Direct measurement of transcription rates reveals multiple mechanisms for configuration of the Arabidopsis ambient temperature response

Background Sensing and responding to ambient temperature is important for controlling growth and development of many organisms, in part by regulating mRNA levels. mRNA abundance can change with temperature, but it is unclear whether this results from changes in transcription or decay rates, and whether passive or active temperature regulation is involved. Results Using a base analog labelling method, we directly measured the temperature coefficient, Q10, of mRNA synthesis and degradation rates of the Arabidopsis transcriptome. We show that for most genes, transcript levels are buffered against passive increases in transcription rates by balancing passive increases in the rate of decay. Strikingly, for temperature-responsive transcripts, increasing temperature raises transcript abundance primarily by promoting faster transcription relative to decay and not vice versa, suggesting a global transcriptional process exists that controls mRNA abundance by temperature. This is partly accounted for by gene body H2A.Z which is associated with low transcription rate Q10, but is also influenced by other marks and transcription factor activities. Conclusions Our data show that less frequent chromatin states can produce temperature responses simply by virtue of their rarity and the difference between their thermal properties and those of the most common states, and underline the advantages of directly measuring transcription rate changes in dynamic systems, rather than inferring rates from changes in mRNA abundance. Background The mechanism for ambient temperature sensing in plants is unclear. Control of transcript levels is believed to be important in responses to temperature [1-4] but affects of ambient temperature on transcription and mRNA decay rates have not been measured. According to the work of Arrhenius [5] the temperature coefficient (Q10) of biochemical reactions is expected to be 2 to 3 at biological temperatures: yet less than 2% of Arabidopsis thaliana genes have a two-fold or greater difference in expression level between 17°C and 27°C [6]. The remaining genes either have rates buffered against changing temperatures, or passive increases in transcription rate must be offset by a balanced increase in decay rate, leading to higher turnover but static steady state levels. Despite this fundamental uncertainty, steady state transcriptomic responses to ambient temperature have been used to infer a role for chromatin modifications in temperature signaling [2,7]. 4-Thiouracil (4SU) is a non-toxic base analogue that has been shown to be incorporated into mammalian and yeast mRNA during transcription [8-12]. Biotinylation and column separation allow 4SU-labeled RNA to be separated from unlabeled RNA, and transcriptomic analysis using the separated samples can be used to simultaneously calculate mRNA synthesis and decay rates [8]. Here we use 4SU labeling to measure transcription rates and determine the Q10 genome-wide of mRNA synthesis and decay rates in Arabidopsis thaliana. We show that ambient temperature has large passive effects on both mRNA synthesis and decay rates, and that where temperature controls transcript abundance it does so by regulating transcription relative to decay and not vice versa. Our analysis suggests that transcription factor binding sites and epigenetic state combine to create a complex network of temperature responses in plants. Results Cells incorporate 4SU into RNA and this has been exploited in mammalian cells [8,11,12] and in yeast [13] to measure mRNA synthesis and decay rates. In order to determine whether plants can take up 4SU we floated intact seedlings in MS medium and monitored 4SU incorporation into RNA by biotinylation and dot blot (Figure S1a in Additional file 1). This clearly showed that plants incorporate 4SU from the environment into RNA and that concentrations as low as 1 mM lead to a signal detectable above background within 1 hour (Figure 1B). The resulting RNA could be separated from unlabeled RNA by biotinylation and passage through a streptavidin column as described previously. At 1.5 mM the flow-through can be depleted of detectable 4SU-labeled RNA, whilst labeled plant RNA is highly concentrated in the fraction recovered from the column [8,13] (Figure S1c in Additional file 1). To maximize recovery we chose a low concentration of 4SU at 1.5 mM [8] as high labeling frequencies are known to lead to binding of fewer more frequently labeled transcripts to the columns and reduce recovery. At this concentration Arabidopsis plants treated with 4SU showed the same growth and survival as control plants (Figure S2a in Additional file 1), suggesting 4SU has low toxicity in plants, as in other organisms. Therefore, 4SU dynamics in Arabidopsis seedlings resemble those described for other experimental systems. Preliminary experiments showed that RNA turnover was faster at 27°C compared to 12°C (Figure S2b in Additional file 1), suggesting that temperature generally affected transcription rates

Evolutionary game dynamics of controlled and automatic decision-making

We integrate dual-process theories of human cognition with evolutionary game theory to study the evolution of automatic and controlled decision-making processes. We introduce a model where agents who make decisions using either automatic or controlled processing compete with each other for survival. Agents using automatic processing act quickly and so are more likely to acquire resources, but agents using controlled processing are better planners and so make more effective use of the resources they have. Using the replicator equation, we characterize the conditions under which automatic or controlled agents dominate, when coexistence is possible, and when bistability occurs. We then extend the replicator equation to consider feedback between the state of the population and the environment. Under conditions where having a greater proportion of controlled agents either enriches the environment or enhances the competitive advantage of automatic agents, we find that limit cycles can occur, leading to persistent oscillations in the population dynamics. Critically, however, these limit cycles only emerge when feedback occurs on a sufficiently long time scale. Our results shed light on the connection between evolution and human cognition, and demonstrate necessary conditions for the rise and fall of rationality.Comment: 9 pages, 7 figure

