Allostery is well documented for proteins but less recognized for DNA-protein interactions. Here we report that specific binding of a protein on DNA is substantially stabilized or destabilized by another protein bound nearby. The ternary complex's free energy oscillates as a function of the separation between the two proteins with a periodicity of ~10 base pairs, the helical pitch of B-form DNA, and a decay length of ~15 base pairs. The binding affinity of a protein near a DNA hairpin is similarly dependent on their separation, which—together with molecular dynamics simulations—suggests that deformation of the double-helical structure is the origin of DNA allostery. The physiological relevance of this phenomenon is illustrated by its effect on gene expression in live bacteria and on a transcription factor's affinity near nucleosomes.

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Genes are often transcribed by multiple RNA polymerases (RNAPs) at densities that can vary widely across genes and environmental conditions. Here, we provide in vitro and in vivo evidence for a built-in mechanism by which co-transcribing RNAPs display either collaborative or antagonistic dynamics over long distances (>2 kb) through transcription-induced DNA supercoiling. In Escherichia coli, when the promoter is active, co-transcribing RNAPs translocate faster than a single RNAP, but their average speed is not altered by large variations in promoter strength and thus RNAP density. Environmentally induced promoter repression reduces the elongation efficiency of already-loaded RNAPs, causing premature termination and quick synthesis arrest of no-longerneeded proteins. This negative effect appears independent of RNAP convoy formation and is abrogated by topoisomerase I activity. Antagonistic dynamics can also occur between RNAPs from divergently transcribed gene pairs. Our findings may be broadly applicable given that transcription on topologically constrained DNA is the norm across organisms.

Members of the genus Thermococcus, sulfur-reducing hyperthermophilic archaea, are ubiquitously present in various deep-sea hydrothermal vent systems and are considered to play a significant role in the microbial consortia. We present the complete genome sequence and feature analysis of Thermococcus onnurineus NA1 isolated from a deep-sea hydrothermal vent area, which reveal clues to its physiology. Based on results of genomic analysis, T. onnurineus NA1 possesses the metabolic pathways for organotrophic growth on peptides, amino acids, or sugars. More interesting was the discovery that the genome encoded unique proteins that are involved in carboxydotrophy to generate energy by oxidation of CO to CO 2 , thereby providing a mechanistic basis for growth with CO as a substrate. This lithotrophic feature in combination with carbon fixation via RuBisCO (ribulose 1,5-bisphosphate carboxylase/oxygenase) introduces a new strategy with a complementing energy supply for T. onnurineus NA1 potentially allowing it to cope with nutrient stress in the surrounding of hydrothermal vents, providing the first genomic evidence for the carboxydotrophy in Thermococcus.Deep-sea hydrothermal vents comprise a plethora of potential habitats, with gradients of nutrient and extreme physicochemical conditions that vary from high to low with respect to temperature (350 to 2°C), oxygenation states, and fluid velocities (13). Many multidisciplinary studies have been carried out to understand the complexities of hydrothermal vent systems. Biological studies have also been accomplished using samples collected from hydrothermal vent areas and culture-dependent and culture-independent techniques, revealing the presence of physiologically, metabolically, and phylogenetically diverse microorganisms (15). These findings have been followed by characterization of many bacterial and archaeal thermophiles (and hyperthermophiles), including both chemolithoautotrophic and chemoorganoheterotrophic strains. Among representative species of the Archaea, sulfur-reducing heterotrophs belonging to the order Thermococcales (encompassing the genera Thermococcus, Pyrococcus, and Palaeococcus) have been reported to be one of the predominant groups (20, 25). Notably, members of the species of Thermococcus were found to be more abundant in the vent ecosystem, with such isolates more frequently reported than the Pyrococcus species (9,11,23,24). Such large populations indicate some significance for the presence of Thermococcus in the microbial consortia that make up the microbial ecology of hydrothermal vent systems.In addition to ecological significance, the hyperthermophilic feature of Thermococcales has fascinated microbiologists interested in fundamental and/or application-based research. To date, the complete genome sequences of three Pyrococcus species, i.e., Pyrococcus horikoshii (16), Pyrococcus furiosus (26), and Pyrococcus abyssi (5), and a Thermococcus strain, Thermococcus kodakaraensis KOD1 (7), have been determined. Analysis of the sequences and the physiolo...

A Gram-negative, yellow-pigmented, halophilic bacterial strain US6-1 T , which degrades high-molecular-mass polycyclic aromatic hydrocarbons of two to five rings, was isolated from muddy sediment of Ulsan Bay, Republic of Korea. The 16S rRNA gene of the isolate showed high sequence similarity to Novosphingobium subarcticum (96?23 %) and Sphingopyxis alaskensis (96?18 %); however, the isolate formed a distinct phyletic line within the genus Novosphingobium. DNA-DNA relatedness between US6-1 T and the closest strain N. subarcticum revealed that strain US6-1 T was independent from this species. Isolate US6-1 T had ubiquinone 10 and a DNA

Supplemental figure legends9 Figure S1. Related to Figure 1. Cell and nucleoid morphology of E. coli cells in different growth 10 media. 11 A. Representative phase contrast and DAPI images of E. coli cells (CJW6324) grown in liquid cultures 12of M9 medium supplemented with the indicated carbon source and other chemicals (CAAT: 0.1% 13 casamino acids and 1 µg/ml thiamine) at 37 °C. For a full description of the growth media, see Table 14 S1. Cell contours (green) were generated using Oufti. 15 B. Bar graph showing the average doubling times of cultures when growing in exponential phase in 16 the indicated growth media. Errors bars indicate the standard deviation between three independent 17 biological replicates. Colors correspond to those used in Figure 1B. 18 C. Scatter plot of growth medium osmolality versus average NC ratio for E. coli cells (CJW6324) grown 19 in the media indicated in B. The color scheme corresponds to the one shown in B. Error bars indicate 20 95% confidence intervals. 21 22

We report a single-molecule assay for nucleic-acid enzymes on flow-stretched DNA templates. To facilitate the detection of slow or intermittent enzymatic activities, we developed the assay with 15-nm spatial resolution at a frame rate of 1 Hz and approximately 10 nm mechanical stability over the timescale of hours. With multiplexed data collection, we applied the assay to phi29 DNA polymerase, HIV-1 reverse transcriptase, lambda exonuclease and Escherichia coli RNA polymerase.

The target of rapamycin (TOR) kinase is an evolutionarily conserved hub of nutrient sensing and metabolic signaling. In plants, a functional connection of TOR activation with glucose availability was demonstrated, while it is yet unclear whether branched-chain amino acids (BCAAs) are a primary input of TOR signaling as they are in yeast and mammalian cells. Here, we report on the characterization of an Arabidopsis mutant over-accumulating BCAAs. Through chemical interventions targeting TOR and by examining mutants of BCAA biosynthesis and TOR signaling, we found that BCAA over-accumulation leads to up-regulation of TOR activity, which causes reorganization of the actin cytoskeleton and actin-associated endomembranes. Finally, we show that activation of TOR is concomitant with alteration of cell expansion, proliferation and specialized metabolism, leading to pleiotropic effects on plant growth and development. These results demonstrate that BCAAs contribute to plant TOR activation and reveal previously uncharted downstream subcellular processes of TOR signaling.