The landscape of the histone-organized chromatin of Bdellovibrionota bacteria
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ABSTRACT: Histone proteins have traditionally been thought to be restricted to eukaryotes and most archaea, with eukaryotic nucleosomal histones deriving from their archaeal ancestors. In contrast, bacteria lack histones as a rule. However, in recent years histone proteins have been identified in a few bacterial clades, in particular the phylum Bdellovibrionota, and these histones have been proposed to exhibit a range of divergent features compared to histones in archaea and eukaryotes. However, no experimental functional genomic studies of the properties of Bdellovibrionota chromatin have been carried out. In this work, we map the landscape of chromatin accessibility, active transcription and three-dimensional genome organization in a member of Bdellovibrionota (a Bacteriovorax strain). We find that Bacteriovorax chromatin is characterized by preferential accessibility around promoter regions, similar to what is observed in eukaryotes with compact genomes such as yeast, and also to some archaea. As in eukaryotes, chromatin accessibility positively correlates with gene expression. Mapping active transcription through single-strand DNA (ssDNA) profiling revealed that Bacteriovorax promoters exhibit very strong polymerase pausing, unlike in yeast, but similar to the state of mammalian and fly promoters. Finally, the Bacteriovorax genome exists in a three-dimensional (3D) conformation analogous to that of other bacteria without histones, organized by the parABS system and along the axis defined by replication origin and termination regions. These results provide a foundation for understanding the chromatin biology of the unique Bdellovibrionota bacteria and the deep evolution of chromatin organization across the tree of life.
Project description:Histones are a principal constituent of chromatin in eukaryotes and fundamental to our understanding of eukaryotic gene regulation. In archaea, histones are phylogenetically widespread but not universal: several archaeal lineages have independently lost histone genes. What prompted or facilitated these losses and how archaea without histones organize their chromatin remains largely unknown. Here, we use micrococcal nuclease digestion of native and reconstituted chromatin to elucidate primary chromatin architecture in an archaeon without histones, the acido-thermophilic archaeon Thermoplasma acidophilum. We confirm and extend prior results showing that T. acidophilum harbours a HU family protein, HTa, that protects part of the genome from MNase digestion. Charting HTa-based chromatin architecture in vitro, in vivo and in an HTa-expressing E. coli strain, we present evidence that HTa is an archaeal histone analog. HTa-protected fragments are GC-rich, display histone-like mono- and dinucleotide patterns around a conspicuous dyad, exhibit relatively invariant positioning throughout the growth cycle, and show archaeal histone-like oligomerization behaviour. Our results suggest that HTa, a DNA-binding protein of bacterial origin, has converged onto an architectural role filled by histones in other archaea.
Project description:Histones are the primary building blocks of chromatin in eukaryotes and many archaea. Bacteria are thought to rely on an orthogonal set of proteins to organize their chromosomes. Several bacterial genomes do, however, encode proteins with putative histone fold domains. Whether these proteins adopt a bona fide histone fold, assemble into higher order complexes that bind DNA, and play a central role in bacterial nucleoid physiology is not known. Here, we demonstrate that histones are major and essential building blocks of chromatin in the predatory bacterium Bdellovibrio bacteriovorus and the human pathogen Leptospira interrogans and likely important in several other bacterial clades. We determine the crystal structure of the B. bacteriovorus histone (Bd0055) dimer at 1.8Å resolution to reveal that histone fold topology, handshake dimer conformation, and the RD clamp motif are conserved between bacteria, archaea, and eukaryotes. However, ostensibly minor differences, including a shorter α2 helix, a less structured α3 helix, and a more acidic surface on one side of the dimer lead to a radically divergent DNA binding mode: instead of wrapping around the outer surface of a multi-subunit histone complex, DNA forms straight fibers, encased by a sheath of tightly packed Bd0055 dimers. Our results demonstrate that bacterial histones have evolved an atypical mode of DNA binding to become integral components of chromatin in distant parts of the bacterial kingdom
Project description:Archaea, together with Bacteria, represent the two main divisions of life on Earth, with many of the defining characteristics of the more complex eukaryotes tracing their origin to evolutionary innovations first made in their archaeal ancestors. One of the most notable such features is nucleosomal chromatin, although archaeal histones and chromatin differ significantly from those of eukaryotes. Despite increased interest in archaeal histones in recent years, the properties of archaeal chromatin have been little studied using genomic tools. Here, we adapt the ATAC-seq assay to the archaeal context and use it to map the accessible landscape of the genome of the euryarchaeote Haloferax volcanii. We integrate the resulting datasets with genome-wide maps of active transcription and single-stranded DNA (ssDNA), and find that while H. volcanii promoters exist in a preferentially accessible state, modulation of transcriptional activity is not associated with changes in promoter accessibility, unlike the typical situation in eukaryotes. Applying orthogonal single-molecule footprinting methods, we quantify the absolute levels of physical protection of H. volcanii, and find that archaeal nucleosomal chromatin is at its baseline comparably to slightly more open than that of eukaryotes. We also evaluate the degree of coordination of transcription within archaeal operons and make the unexpected observation that some CRISPR arrays are associated with highly prevalent ssDNA structures. These results provide a foundation for the future functional studies of archaeal chromatin.
