Nucleoid-Associated Proteins Undergo Liquid-Liquid Phase Separation with DNA into Multiphasic Condensates Resembling Bacterial Nucleoids

Abstract

Liquid-liquid phase separation offers unique spatiotemporal control over myriad complex intracellular biochemical processes through compartmentalization of biomolecules into highly dynamic, liquid-like condensates known as membrane-less organelles. The bacterial nucleoid is thought to be one such phase-separated condensate; however, its formation, regulation, and biophysical characteristics are poorly understood. Our super-resolution imaging data suggests that nucleoids are dynamic assemblages of sub-micron-sized liquid-like droplets. We demonstrate that non-sequence-specific Nucleoid-Associated Proteins (NAPs) such as HU-A, HU-B and Dps, accrete nucleic acids and spontaneously condense with them into liquid-like, multicomponent, multiphasic, heterotypic phase-separated droplets. Upon mixing of HU-B, DNA and Dps, HU-B-enriched droplets are seen to contain demixed Dps-enriched droplets. Our findings indicate scope for the possible existence of multiphasic liquid-like compartments within nucleoids, providing insights into bacterial growth phase-dependent variations in the levels of different NAPs.

Morphological Diversity of Dps Complex with Genomic DNA

“…HU and Dps together form complex multiphasic droplets with DNA, without undergoing complete mixing, with evidence of the formation of domains of differential HU or Dps content. This ability of HU condensates to accommodate Dps-rich condensates could prevent the premature crystallization of Dps during stress [44]. HU and IHF are close homologues with similar structures and functions.…”