Lightweight, Self-Deploying Foam Antenna Structures

Lightweight, deployable antennas for a variety of outer-space and terrestrial applications would be designed and fabricated according to the concept of cold hibernated elastic memory (CHEM) structures, according to a proposal. Mechanically deployable antennas now in use are heavy, complex, and unreliable, and they utilize packaging volume inefficiently. The proposed CHEM antenna structures would be simple and would deploy themselves without need for any mechanisms and, therefore, would be more reliable. The proposed CHEM antenna structures would also weigh less, could be packaged in smaller volumes, and would cost less, relative to mechanically deployable antennas. The CHEM concept was described in two prior NASA Tech Briefs articles: "Cold Hibernated Elastic Memory (CHEM) Expandable Structures" (NPO-20394), Vol. 23, No. 2 (February 1999), page 56; and "Solar Heating for Deployment of Foam Structures" (NPO-20961), Vol. 25, No. 10 (October 2001), page 36. To recapitulate from the cited prior articles: The CHEM concept is one of utilizing opencell foams of shape-memory polymers (SMPs) to make lightweight, reliable, simple, and inexpensive structures that can be alternately (1) compressed and stowed compactly or (2) expanded, then rigidified for use. A CHEM structure is fabricated at full size from a block of SMP foam in its glassy state [at a temperature below the glass-transition temperature (Tg) of the SMP]. The structure is heated to the rubbery state of the SMP (that is, to a temperature above Tg) and compacted to a small volume. After compaction, the structure is cooled to the glassy state of the SMP. The compacting force can then be released and the structure remains compact as long as the temperature is kept below Tg. Upon subsequent heating of the structure above Tg, the simultaneous elastic recovery of the foam and its shape-memory effect cause the structure to expand to its original size and shape. Once thus deployed, the structure can be rigidified by cooling below Tg. Once deployed and rigidified, the structure could be heated and recompacted. In principle, there should be no limit on the achievable number of compaction/deployment/ rigidification cycles. Thus far, several different designs of a 3.5-m-long CHEM conical corrugated horn antenna have been analyzed (see figure). A small CHEM structural antenna model was fabricated and a thin, electrically conductive layer of aluminum was deposited on the inner surface of the model. This structural model was then subjected to the compaction and deployment treatments described above to demonstrate the feasibility of a CHEM corrugated horn antenna

TreeToReads - a pipeline for simulating raw reads from phylogenies.

BackgroundUsing phylogenomic analysis tools for tracking pathogens has become standard practice in academia, public health agencies, and large industries. Using the same raw read genomic data as input, there are several different approaches being used to infer phylogenetic tree. These include many different SNP pipelines, wgMLST approaches, k-mer algorithms, whole genome alignment and others; each of these has advantages and disadvantages, some have been extensively validated, some are faster, some have higher resolution. A few of these analysis approaches are well-integrated into the regulatory process of US Federal agencies (e.g. the FDA's SNP pipeline for tracking foodborne pathogens). However, despite extensive validation on benchmark datasets and comparison with other pipelines, we lack methods for fully exploring the effects of multiple parameter values in each pipeline that can potentially have an effect on whether the correct phylogenetic tree is recovered.ResultsTo resolve this problem, we offer a program, TreeToReads, which can generate raw read data from mutated genomes simulated under a known phylogeny. This simulation pipeline allows direct comparisons of simulated and observed data in a controlled environment. At each step of these simulations, researchers can vary parameters of interest (e.g., input tree topology, amount of sequence divergence, rate of indels, read coverage, distance of reference genome, etc) to assess the effects of various parameter values on correctly calling SNPs and reconstructing an accurate tree.ConclusionsSuch critical assessments of the accuracy and robustness of analytical pipelines are essential to progress in both research and applied settings

Dual positive and negative regulation of GPCR signaling by GTP hydrolysis

G protein-coupled receptors (GPCRs) regulate a variety of intracellular pathways through their ability to promote the binding of GTP to heterotrimeric G proteins. Regulator of G protein signaling (RGS) proteins increase the intrinsic GTPase activity of G-subunits and are widely regarded as negative regulators of G protein signaling. Using yeast we demonstrate that GTP hydrolysis is not only required for desensitization, but is essential for achieving a high maximal (saturated level) response. Thus RGS-mediated GTP hydrolysis acts as both a negative (low stimulation) and positive (high stimulation) regulator of signaling. To account for this we generated a new kinetic model of the G protein cycle where GGTP enters an inactive GTP-bound state following effector activation. Furthermore, in vivo and in silico experimentation demonstrates that maximum signaling output first increases and then decreases with RGS concentration. This unimodal, non-monotone dependence on RGS concentration is novel. Analysis of the kinetic model has revealed a dynamic network motif that shows precisely how inclusion of the inactive GTP-bound state for the G produces this unimodal relationship