Project description:Histones and associated chromatin proteins have essential functions in eukaryotic genome organization and regulation. Despite this fundamental role in eukaryotic cell biology, we lack a phylogenetically-comprehensive understanding of chromatin evolution. Here, we combine comparative proteomics and genomics analysis of chromatin in eukaryotes and archaea. Proteomics uncovers the existence of histone post-translational modifications in Archaea. However, archaeal histone modifications are scarce, in contrast with the highly conserved and abundant marks we identify across eukaryotes. Phylogenetic analysis reveals that chromatin-associated catalytic functions (e.g., methyltransferases) have pre-eukaryotic origins, whereas histone mark readers and chaperones are eukaryotic innovations. We show that further chromatin evolution is characterized by expansion of readers, including capture by transposable elements and viruses. Overall, our study infers detailed evolutionary history of eukaryotic chromatin: from its archaeal roots, through the emergence of nucleosome-based regulation in the eukaryotic ancestor, to the diversification of chromatin regulators and their hijacking by genomic parasites
Project description:Histones are the primary building blocks of chromatin in eukaryotes and many archaea. Bacteria are thought to rely on an orthogonal set of proteins to organize their chromosomes. Several bacterial genomes do, however, encode proteins with putative histone fold domains. Whether these proteins adopt a bona fide histone fold, assemble into higher order complexes that bind DNA, and play a central role in bacterial nucleoid physiology is not known. Here, we demonstrate that histones are major and essential building blocks of chromatin in the predatory bacterium Bdellovibrio bacteriovorus and the human pathogen Leptospira interrogans and likely important in several other bacterial clades. We determine the crystal structure of the B. bacteriovorus histone (Bd0055) dimer at 1.8Å resolution to reveal that histone fold topology, handshake dimer conformation, and the RD clamp motif are conserved between bacteria, archaea, and eukaryotes. However, ostensibly minor differences, including a shorter α2 helix, a less structured α3 helix, and a more acidic surface on one side of the dimer lead to a radically divergent DNA binding mode: instead of wrapping around the outer surface of a multi-subunit histone complex, DNA forms straight fibers, encased by a sheath of tightly packed Bd0055 dimers. Our results demonstrate that bacterial histones have evolved an atypical mode of DNA binding to become integral components of chromatin in distant parts of the bacterial kingdom.
Project description:Across the tree of life, DNA in living cells is associated with proteins that coat chromosomes, constrain their structure and influence DNA-templated processes such as transcription and replication. In bacteria and eukaryotes, HU and histones, respectively, are the principal constituents of chromatin, with few exceptions. Archaea, in contrast, have more diverse repertoires of nucleoid-associated proteins (NAPs). The evolutionary and ecological drivers behind this diversity are poorly understood. Here, we combine a systematic phylogenomic survey of known and predicted NAPs with quantitative protein abundance data to shed light on the forces governing the evolution of archaeal chromatin. Our survey identifies the Diaforarchaea as a hotbed of NAP gain and loss and we validate novel candidate NAPs in two members of this clade, Thermoplasma volcanium and Methanomassiliicoccus luminyensis, using sucrose gradient-based nucleoid enrichment coupled to quantitative mass spectrometry. Comparative analysis across a panel of 19 archaea revealed that investment in NAP production varies over two orders of magnitude, from <0.03% to >5% of total protein. Integrating genomic and ecological data, we demonstrate that growth temperature is an excellent predictor of relative NAP investment across archaea. Our results suggest that high levels of chromatinization have evolved as a mechanism toprevent uncontrolled helix opening and runaway denaturation – rather than, for example, to globally orchestrate gene expression – with implications for the origin of chromatin in both archaea and eukaryotes.
