Nε-Lysine Acetylation of the Histone-Like Protein HBsu Regulates the Process of Sporulation and Affects the Resistance Properties of Bacillus subtilis Spores

Bacillus subtilis produces dormant, highly resistant endospores in response to extreme environmental stresses or starvation. These spores are capable of persisting in harsh environments for many years, even decades, without essential nutrients. Part of the reason that these spores can survive such extreme conditions is because their chromosomal DNA is well protected from environmental insults. The α/β-type small acid-soluble proteins (SASPs) coat the spore chromosome, which leads to condensation and protection from such insults. The histone-like protein HBsu has been implicated in the packaging of the spore chromosome and is believed to be important in modulating SASP-mediated alterations to the DNA, including supercoiling and stiffness. Previously, we demonstrated that HBsu is acetylated at seven lysine residues, and one physiological function of acetylation is to regulate chromosomal compaction. Here, we investigate if the process of sporulation or the resistance properties of mature spores are influenced by the acetylation state of HBsu. Using our collection of point mutations that mimic the acetylated and unacetylated forms of HBsu, we first determined if acetylation affects the process of sporulation, by determining the overall sporulation frequencies. We found that specific mutations led to decreases in sporulation frequency, suggesting that acetylation of HBsu at some sites, but not all, is required to regulate the process of sporulation. Next, we determined if the spores produced from the mutant strains were more susceptible to heat, ultraviolet (UV) radiation and formaldehyde exposure. We again found that altering acetylation at specific sites led to less resistance to these stresses, suggesting that proper HBsu acetylation is important for chromosomal packaging and protection in the mature spore. Interestingly, the specific acetylation patterns were different for the sporulation process and resistance properties of spores, which is consistent with the notion that a histone-like code exists in bacteria. We propose that specific acetylation patterns of HBsu are required to ensure proper chromosomal arrangement, packaging, and protection during the process of sporulation.

An optimal growth law for RNA composition and its partial implementation through ribosomal and tRNA gene locations in bacterial genomes

The distribution of cellular resources across bacterial proteins has been quantified through phenomenological growth laws. Here, we describe a complementary bacterial growth law for RNA composition, emerging from optimal cellular resource allocation into ribosomes and ternary complexes. The predicted decline of the tRNA/rRNA ratio with growth rate agrees quantitatively with experimental data. Its regulation appears to be implemented in part through chromosomal localization, as rRNA genes are typically closer to the origin of replication than tRNA genes and thus have increasingly higher gene dosage at faster growth. At the highest growth rates in E. coli, the tRNA/rRNA gene dosage ratio based on chromosomal positions is almost identical to the observed and theoretically optimal tRNA/rRNA expression ratio, indicating that the chromosomal arrangement has evolved to favor maximal transcription of both types of genes at this condition.

Chromosomal arrangement of synthetic lethal gene pairs: repulsion or attraction?

Synthetic lethal interactions are of paramount importance both in biology and in medicine, and hence increasing efforts have been devoted to their systematic identification. Our previous computational analysis revealed that in prokaryotic species, synthetic lethal genes tend to be further away in chromosomes than random (i.e. repulsion), which was shown to provide bacterial genomes with greater robustness to large-scale DNA deletions. To test the generalizability of this observation in eukaryotic genomes, we leveraged the wealth of experimentally determined synthetic lethal genetic interactions of yeast that are curated in the BioGRID (Biological General Repository for Interaction Datasets) database. We observed an opposite trend that is the genomic proximity of synthetic lethal gene pairs both on the 2D and 3D chromosomal space of the yeast genome (i.e. 2D and 3D attraction). To gain mechanistic insights into the origin of the attraction of synthetic lethal gene pairs in S. cerevisiae, we characterized four classes of genes, in which synthetic lethal interactions are enriched and partly explain the observed patterns of genomic attraction: i) gene pairs operating on the same pathways, 2) co-expressed genes, 3) gene pairs whose protein products physically interact and 4) the paralogs. However, our analysis revealed that the contribution of these four types of genes is not sufficient to fully explain the observed 2D and 3D attraction of synthetic lethal gene pairs and hence its evolutionary origin still remains as an open question.Significance statementUnravelling the organizing principles underlying gene arrangements is one of the fundamental questions of research in evolutionary biology. One understudied aspect of this organization is the relative chromosomal arrangement of synthetic lethal gene pairs. In this study, by analyzing a wealth of synthetic lethality data in yeast, we provide evidence that synthetic lethal gene pairs tend to be attracted to each other both on 2D and 3D chromosomal space of the yeast genome. This observation is in sharp contrast with the repulsion of synthetic lethal (metabolic) gene pairs that we observed previously in bacterial genomes. Characterizing the evolutionary forces underlying this genomic pattern in yeast can open the door towards an evolutionary theory of genome organization in eukaryotes.

