Partial characterization of the transcription factor rlmA and its subcellular localization in the human opportunistic fungus Aspergillus fumigatus

Abstract

Aspergillus fumigatus is a ubiquitous filamentous fungus with an important role in nature in the recycling of carbon and nitrogen. Nowaday is the most prevalent airborne fungal pathogen, and causes severe invasive infections (aspergillosis), however, it is not fully understood why this specie has become one of the most prevalent opportunistic pathogens. To adapt to a changing environment and to maintain a state of homeostasis, microorganisms have developed a sophisticated regulatory system that controls the cellular response upon external signal through transducing proteins for an effective cell response. In particular, the cell wall integrity (CWI) is responsible for cell wall remodelind and reinforcement upon cell wall stress in fungus. CWI works on fungal cell connecting cell wall damage to a signaling pathway mediated axis by protein kinase C (PKC) and MAPK phosphorylation. This circuit culminates in the activation of the RLM1 transcription factor in Saccharomyces cerevisiae. RLM1 plays a key role in the regulation of genes involved in biosynthesis and cell wall reinforcement in S. cerevisiae. In A. fumigatus through similarity analysis, it was possible to identify the homologue of RLM1 named here as rlmA (Afu3g08520). However, the function of rlmA as an effector of CWI has not been elucidated yet. Therefore, the main objective of this project is the study the function and the importance of rlmA in maintaining the integrity of the cell wall in this organism. The cell wall is a rigid and essential structure for the survival of the fungal cells so, the understanding of how rlmA relates to its maintenance in A. fumigatus may be useful for the development of new antifungal drugs. Recently, our laboratory isolated null mutant DrlmA which will be employed in this study for the phenotypic analyses proposed here. In addition two strains were constructed: DrlmA:: rlmA (complementing strain) which will be used in the experiments of phenotypic characterization and strain rlmA::gfp aiming: (i) to study the subcellular localization of the protein RlmA in the absence and presence of cell wall stress and (ii) its abundance at the protein level by western blot under the same conditions.