Project description:Structure probing combined with next-generation sequencing (NGS) has provided novel insights into RNA structure-function relationships. To date such studies have focused largely on bacteria and eukaryotes, with little attention given to the third domain of life, archaea. Furthermore, functional RNAs have not been extensively studied in archaea, leaving open questions about RNA structure and function within this domain of life. With archaeal species being diverse and having many similarities to both bacteria and eukaryotes, the archaea domain has the potential to be an evolutionary bridge. In this study, we introduce a method for probing RNA structure in vivo in the archaea domain of life. We investigated the structure of ribosomal RNA (rRNA) from Methanosarcina acetivorans, a well-studied anaerobic archaeal species, grown with either methanol or acetate. After probing the RNA in vivo with dimethyl sulfate (DMS), Structure-seq2 libraries were generated, sequenced, and analyzed. We mapped the reactivity of DMS onto the secondary structure of the ribosome, which we determined independently with comparative analysis, and confirmed the accuracy of DMS probing in M. acetivorans. Accessibility of the rRNA to DMS in the two carbon sources was found to be quite similar, although some differences were found. Overall, this study establishes the Structure-seq2 pipeline in the archaea domain of life and informs about ribosomal structure within M. acetivorans.
Project description:We use MNase-Seq to elucidate primary chromatin architecture in an archaeon without histones, the acido-thermophilic archaeon Thermoplasma acidophilum. Like all members of the Thermoplasmatales, T. acidophilum harbours a HU family protein, HTa, that is highly expressed and protects - like histones but unlike well-characterized bacterial HU proteins – a sizeable fraction of the genome from MNase digestion. Comparing HTa-based chromatin architecture to that of three histone-encoding archaea, Methanothermus fervidus, Haloferax volcanii, and Thermococcus kodakkarensis, we present evidence that HTa is an archaeal histone analog. HTa-protected fragments are GC-rich, display histone-like mono- and dinucleotide patterns around the dyad, exhibit relatively invariant positioning throughout the growth cycle, and show archaeal histone-like oligomerization dynamics. Our results suggest that HTa, a DNA-binding protein of bacterial origin, has converged onto an architectural role filled by histones in other archaea.
Project description:We use MNase-Seq to elucidate primary chromatin architecture in an archaeon without histones, the acido-thermophilic archaeon Thermoplasma acidophilum. Like all members of the Thermoplasmatales, T. acidophilum harbours a HU family protein, HTa, that is highly expressed and protects - like histones but unlike well-characterized bacterial HU proteins – a sizeable fraction of the genome from MNase digestion. Comparing HTa-based chromatin architecture to that of three histone-encoding archaea, Methanothermus fervidus, Haloferax volcanii, and Thermococcus kodakkarensis, we present evidence that HTa is an archaeal histone analog. HTa-protected fragments are GC-rich, display histone-like mono- and dinucleotide patterns around the dyad, exhibit relatively invariant positioning throughout the growth cycle, and show archaeal histone-like oligomerization dynamics. Our results suggest that HTa, a DNA-binding protein of bacterial origin, has converged onto an architectural role filled by histones in other archaea.
Project description:ChIP-chip analyses of Psc3 in wild-type and mutant fission yeast cells. Eukaryotic genomes are folded into three-dimensional structures that govern diverse hromosomal procsses. Studeis in Drosophila and mammals have revealed large self-associating tomological domains whose borders are enriched in cohesin/CTCF factors that are required for long-range intrations. However, mechanisms governing higher-order folding of chromatin fivbers and the exact function of cohesin in this process remain poor understood. Here we perform Hi-C to explore the organization of the Schizosaccharomyces pombe genome at high-resolution, which despite its small size comprises fundamental features found in higher eukaryotes. Our analyses reveal that in addition to determinants of Rabl-like chromosome architecture, smaller locally interacting regions of chromatin, referred to as globules, are a distinctive features of S. pombe chromosome organization. This feature of chromatin architecture requires a function of cohesin distinct from its role in sister chromatid cohesion. Cohesin is enriched at globule boundaries and its loss causes disruption of local globule structure and global chromosome territories. Heterochromatin, which selectively loads cohesin at specific loci including pericentromric and subtelomeric domains, is dispensable for globule formation but uniquely impacts genome organization through chromatin compaction by enforcing Rabl configuration. Genome-wide distribution of Psc3 were determined by ChIP-chip analysis in wild-type and mutant fission yeast cells.