrient status within host niches. Studies of C. albicans stress adaptation have generally been performed on glucose-grown cells, leaving the effects of alternative carbon sources upon stress resistance largely unexplored as well alterations in its virulence. In a previous work both copies of the RLM1 gene of C. albicans were deleted and phenotypic analysis of Δrlm1/Δrlm1 mutants showed typical cell wall weakening phenotypes, such as hypersensitivity to Congo Red, Calcofluor White and caspofungin, showing its involvement in the cell wall remodelling. To understand the role of RLM1 under the influence of different carbon source, phenotypic characterization against cell wall damaging stress agents, modulation of virulence factors as well as their involvement in host-pathogen interaction was preformed in the present work. Candida albicans Δrlm1/Δrlm1 mutant displayed phenotypes associated to cell wall deficiency such as, hypersensitivity to Congo red, caspofungin in glucose- and lactate- grown cells. However the RLM1 mutants were slight more resistant to Congo Red when grown in lactate. On the other hand lactate-grown cells were not able to growth al all in the presence of SDS and presented sensitivity to caffeine, in comparison with glucose-grown cells. The increased transcription of genes already reported to be involved in cell adhesion correlated well with adhesion and biofilm assays, in which RLM1 mutant presented greater biofilm formation than WT in both growth condition. However cell adapted to lactate adhered more and biofilm formation was more pronounced. The host-pathogen interaction was accessed by co-incubation with murine macrophages-like cell line (J774). In general, lactate-grown cells were less efficiently killed in comparison to glucose-grown cells. However the mutant presented distinct behaviors, they were more resistant when adapted to glucose than to lactate. The TNF-α and IL-10 were lower in response to Δrlm1/Δrlm1 mutant and the cellular toxicity, measured as extracellular lactate dehydrogenase activity, was significantly lower in comparison with the WT and complemented strains in glucose-grown cells. The effect observed before was reverted when C. albicans cells were grown on lactate. In conclusion, C. albicans cells adapted to different carbon sources behave differently, particularly in the interaction with macrophages, in which the RLM1 mutation plays a decisive effect since it affects the cell wall integrity. |
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