TEnvR: MATLAB-Based Toolbox for Environmental Research

With the advancements in science and technology, datasets become larger and more multivariate, which warrants the need for programming tools for fast data processing and multivariate statistical analysis. Here, the MATLAB-based Toolbox for Environmental Research TEnvR (pronounced ten-ver ) is introduced. This novel toolbox includes 44 open-source codes for automated data analysis from a multitude of techniques, such as ultraviolet-visible, fluorescence, and nuclear magnetic resonance spectroscopies, as well as from ultrahigh resolution mass spectrometry. Provided are codes for processing data (e.g., spectral corrections, formula assignment), visualization of figures, calculation of metrics, multivariate statistics, and automated work-up of large datasets. TEnvR allows for efficient data analysis with minimal by-hand manual work by the user, which allows scientists to do research more efficiently. This manuscript is supplemented with a detailed tutorial, example data, and screenshots, which collectively provide instructions on how to use all codes. TEnvR is novice-friendly and experience in programming with MATLAB is not required. TEnvR fulfills the need for a concise MATLAB-based toolbox for working with environmental data and will be updated annually to keep pace with the latest advances and needs for computational work in the environmental sciences

Biolability of Fresh and Photodegraded Pyrogenic Dissolved Organic Matter From Laboratory-Prepared Chars

Pyrogenic dissolved organic matter (pyDOM) is known to be an important biogeochemical constituent of aquatic ecosystems and the carbon cycle. While recent studies have examined how pyDOM production, composition, and photolability varies with parent pyrogenic solid material type, we lack an understanding of potential microbial mineralization and transformation of pyDOM in the biogeosphere. Thus, leachates of oak, charred at 400 °C and 650 °C, as well as their photodegraded counterparts were incubated with a soil‐extracted microbial consortium over 96 days. During the incubation, significantly more carbon was biomineralized from the lower versus higher temperature char leachate (45% vs. 37% lost, respectively). Further, the photodegraded leachates were biomineralized to significantly greater extents than their fresh non‐photodegraded counterparts. Kinetic modeling identified the mineralizable pyDOC fractions to have half‐lives of 9–13 days. Proton nuclear magnetic resonance spectroscopy indicated that the majority of this loss could be attributed to low molecular weight constituents of pyDOM (i.e., simple alcohols and acids). Further, the quantification of benzenepolycarboxylic acid (BPCA) molecular markers indicated that condensed aromatic compounds in pyDOM were biomineralized to much lesser extents (4.4% and 10.1% decrease in yields of ΣBPCA‐C over 66 days from 400 °C and 650 °C oak pyDOM, respectively), but most of this loss could be attributed to the biomineralization of smaller condensed clusters (four aromatic rings or less). These results highlight the contrasting bioavailability of different portions of pyDOM, and the need to examine both to evaluate its role in soil or aquatic heterotrophy and its environmental fate in the hydrogeosphere

Biolability of Fresh and Photodegraded Pyrogenic Dissolved Organic Matter From Laboratory-Prepared Chars

Pyrogenic dissolved organic matter (pyDOM) is known to be an important biogeochemical constituent of aquatic ecosystems and the carbon cycle. While recent studies have examined how pyDOM production, composition, and photolability varies with parent pyrogenic solid material type, we lack an understanding of potential microbial mineralization and transformation of pyDOM in the biogeosphere. Thus, leachates of oak, charred at 400 °C and 650 °C, as well as their photodegraded counterparts were incubated with a soil‐extracted microbial consortium over 96 days. During the incubation, significantly more carbon was biomineralized from the lower versus higher temperature char leachate (45% vs. 37% lost, respectively). Further, the photodegraded leachates were biomineralized to significantly greater extents than their fresh non‐photodegraded counterparts. Kinetic modeling identified the mineralizable pyDOC fractions to have half‐lives of 9–13 days. Proton nuclear magnetic resonance spectroscopy indicated that the majority of this loss could be attributed to low molecular weight constituents of pyDOM (i.e., simple alcohols and acids). Further, the quantification of benzenepolycarboxylic acid (BPCA) molecular markers indicated that condensed aromatic compounds in pyDOM were biomineralized to much lesser extents (4.4% and 10.1% decrease in yields of ΣBPCA‐C over 66 days from 400 °C and 650 °C oak pyDOM, respectively), but most of this loss could be attributed to the biomineralization of smaller condensed clusters (four aromatic rings or less). These results highlight the contrasting bioavailability of different portions of pyDOM, and the need to examine both to evaluate its role in soil or aquatic heterotrophy and its environmental fate in the hydrogeosphere

Selective Pressures to Maintain Attachment Site Specificity of Integrative and Conjugative Elements

Integrative and conjugative elements (ICEs) are widespread mobile genetic elements that are usually found integrated in bacterial chromosomes. They are important agents of evolution and contribute to the acquisition of new traits, including antibiotic resistances. ICEs can excise from the chromosome and transfer to recipients by conjugation. Many ICEs are site-specific in that they integrate preferentially into a primary attachment site in the bacterial genome. Site-specific ICEs can also integrate into secondary locations, particularly if the primary site is absent. However, little is known about the consequences of integration of ICEs into alternative attachment sites or what drives the apparent maintenance and prevalence of the many ICEs that use a single attachment site. Using ICEBs1, a site-specific ICE from Bacillus subtilis that integrates into a tRNA gene, we found that integration into secondary sites was detrimental to both ICEBs1 and the host cell. Excision of ICEBs1 from secondary sites was impaired either partially or completely, limiting the spread of ICEBs1. Furthermore, induction of ICEBs1 gene expression caused a substantial drop in proliferation and cell viability within three hours. This drop was dependent on rolling circle replication of ICEBs1 that was unable to excise from the chromosome. Together, these detrimental effects provide selective pressure against the survival and dissemination of ICEs that have integrated into alternative sites and may explain the maintenance of site-specific integration for many ICEs.United States. Public Health Service (Grant GM050895

