The effects of polydisperse crowders on the compaction of the Escherichia coli nucleoid

Abstract

DNA binding proteins, supercoiling, macromolecular crowders, and transient DNA attachments to the cell membrane have all been implicated in the organization of the bacterial chromosome. However, it is unclear what role these factors play in compacting the bacterial DNA into a distinct organelle-like entity, the nucleoid. By analyzing the effects of osmotic shock and mechanical squeezing on Escherichia coli, we show that macromolecular crowders play a dominant role in the compaction of the DNA into the nucleoid. We find that a 30% increase in the crowder concentration from physiological levels leads to a 3-fold decrease in the nucleoid's volume. The compaction is anisotropic, being higher along the long axes of the cell at low crowding levels. At higher crowding levels the compression becomes isotropic, implying that E. coli nucleoids lack a well-defined backbone. We furthermore show that the compressibility of the nucleoid is not significantly affected by cell growth rates and by prior treatment with rifampicin. The latter results point out that in addition to poly-ribosomes, soluble cytoplasmic proteins have a significant contribution in determining the size of the nucleoid.

Interconnecting solvent quality, transcription, and chromosome folding in Escherichia coli

“…Among these factors, we find RNAs to be attractive candidates. Indeed, treatment of E. coli with rifampicin (Rif), which blocks transcription initiation and leads to mRNA depletion due to the short lifetime of most bacterial mRNAs (Bernstein et al, 2002;Chen et al, 2015;Selinger et al, 2003), is known to result in nucleoid expansion (Bakshi et al, 2012(Bakshi et al, , 2014Cabrera et al, 2009;Cabrera and Jin, 2003;Dworsky and Schaechter, 1973;Pettijohn and Hecht, 1974;Sun and Margolin, 2004;Yang et al, 2020). Conversely, treatment with chloramphenicol (Cpl), which inhibits ribosome translocation and stabilizes mRNAs (Lopez et al, 1998;Pato et al, 1973;Schneider et al, 1978), is associated with nucleoid compaction ll 3632 Cell 184, 3626-3642, July 8, 2021 Article (Bakshi et al, 2012(Bakshi et al, , 2014Cabrera et al, 2009;van Helvoort et al, 1996;Zimmerman, 2002).…”

“…Transertion, which is the coupling between transcription, translation, and insertion of nascent proteins into the cytoplasmic membrane, is thought to provide an expanding force on the nucleoid (Bakshi et al, 2014;Woldringh, 2002;Woldringh et al, 1995). Conversely, macromolecular crowding is frequently proposed to generate a compacting force through steric (size-dependent) effects (Bakshi et al, 2014;Cunha et al, 2001;de Vries, 2010;Jeon et al, 2016;Jun, 2015;Kim et al, 2015;Mondal et al, 2011;Odijk, 1998;Pelletier et al, 2012;Shendruk et al, 2015;Wegner et al, 2016;Wu et al, 2019;Yang et al, 2020;Yoshikawa et al, 2010;Zhang et al, 2009;Zimmerman, 1993;Zimmerman and Minton, 1993). The idea is that entropic forces drive depletion of crowders (e.g., proteins and ribosomes) from the nucleoid to reduce the excluded volume.…”

“…The imbalance in crowder concentration between the nucleoid region and the rest of the cytoplasm results in an effec-tive osmotic pressure that pushes DNA segments closer to each other. The magnitude of the entropic (depletion) forces, which remains to be determined inside cells, will depend on the abundance, size, and polydispersity of the crowders (Kim et al, 2015;Yang et al, 2020). While steric interactions are primarily emphasized in current models of chromosome compaction, DNA and other cytoplasmic components are not chemically inert, and non-steric interactions may also contribute to chromosome folding in vivo (Joyeux, 2015).…”

Transient Membrane-Linked FtsZ Assemblies Precede Z-Ring Formation in Escherichia coli

“…Power of the Hg lamp prior to each measurement (P GFP and P cells , respectively) was measured by a Thorlabs PM121D power meter. The final formula used in calculations of cellular Here R cell is the cytosolic radius of the cells estimated to be 0.32 mm in these growth conditions for this strain [49]. Additionally, a correction factor, corr, was introduced to the formula to account for cells that were not forming a continuous tube-like volume but consisted of individual cylindrical cells with hemispherical endcaps.…”

Transient Membrane-Linked FtsZ Assemblies Precede Z-Ring Formation in <i>Escherichia Coli</i>

“…Power of the Hg lamp prior to each measurement (P GFP and P cells , respectively) was measured by a Thorlabs PM121D power meter. The final formula used in calculations of cellular FtsZ was: Here R cell is the cytosolic radius of the cells estimated to be 0.32 μm in these growth conditions for this strain [49]. Additionally, a correction factor, corr, was introduced to the formula to account for cells that were not forming a continuous tube-like volume but consisted of individual cylindrical cells with hemispherical endcaps.…”