Project description:One of the most complex prokaryotic organelles are magnetosomes, which are formed by magnetotactic bacteria as sensors for navigation in the Earth’s magnetic field. In the alphaproteobacterium Magnetospirillum gryphiswaldense magnetosomes consist of chains of magnetite crystals (Fe3O4) that under suboxic conditions are biomineralized within membrane vesicles. To form such an intricate structure, the transcription of >30 specific structural genes clustered within the genomic magnetosome island (MAI) has to be coordinated with the expression of an as-yet unknown number of auxiliary genes encoding several generic metabolic functions. However, their global regulation and transcriptional organization in response to anoxic conditions most favorable for magnetite biomineralization are still unclear. Here, we compared transcriptional profiles of anaerobically grown magnetosome forming cells with those in which magnetosome biosynthesis has been suppressed by aerobic condition. Using whole transcriptome shotgun sequencing, we found that transcription of about 300 of the >4300 genes was significantly enhanced during magnetosome formation. The about 40 top upregulated genes are directly or indirectly linked to aerobic and anaerobic respiration (denitrification) or unknown functions. mam and mms gene clusters specifically controlling magnetosome biosynthesis were highly transcribed, but constitutively expressed irrespective of the growth condition. By Cappable-sequencing, we show that the transcriptional complexity of both the MAI and the entire genome decreased under anaerobic conditions optimal for magnetosome formation. In addition, predominant promoter structures were highly similar to sigma factor σ70 dependent promoters in other Alphaproteobacteria. Our transcriptome-wide analysis revealed that magnetite biomineralization relies on a complex interplay between generic metabolic processes such as aerobic and anaerobic respiration, cellular redox control, and the biosynthesis of specific magnetosome structures. In addition, we provide insights into global regulatory features that have remained uncharacterized in the widely studied model organism M. gryphiswaldense, including a comprehensive dataset of newly annotated transcription start sites and genome-wide operon detection as a community resource.
Project description:Magnetosomes are complex membrane organelles synthesized by magnetotactic bacteria (MTB) for navigation in the magnetic field. Their formation is tightly controlled by ˃30 specific magnetosome genes arranged in operons. The transcriptional organization of these operons in the model MTB Magnetospirillum gryphiswaldense MSR-1 has been long viewed to be simple, having each a convenient promoter that enable transcription of the operon as a single transcriptional unit. Here, by applying Cappable-seq and whole transcriptome shotgun RNA sequencing, we revealed multiple additional transcription starting sites (TSS) within the magnetosome operons mamABop, mms6op and mamXYop. Using experimental validation by bioluminescence reporter, we demonstrated that most of the new TSS are generated by biologically meaningful promoters. Furthermore, the knockout of the primary promoters in mamABop and mms6op resulted only in mild impairments of the magnetosome formation, suggesting that additional transcripts are indeed generated within these operons. Besides, we identified a strong promoter in the intergenic region within mamXYop, which transcript likely represents a non-coding RNA important for proper expression of the genes in this operon. The promoter sequences from MSR-1 are conservative in most magnetotactic Magnetospirillum spp., but not in other MTB, suggesting the functional conservation in magnetospirilla but independent evolution of the transcription circuits within the magnetosome operons in different phylogenetic groups. Taken together, our data suggest much more complex transcriptional architecture of the magnetosome operons in MSR-1 than deemed before and contribute to unraveling the fundamentals of magnetosome biosynthesis at transcriptional level.
Project description:3 samples of R1, R2 and R3 bone marrow monocytes were compared from 3 biological replicates in 3 separate experiments. R1, R2 and R3 were sorted from triplicate experiments from pools of mice