Cell morphology and nucleoid dynamics in dividing D. radiodurans

Our knowledge of bacterial nucleoids originates mostly from studies of rod- or crescent-shaped bacteria. Here, we reveal that Deinococcus radiodurans, a relatively large, spherical bacterium, possessing a multipartite genome, and well-known for its radioresistance, constitutes a valuable system for the study of nucleoids in cocci. Using advanced microscopy, we show that as D. radiodurans progresses through its cell cycle, it undergoes coordinated morphological changes at both the cellular and nucleoid level. D. radiodurans nucleoids were found to be highly condensed, but also surprisingly dynamic, adopting multiple distinct configurations and presenting a novel chromosomal arrangement in which oriC loci are radially distributed around clustered ter sites maintained at the centre of cells. Single-molecule and ensemble studies of the abundant histone-like HU protein suggest that its loose binding to DNA may contribute to this remarkable plasticity. These findings clearly demonstrate that nucleoid organization is complex and tightly coupled to cell cycle progression.

The Staphylococcus aureus Two-Component System AgrAC Displays Four Distinct Genomic Arrangements That Delineate Genomic Virulence Factor Signatures

Two-component systems (TCSs) consist of a histidine kinase and a response regulator. Here, we evaluated the conservation of the AgrAC TCS among 149 completely sequenced S. aureus strains. It is composed of four genes: agrBDCA. We found that: i) AgrAC system (agr) was found in all but one of the 149 strains; ii) The agr positive strains were further classified into four agr types based on AgrD protein sequences, iii) the four agr types not only specified the chromosomal arrangement of the agr genes but also the sequence divergence of AgrC histidine kinase protein, which confers signal specificity, iv) the sequence divergence was reflected in distinct structural properties especially in the transmembrane region and second extracellular binding domain, and v) there was a strong correlation between the agr type and the virulence genomic profile of the organism. Taken together, these results demonstrate that bioinformatic analysis of the agr locus leads to a classification system that correlates with the presence of virulence factors and protein structural properties.

Genome-level parameters describe the pan-nuclear fractal nature of eukaryotic interphase chromosomal arrangement

Long-range inter-chromosomal interactions in the interphase nucleus subsume critical genome-level regulatory functions such as transcription and gene expression. To decipher a physical basis of diverse pan-nuclear inter-chromosomal arrangement that facilitates these processes, we investigate the scaling effects as obtained from disparate eukaryotic genomes and compare their total number of genes with chromosome size. First, we derived the pan-nuclear average fractal dimension of inter-chromosomal arrangement in interphase nuclei of different species and corroborated our predictions with independently reported results. Then, we described the different patterns across disparate unicellular and multicellular eukaryotes. We report that, unicellular lower eukaryotes have inter-chromosomal fractal dimension ≈ 1 at pan-nuclear dimensions, which is analogous to the multi-polymer crumpled globule model. Multi-fractal dimensions, corresponding to different inter-chromosomal arrangements emerged from multicellular eukaryotes, such that closely related species have relatively similar patterns. Using this theoretical approach, we computed that the average fractal dimension from human acrocentric versus metacentric chromosomes was distinct, implying that a multi-fractal nature of inter-chromosomal geometry may facilitate viable large-scale chromosomal aberrations, such as Robertsonian translocation. Next, based on inter-chromosomal geometry, we also report that this average multi-fractal dimension in nocturnal mammals is distinct from diurnal ones, and our result seems to corroborate the plasticity of the inter-chromosomal arrangement reported among nocturnal species. (For example, the arrangement of heterochromatin versus euchromatin in rod photoreceptor and fibroblast cells of Mus musculus is inverted.) Altogether, our results substantiate that genome-level constraints have also co-evolved with the average pan-nuclear fractal dimension of inter-chromosomal folding during eukaryotic evolution.

Systems-level chromosomal parameters represent the suprachromosomal basis for a non-random chromosomal arrangement in human interphase nuclei

Forty-six chromosome territories (CTs) are positioned uniquely in the human interphase nuclei, wherein each of their position can range from the center of the nucleus to its periphery. A non-empirical basis for their non-random arrangement remains unreported. Here, we derive a suprachromosomal basis of that overall arrangement (which we refer to as a CT constellation), and report a hierarchical nature of the same. Using matrix algebra, we unify intrinsic chromosomal parameters (e.g., chromosomal length, gene density, the number of genes per chromosome), to derive an extrinsic effective gene density matrix, the hierarchy of which is dominated by the extrinsic mathematical coupling of HSA19, followed by HSA17 (human chromosome 19 and 17, both preferentially interior CTs) with all CTs. We corroborate predicted constellations and the effective gene density hierarchy with published reports from fluorescent in situ hybridization based microscopy and Hi-C techniques, and delineate analogous hierarchy in disparate vertebrates. Our theory accurately predicts CTs localized to the nuclear interior, which interestingly share conserved synteny with HSA19 and/or HSA17. Finally, the effective gene density hierarchy dictates how inter-CT permutations represent plasticity within constellations and we suggest that a differential mix of coding with noncoding genome may modulate the same.