Publications by authors named "Chris Cox"

31 Publications

Discovery, Optimization, and Biological Characterization of 2,3,6-Trisubstituted Pyridine-Containing M Positive Allosteric Modulators.

ChemMedChem 2019 05 28;14(9):943-951. Epub 2019 Mar 28.

Department of Medicinal Chemistry, Merck & Co., Inc., West Point, PA, USA.

Herein we describe the discovery and optimization of a new series of 2,3-disubstituted and 2,3,6-trisubstituted muscarinic acetylcholine receptor 4 (M ) positive allosteric modulators (PAMs). Iterative libraries enabled rapid exploration of one-dimensional structure-activity relationships (SAR) and identification of potency-enhancing heterocycle and N-alkyl pyrazole substituents. Further optimization led to identification of the potent, receptor-subtype-selective, brain-penetrant tool compound 24 (7-[3-[1-[(1-fluorocyclopentyl)methyl]pyrazol-4-yl]-6-methyl-2-pyridyl]-3-methoxycinnoline). It is efficacious in preclinical assays that are predictive of antipsychotic effects, producing dose-dependent reversal of amphetamine-induced hyperlocomotion in rats and mice, but not in M knockout mice. Cholinergic-related adverse effects observed in rats treated with 24 at unbound plasma concentrations more than 3-fold higher than an efficacious dose in the hyperlocomotion assay were fewer and less severe than those observed in rats treated with the nonselective M agonist xanomeline, suggesting a receptor-subtype-selective PAM has the potential for an improved safety profile.
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http://dx.doi.org/10.1002/cmdc.201900088DOI Listing
May 2019

The LacI family protein GlyR3 co-regulates the operon and in .

Biotechnol Biofuels 2017 24;10:163. Epub 2017 Jun 24.

Department of Chemical and Biomolecular Engineering, University of Tennessee, Knoxville, TN 37996 USA.

Background: utilizes a wide variety of free and cellulosomal cellulases and accessory enzymes to hydrolyze polysaccharides present in complex substrates. To date only a few studies have unveiled the details by which the expression of these cellulases are regulated. Recent studies have described the auto regulation of the operon and determined that the -- gene cluster and nearby - gene cluster are co-transcribed as polycistronic mRNA.

Results: In this paper, we demonstrate that the GlyR3 protein mediates the regulation of We first identify putative GlyR3 binding sites within or just upstream of the coding regions of and . Using an electrophoretic mobility shift assay (EMSA), we determined that a higher concentration of GlyR3 is required to effectively bind to the putative site in comparison to the site. Neither the putative site nor random DNA significantly binds GlyR3. While laminaribiose interfered with GlyR3 binding to the binding site, binding to the site was unaffected. In the presence of laminaribiose, in vivo transcription of the -- gene cluster increases, while expression is repressed, compared to in the absence of laminaribiose, consistent with the results from the EMSA. An in vitro transcription assay demonstrated that GlyR3 and laminaribiose interactions were responsible for the observed patters of in vivo transcription.

Conclusions: Together these results reveal a mechanism by which is expressed at low concentrations of GlyR3 but repressed at high concentrations. In this way, is able to co-regulate both the and gene clusters in response to the availability of β-1,3-polysaccharides in its environment.
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http://dx.doi.org/10.1186/s13068-017-0849-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5483248PMC
June 2017

Evolutionary Connectionism: Algorithmic Principles Underlying the Evolution of Biological Organisation in Evo-Devo, Evo-Eco and Evolutionary Transitions.

Evol Biol 2016 8;43(4):553-581. Epub 2015 Dec 8.

Agents, Interactions and Complexity, ECS, University of Southampton, Southampton, UK.

The mechanisms of variation, selection and inheritance, on which evolution by natural selection depends, are not fixed over evolutionary time. Current evolutionary biology is increasingly focussed on understanding how the evolution of developmental organisations modifies the distribution of phenotypic variation, the evolution of ecological relationships modifies the selective environment, and the evolution of reproductive relationships modifies the heritability of the evolutionary unit. The major transitions in evolution, in particular, involve radical changes in developmental, ecological and reproductive organisations that instantiate variation, selection and inheritance at a higher level of biological organisation. However, current evolutionary theory is poorly equipped to describe how these organisations change over evolutionary time and especially how that results in adaptive complexes at successive scales of organisation (the key problem is that evolution is self-referential, i.e. the products of evolution change the parameters of the evolutionary process). Here we first reinterpret the central open questions in these domains from a perspective that emphasises the common underlying themes. We then synthesise the findings from a developing body of work that is building a new theoretical approach to these questions by converting well-understood theory and results from models of cognitive learning. Specifically, connectionist models of memory and learning demonstrate how simple incremental mechanisms, adjusting the relationships between individually-simple components, can produce organisations that exhibit complex system-level behaviours and improve the adaptive capabilities of the system. We use the term "evolutionary connectionism" to recognise that, by functionally equivalent processes, natural selection acting on the relationships within and between evolutionary entities can result in organisations that produce complex system-level behaviours in evolutionary systems and modify the adaptive capabilities of natural selection over time. We review the evidence supporting the functional equivalences between the domains of learning and of evolution, and discuss the potential for this to resolve conceptual problems in our understanding of the evolution of developmental, ecological and reproductive organisations and, in particular, the major evolutionary transitions.
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http://dx.doi.org/10.1007/s11692-015-9358-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5119841PMC
December 2015