DnaC Inactivation in Escherichia coli K-12 Induces the SOS Response and Expression of Nucleotide Biosynthesis Genes

Background: Initiation of chromosome replication in E. coli requires the DnaA and DnaC proteins and conditionally-lethal dnaA and dnaC mutants are often used to synchronize cell populations. Methodology/Principal Findings: DNA microarrays were used to measure mRNA steady-state levels in initiation-deficient dnaA46 and dnaC2 bacteria at permissive and non-permissive temperatures and their expression profiles were compared to MG1655 wildtype cells. For both mutants there was altered expression of genes involved in nucleotide biosynthesis at the non-permissive temperature. Transcription of the dnaA and dnaC genes was increased at the non-permissive temperature in the respective mutant strains indicating auto-regulation of both genes. Induction of the SOS regulon was observed in dnaC2 cells at 38uC and 42uC. Flow cytometric analysis revealed that dnaC2 mutant cells at non-permissive temperature had completed the early stages of chromosome replication initiation. Conclusion/Significance: We suggest that in dnaC2 cells the SOS response is triggered by persistent open-complex formation at oriC and/or by arrested forks that require DnaC for replication restart

Control of Transcription by Cell Size

Cell size increases significantly with increasing ploidy. Differences in cell size and ploidy are associated with alterations in gene expression, although no direct connection has been made between cell size and transcription. Here we show that ploidy-associated changes in gene expression reflect transcriptional adjustment to a larger cell size, implicating cellular geometry as a key parameter in gene regulation. Using RNA-seq, we identified genes whose expression was altered in a tetraploid as compared with the isogenic haploid. A significant fraction of these genes encode cell surface proteins, suggesting an effect of the enlarged cell size on the differential regulation of these genes. To test this hypothesis, we examined expression of these genes in haploid mutants that also produce enlarged size. Surprisingly, many genes differentially regulated in the tetraploid are identically regulated in the enlarged haploids, and the magnitude of change in gene expression correlates with the degree of size enlargement. These results indicate a causal relationship between cell size and transcription, with a size-sensing mechanism that alters transcription in response to size. The genes responding to cell size are enriched for those regulated by two mitogen-activated protein kinase pathways, and components in those pathways were found to mediate size-dependent gene regulation. Transcriptional adjustment to enlarged cell size could underlie other cellular changes associated with polyploidy. The causal relationship between cell size and transcription suggests that cell size homeostasis serves a regulatory role in transcriptome maintenance.National Institutes of Health (U.S.) (grant GM035010)National Institutes of Health (U.S.) (grant GM040266

Cell-Cycle Dependence of Transcription Dominates Noise in Gene Expression

The large variability in mRNA and protein levels found from both static and dynamic measurements in single cells has been largely attributed to random periods of transcription, often occurring in bursts. The cell cycle has a pronounced global role in affecting transcriptional and translational output, but how this influences transcriptional statistics from noisy promoters is unknown and generally ignored by current stochastic models. Here we show that variable transcription from the synthetic tetO promoter in S. cerevisiae is dominated by its dependence on the cell cycle. Real-time measurements of fluorescent protein at high expression levels indicate tetO promoters increase transcription rate ~2-fold in S/G2/M similar to constitutive genes. At low expression levels, where tetO promoters are thought to generate infrequent bursts of transcription, we observe random pulses of expression restricted to S/G2/M, which are correlated between homologous promoters present in the same cell. The analysis of static, single-cell mRNA measurements at different points along the cell cycle corroborates these findings. Our results demonstrate that highly variable mRNA distributions in yeast are not solely the result of randomly switching between periods of active and inactive gene expression, but instead largely driven by differences in transcriptional activity between G1 and S/G2/M.GM095733BBBE 103316MIT Startup Fun

The Min System and Nucleoid Occlusion Are Not Required for Identifying the Division Site in Bacillus subtilis but Ensure Its Efficient Utilization

Precise temporal and spatial control of cell division is essential for progeny survival. The current general view is that precise positioning of the division site at midcell in rod-shaped bacteria is a result of the combined action of the Min system and nucleoid (chromosome) occlusion. Both systems prevent assembly of the cytokinetic Z ring at inappropriate places in the cell, restricting Z rings to the correct site at midcell. Here we show that in the bacterium Bacillus subtilis Z rings are positioned precisely at midcell in the complete absence of both these systems, revealing the existence of a mechanism independent of Min and nucleoid occlusion that identifies midcell in this organism. We further show that Z ring assembly at midcell is delayed in the absence of Min and Noc proteins, while at the same time FtsZ accumulates at other potential division sites. This suggests that a major role for Min and Noc is to ensure efficient utilization of the midcell division site by preventing Z ring assembly at potential division sites, including the cell poles. Our data lead us to propose a model in which spatial regulation of division in B. subtilis involves identification of the division site at midcell that requires Min and nucleoid occlusion to ensure efficient Z ring assembly there and only there, at the right time in the cell cycle