Histones with an unconventional DNA-binding mode in vitro are major chromatin constituents in the bacterium Bdellovibrio bacteriovorus.

Histone proteins bind DNA and organize the genomes of eukaryotes and most archaea, whereas bacteria rely on different nucleoid-associated proteins. Homology searches have detected putative histone-fold domains in a few bacteria, but whether these function like archaeal/eukaryotic histones is unknown. Here we report that histones are major chromatin components in the bacteria Bdellovibrio bacteriovorus and Leptospira interrogans. Patterns of sequence evolution suggest important roles for histones in additional bacterial clades. Crystal structures (<2.0 Å) of the B. bacteriovorus histone (Bd0055) dimer and the histone-DNA complex confirm conserved histone-fold topology but indicate a distinct DNA-binding mode. Unlike known histones in eukaryotes, archaea and viruses, Bd0055 binds DNA end-on, forming a sheath of dimers encasing straight DNA rather than wrapping DNA around their outer surface. Our results demonstrate that histones are present across the tree of life and highlight potential evolutionary innovation in how they associate with DNA.

Histone variants in archaea - An undiscovered country.

Exchanging core histones in the nucleosome for paralogous variants can have important functional ramifications. Many of these variants, and their physiological roles, have been characterized in exquisite detail in model eukaryotes, including humans. In comparison, our knowledge of histone biology in archaea remains rudimentary. This is true in particular for our knowledge of histone variants. Many archaea encode several histone genes that differ in sequence, but do these paralogs make distinct, adaptive contributions to genome organization and regulation in a manner comparable to eukaryotes? Below, we review what we know about histone variants in archaea at the level of structure, regulation, and evolution. In all areas, our knowledge pales when compared to the wealth of insight that has been gathered for eukaryotes. Recent findings, however, provide tantalizing glimpses into a rich and largely undiscovered country that is at times familiar and eukaryote-like and at times strange and uniquely archaeal. We sketch a preliminary roadmap for further exploration of this country; an undertaking that may ultimately shed light not only on chromatin biology in archaea but also on the origin of histone-based chromatin in eukaryotes.

The structural basis of Cdc7-Dbf4 kinase dependent targeting and phosphorylation of the MCM2-7 double hexamer.

The controlled assembly of replication forks is critical for genome stability. The Dbf4-dependent Cdc7 kinase (DDK) initiates replisome assembly by phosphorylating the MCM2-7 replicative helicase at the N-terminal tails of Mcm2, Mcm4 and Mcm6. At present, it remains poorly understood how DDK docks onto the helicase and how the kinase targets distal Mcm subunits for phosphorylation. Using cryo-electron microscopy and biochemical analysis we discovered that an interaction between the HBRCT domain of Dbf4 with Mcm2 serves as an anchoring point, which supports binding of DDK across the MCM2-7 double-hexamer interface and phosphorylation of Mcm4 on the opposite hexamer. Moreover, a rotation of DDK along its anchoring point allows phosphorylation of Mcm2 and Mcm6. In summary, our work provides fundamental insights into DDK structure, control and selective activation of the MCM2-7 helicase during DNA replication. Importantly, these insights can be exploited for development of novel DDK inhibitors.

Deep Conservation of Histone Variants in Thermococcales Archaea.

Histones are ubiquitous in eukaryotes where they assemble into nucleosomes, binding and wrapping DNA to form chromatin. One process to modify chromatin and regulate DNA accessibility is the replacement of histones in the nucleosome with paralogous variants. Histones are also present in archaea but whether and how histone variants contribute to the generation of different physiologically relevant chromatin states in these organisms remains largely unknown. Conservation of paralogs with distinct properties can provide prima facie evidence for defined functional roles. We recently revealed deep conservation of histone paralogs with different properties in the Methanobacteriales, but little is known experimentally about these histones. In contrast, the two histones of the model archaeon Thermococcus kodakarensis, HTkA and HTkB, have been examined in some depth, both in vitro and in vivo. HTkA and HTkB exhibit distinct DNA-binding behaviors and elicit unique transcriptional responses when deleted. Here, we consider the evolution of HTkA/B and their orthologs across the order Thermococcales. We find histones with signature HTkA- and HTkB-like properties to be present in almost all Thermococcales genomes. Phylogenetic analysis indicates the presence of one HTkA- and one HTkB-like histone in the ancestor of Thermococcales and long-term maintenance of these two paralogs throughout Thermococcales diversification. Our results support the notion that archaea and eukaryotes have convergently evolved histone variants that carry out distinct adaptive functions. Intriguingly, we also detect more highly diverged histone-fold proteins, related to those found in some bacteria, in several Thermococcales genomes. The functions of these bacteria-type histones remain unknown, but structural modeling suggests that they can form heterodimers with HTkA/B-like histones.

Histone variants in archaea and the evolution of combinatorial chromatin complexity.

Nucleosomes in eukaryotes act as platforms for the dynamic integration of epigenetic information. Posttranslational modifications are reversibly added or removed and core histones exchanged for paralogous variants, in concert with changing demands on transcription and genome accessibility. Histones are also common in archaea. Their role in genome regulation, however, and the capacity of individual paralogs to assemble into histone-DNA complexes with distinct properties remain poorly understood. Here, we combine structural modeling with phylogenetic analysis to shed light on archaeal histone paralogs, their evolutionary history, and capacity to generate combinatorial chromatin states through hetero-oligomeric assembly. Focusing on the human commensal Methanosphaera stadtmanae as a model archaeal system, we show that the heteromeric complexes that can be assembled from its seven histone paralogs vary substantially in DNA binding affinity and tetramer stability. Using molecular dynamics simulations, we go on to identify unique paralogs in M. stadtmanae and Methanobrevibacter smithii that are characterized by unstable interfaces between dimers. We propose that these paralogs act as capstones that prevent stable tetramer formation and extension into longer oligomers characteristic of model archaeal histones. Importantly, we provide evidence from phylogeny and genome architecture that these capstones, as well as other paralogs in the Methanobacteriales, have been maintained for hundreds of millions of years following ancient duplication events. Taken together, our findings indicate that at least some archaeal histone paralogs have evolved to play distinct and conserved functional roles, reminiscent of eukaryotic histone variants. We conclude that combinatorially complex histone-based chromatin is not restricted to eukaryotes and likely predates their emergence.

Chromatinization of Escherichia coli with archaeal histones.

Nucleosomes restrict DNA accessibility throughout eukaryotic genomes, with repercussions for replication, transcription, and other DNA-templated processes. How this globally restrictive organization emerged during evolution remains poorly understood. Here, to better understand the challenges associated with establishing globally restrictive chromatin, we express histones in a naive system that has not evolved to deal with nucleosomal structures: Escherichia coli. We find that histone proteins from the archaeon Methanothermus fervidus assemble on the E. coli chromosome in vivo and protect DNA from micrococcal nuclease digestion, allowing us to map binding footprints genome-wide. We show that higher nucleosome occupancy at promoters is associated with lower transcript levels, consistent with local repressive effects. Surprisingly, however, this sudden enforced chromatinization has only mild repercussions for growth unless cells experience topological stress. Our results suggest that histones can become established as ubiquitous chromatin proteins without interfering critically with key DNA-templated processes.