Geometric properties of Deinococcus radiodurans cells and nucleoids, and the spatial distribution of proteins with respect to cells and nucleoids, reflect effects of radiation exposure or the deletion of key genome-associated proteins

Unusually, the extremophile bacterium Deinococcus radiodurans constituitively maintains a highly-organized and condensed nucleoid; this may contribute to D. radiodurans’ extraordinarily high tolerance of ionizing radiation (IR) and other stresses. What role different proteins play in regulating the compaction of DNA in D. radiodurans is unknown. Thus, there is a need for a screening tool to identify proteins involved in nuclear compaction and stress response. Previous studies of the D. radiodurans nucleoid relied on manual annotation and largely qualitative metrics. Here, we introduce a high-throughput approach to quantifying the geometric properties of cells and nucleoids, using confocal microscopy, digital reconstructions of bacterial cells, and machine learning. We measure pronounced changes in D. radiodurans cells and nucleoids following either IR or genetic knocking-out of candidate proteins. We find that the nucleoid becomes a more-compact and more-spherical shape as IR increases, and that cells and nucleoids adopt morphological extremes upon deletin of protein-encoding genes. We also find that different protein types have different spatial distributions within the cell and the nucleoid. These are also altered by IR stress and by changes in protein expression. Thus, here we demonstrate an adaptable, quantitative methodology for analyzing D. radiodurans’ robust response to stressors and for screening candidate proteins for their role in the organization of the bacterial nucleoid.