The Low Noise Limit in Gene Expression.

PLoS One 2015 21;10(10):e0140969. Epub 2015 Oct 21.

Center for Nanophase Materials Sciences, Oak Ridge National Laboratory, Oak Ridge, Tennessee, United States of America; Bredesen Center for Interdisciplinary Research and Graduate Education, University of Tennessee, Knoxville, Tennessee, United States of America; Department of Materials Science and Engineering, University of Tennessee, Knoxville, Tennessee, United States of America.

Protein noise measurements are increasingly used to elucidate biophysical parameters. Unfortunately noise analyses are often at odds with directly measured parameters. Here we show that these inconsistencies arise from two problematic analytical choices: (i) the assumption that protein translation rate is invariant for different proteins of different abundances, which has inadvertently led to (ii) the assumption that a large constitutive extrinsic noise sets the low noise limit in gene expression. While growing evidence suggests that transcriptional bursting may set the low noise limit, variability in translational bursting has been largely ignored. We show that genome-wide systematic variation in translational efficiency can-and in the case of E. coli does-control the low noise limit in gene expression. Therefore constitutive extrinsic noise is small and only plays a role in the absence of a systematic variation in translational efficiency. These results show the existence of two distinct expression noise patterns: (1) a global noise floor uniformly imposed on all genes by expression bursting; and (2) high noise distributed to only a select group of genes.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0140969PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4619080PMC
July 2016

Transcriptomic analysis of Clostridium thermocellum Populus hydrolysate-tolerant mutant strain shows increased cellular efficiency in response to Populus hydrolysate compared to the wild type strain.

BMC Microbiol 2014 Aug 16;14:215. Epub 2014 Aug 16.

Department of Civil and Environmental Engineering, University of Tennessee, Knoxville, TN, USA.

Background: The thermophilic, anaerobic bacterium, Clostridium thermocellum is a model organism for consolidated processing due to its efficient fermentation of cellulose. Constituents of dilute acid pretreatment hydrolysate are known to inhibit C. thermocellum and other microorganisms. To evaluate the biological impact of this type of hydrolysate, a transcriptomic analysis of growth in hydrolysate-containing medium was conducted on 17.5% v/v Populus hydrolysate-tolerant mutant (PM) and wild type (WT) strains of C. thermocellum.

Results: In two levels of Populus hydrolysate medium (0% and 10% v/v), the PM showed both gene specific increases and decreases of gene expression compared to the wild-type strain. The PM had increased expression of genes in energy production and conversion, and amino acid transport and metabolism in both standard and 10% v/v Populus hydrolysate media. In particular, expression of the histidine metabolism increased up to 100 fold. In contrast, the PM decreased gene expression in cell division and sporulation (standard medium only), cell defense mechanisms, cell envelope, cell motility, and cellulosome in both media. The PM downregulated inorganic ion transport and metabolism in standard medium but upregulated it in the hydrolysate media when compared to the WT. The WT differentially expressed 1072 genes in response to the hydrolysate medium which included increased transcription of cell defense mechanisms, cell motility, and cellulosome, and decreased expression in cell envelope, amino acid transport and metabolism, inorganic ion transport and metabolism, and lipid metabolism, while the PM only differentially expressed 92 genes. The PM tolerates up to 17.5% v/v Populus hydrolysate and growth in it elicited 489 genes with differential expression, which included increased expression in energy production and conversion, cellulosome production, and inorganic ion transport and metabolism and decreased expression in transcription and cell defense mechanisms.

