Direct observation of a crescent-shape chromosome in expanded Bacillus subtilis cells
Miloš Tišma (BN/Cees Dekker Lab)
Florian Patrick Bock (University of Lausanne)
Jacob Kerssemakers (BN/Cees Dekker Lab)
Hammam Antar (University of Lausanne)
Aleksandre Japaridze (TU Delft - Dynamics of Micro and Nano Systems)
Stephan Gruber (University of Lausanne)
Cees Dekker (BN/Cees Dekker Lab)
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Abstract
Bacterial chromosomes are folded into tightly regulated three-dimensional structures to ensure proper transcription, replication, and segregation of the genetic information. Direct visualization of chromosomal shape within bacterial cells is hampered by cell-wall confinement and the optical diffraction limit. Here, we combine cell-shape manipulation strategies, high-resolution fluorescence microscopy techniques, and genetic engineering to visualize the shape of unconfined bacterial chromosome in real-time in live Bacillus subtilis cells that are expanded in volume. We show that the chromosomes predominantly exhibit crescent shapes with a non-uniform DNA density that is increased near the origin of replication (oriC). Additionally, we localized ParB and BsSMC proteins – the key drivers of chromosomal organization – along the contour of the crescent chromosome, showing the highest density near oriC. Opening of the BsSMC ring complex disrupted the crescent chromosome shape and instead yielded a torus shape. These findings help to understand the threedimensional organization of the chromosome and the main protein complexes that underlie its structure.
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