Properties of Turbulence in the Very Local Interstellar Clouds

We have investigated the degree to which turbulence in the Very Local Interstellar Clouds resembles the highly-studied turbulence in the solar corona and the solar wind. The turbulence diagnostics for the Local Clouds are the absorption line widths measured along 32 lines of sight to nearby stars, yielding measurements for 53 absorption components (Redfield and Linsky 2004). We have tested whether the Local Cloud turbulence has the following properties of turbulence in the solar corona or the solar wind: (a) velocity fluctuations mainly perpendicular to the average magnetic field, (b) a temperature anisotropy in the sense that the perpendicular temperature is larger than the parallel temperature (or at least enhanced relative to expectation), and (c) an ion temperature which is dependent on the ion Larmor radius, in the sense that more massive ions have higher temperatures. Our analysis of the data does not show compelling evidence for any of these properties in Local Cloud turbulence, indicating possible differences with heliospheric plasmas. In the case of anisotropy of velocity fluctuations, although the expected observational signature is not seen, we cannot exclude the possibility of relatively high degrees of anisotropy (anisotropy parameter $\epsilon \sim 0.50 - 0.70$), if some other process in the the Local Clouds is causing variations in the turbulent line width from one line of sight to another. We briefly consider possible reasons for differences between coronal and solar wind turbulence and that in the Local Clouds.Comment: Submitted to the Astrophysical Journa

High-resolution temporal profiling of transcripts during Arabidopsis leaf senescence reveals a distinct chronology of processes and regulation

Leaf senescence is an essential developmental process that impacts dramatically on crop yields and involves altered regulation of thousands of genes and many metabolic and signaling pathways, resulting in major changes in the leaf. The regulation of senescence is complex, and although senescence regulatory genes have been characterized, there is little information on how these function in the global control of the process. We used microarray analysis to obtain a highresolution time-course profile of gene expression during development of a single leaf over a 3-week period to senescence. A complex experimental design approach and a combination of methods were used to extract high-quality replicated data and to identify differentially expressed genes. The multiple time points enable the use of highly informative clustering to reveal distinct time points at which signaling and metabolic pathways change. Analysis of motif enrichment, as well as comparison of transcription factor (TF) families showing altered expression over the time course, identify clear groups of TFs active at different stages of leaf development and senescence. These data enable connection of metabolic processes, signaling pathways, and specific TF activity, which will underpin the development of network models to elucidate the process of senescence

The Eastern Arm of M83 Revisited: High-Resolution Mapping of 12CO 1-0 Emission

We have used the Owens Valley Millimeter Array to map 12CO (J=1-0) along a 3.5 kpc segment of M83's eastern spiral arm at resolutions of 6.5"x3.5", 10", and 16". The CO emission in most of this segment lies along the sharp dust lane demarking the inner edge of the spiral arm, but beyond a certain point along the arm the emission shifts downstream from the dust lane to become better aligned with the young stars seen in blue and H-beta images. This morphology resembles that of the western arm of M100. Three possibilities, none of which is wholly satisfactory, are considered to explain the deviation of the CO arm from the dust lane: heating of the CO by UV radiation from young stars, heating by low-energy cosmic rays, and a molecular medium consisting of two (diffuse and dense) components which react differently to the density wave. Regardless, the question of what CO emission traces along this spiral arm is a complicated one. Strong tangential streaming is observed where the arm crosses the kinematic major axis of the galaxy, implying that the shear becomes locally prograde in the arms. Inferred from the streaming is a very high gas surface density of about 230 solar masses/pc**2 and an arm-interarm contrast greater than 2.3 in the part of the arm near the major axis. Using two different criteria, we find that the gas at this location is well above the threshold for gravitational instability -- much more clearly so than in either M51 or M100.Comment: Accepted for publication in ApJ. 25 pages, 5 figures. Manuscript in LaTeX, figures in pdf. Fig 3 in colo

Is Title VI a Magic Bullet? Environmental Racism in the Context of Political-Economic Processes and Imperatives

This Article examines avenues of redress and pollution prevention for impoverished people of color that flow from Title VI litigation strategies within the larger context of the environmental justice movement. Environmental justice issues can serve as tools with which to question status quo distributive policymaking processes and outcomes. Specifically, this Article concerns itself with practical routes toward increasing distributive justice and democratic efficacy