Batista Marcelo Bueno, Wassem Roseli, Pedrosa Fábio de Oliveira, de Souza Emanuel Maltempi, Dixon Ray, Monteiro Rose Adele
Department of Biochemistry and Molecular Biology, Universidade Federal do Paraná, P.O. Box 19046, Curitiba, PR, 81531-990, Brazil.
Department of Genetics, Universidade Federal do Paraná, P.O. Box 19071, Curitiba, PR, 81531-990, Brazil.
BMC Microbiol. 2015 May 7;15:95. doi: 10.1186/s12866-015-0432-6.
Orthologous proteins of the Crp/Fnr family have been previously implicated in controlling expression and/or activity of the NifA transcriptional activator in some diazotrophs. This study aimed to address the role of three Fnr-like proteins from H. seropedicae SmR1 in controlling NifA activity and consequent NifA-mediated transcription activation.
The activity of NifA-dependent transcriptional fusions (nifA::lacZ and nifB::lacZ) was analysed in a series of H. seropedicae fnr deletion mutant backgrounds. We found that combined deletions in both the fnr1 and fnr3 genes lead to higher expression of both the nifA and nifB genes and also an increased level of nifH transcripts. Expression profiles of nifB under different oxygen concentrations, together with oxygen consumption measurements suggest that the triple fnr mutant has higher respiratory activity when compared to the wild type, which we believe to be responsible for greater stability of the oxygen sensitive NifA protein. This conclusion was further substantiated by measuring the levels of NifA protein and its activity in fnr deletion strains in comparison with the wild-type.
Fnr proteins are indirectly involved in controlling the activity of NifA in H. seropedicae, probably as a consequence of their influence on respiratory activity in relation to oxygen availability. Additionally we can suggest that there is some redundancy in the physiological function of the three Fnr paralogs in this organism, since altered respiration and effects on NifA activity are only observed in deletion strains lacking both fnr1 and fnr3.
此前已表明，Crp/Fnr家族的直系同源蛋白在某些固氮菌中参与控制NifA转录激活因子的表达和/或活性。本研究旨在探讨来自巴西固氮螺菌SmR1的三种Fnr样蛋白在控制NifA活性以及随后的NifA介导的转录激活中的作用。
在一系列巴西固氮螺菌fnr缺失突变体背景中分析了NifA依赖性转录融合体（nifA::lacZ和nifB::lacZ）的活性。我们发现，fnr1和fnr3基因的联合缺失导致nifA和nifB基因的表达更高，同时nifH转录本水平也增加。nifB在不同氧浓度下的表达谱以及耗氧量测量表明，与野生型相比，三重fnr突变体具有更高的呼吸活性，我们认为这是氧敏感的NifA蛋白更稳定的原因。通过测量fnr缺失菌株与野生型相比的NifA蛋白水平及其活性，进一步证实了这一结论。
Fnr蛋白间接参与控制巴西固氮螺菌中NifA的活性，这可能是由于它们对与氧可用性相关的呼吸活性的影响。此外，我们可以认为，该生物体中三种Fnr旁系同源物的生理功能存在一些冗余，因为只有在缺乏fnr1和fnr3的缺失菌株中才观察到呼吸改变和对NifA活性的影响。