Conclusion: These results suggest the mechanisms of tolerance for the Populus hydrolysate-tolerant mutant strain of C. thermocellum are based on increased cellular efficiency caused apparently by downregulation of non-critical genes and increasing the expression of genes in energy production and conversion rather than tolerance to specific hydrolysate components. The wild type, conversely, responds to hydrolysate media by down-regulating growth genes and up-regulating stress response genes.
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http://dx.doi.org/10.1186/s12866-014-0215-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4236516PMC
August 2014

Industrial robustness: understanding the mechanism of tolerance for the Populus hydrolysate-tolerant mutant strain of Clostridium thermocellum.

PLoS One 2013 21;8(10):e78829. Epub 2013 Oct 21.

Department of Civil and Environmental Engineering, University of Tennessee, Knoxville, Tennessee, United States of America ; Bioenergy Science Center, Oak Ridge National Laboratory, Oak Ridge, Tennessee, United States of America.

Background: An industrially robust microorganism that can efficiently degrade and convert lignocellulosic biomass into ethanol and next-generation fuels is required to economically produce future sustainable liquid transportation fuels. The anaerobic, thermophilic, cellulolytic bacterium Clostridium thermocellum is a candidate microorganism for such conversions but it, like many bacteria, is sensitive to potential toxic inhibitors developed in the liquid hydrolysate produced during biomass processing. Microbial processes leading to tolerance of these inhibitory compounds found in the pretreated biomass hydrolysate are likely complex and involve multiple genes.

Methodology/principal Findings: In this study, we developed a 17.5% v/v Populus hydrolysate tolerant mutant strain of C. thermocellum by directed evolution. The genome of the wild type strain, six intermediate population samples and seven single colony isolates were sequenced to elucidate the mechanism of tolerance. Analysis of the 224 putative mutations revealed 73 high confidence mutations. A longitudinal analysis of the intermediate population samples, a pan-genomic analysis of the isolates, and a hotspot analysis revealed 24 core genes common to all seven isolates and 8 hotspots. Genetic mutations were matched with the observed phenotype through comparison of RNA expression levels during fermentation by the wild type strain and mutant isolate 6 in various concentrations of Populus hydrolysate (0%, 10%, and 17.5% v/v).

Conclusion/significance: The findings suggest that there are multiple mutations responsible for the Populus hydrolysate tolerant phenotype resulting in several simultaneous mechanisms of action, including increases in cellular repair, and altered energy metabolism. To date, this study provides the most comprehensive elucidation of the mechanism of tolerance to a pretreated biomass hydrolysate by C. thermocellum. These findings make important contributions to the development of industrially robust strains of consolidated bioprocessing microorganisms.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0078829PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3804516PMC
August 2014

Kinetic modeling of batch fermentation for Populus hydrolysate tolerant mutant and wild type strains of Clostridium thermocellum.

Bioresour Technol 2013 Nov 20;147:605-613. Epub 2013 Aug 20.

Department of Civil and Environmental Engineering, University of Tennessee, Knoxville, TN, United States; Bioenergy Science Center, Oak Ridge National Laboratory, Oak Ridge, TN, United States; Center for Environmental Biotechnology, University of Tennessee, Knoxville, TN, United States. Electronic address:

The extent of inhibition of two strains of Clostridium thermocellum by a Populus hydrolysate was investigated. A Monod-based model of wild type (WT) and Populus hydrolysate tolerant mutant (PM) strains of the cellulolytic bacterium C. thermocellum was developed to quantify growth kinetics in standard media and the extent of inhibition to a Populus hydrolysate. The PM was characterized by a higher growth rate (μmax=1.223 vs. 0.571 h(-1)) and less inhibition (KI,gen=0.991 vs. 0.757) in 10% v/v Populus hydrolysate compared to the WT. In 17.5% v/v Populus hydrolysate inhibition of PM increased slightly (KI,gen=0.888), whereas the WT was strongly inhibited and did not grow in a reproducible manner. Of the individual inhibitors tested, 4-hydroxybenzoic acid was the most inhibitory, followed by galacturonic acid. The PM did not have a greater ability to detoxify the hydrolysate than the WT.
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http://dx.doi.org/10.1016/j.biortech.2013.08.086DOI Listing
November 2013

Transcriptional burst frequency and burst size are equally modulated across the human genome.

Proc Natl Acad Sci U S A 2012 Oct 11;109(43):17454-9. Epub 2012 Oct 11.

Gladstone Institutes, San Francisco, CA 94158, USA.

Gene expression occurs either as an episodic process, characterized by pulsatile bursts, or as a constitutive process, characterized by a Poisson-like accumulation of gene products. It is not clear which mode of gene expression (constitutive versus bursty) predominates across a genome or how transcriptional dynamics are influenced by genomic position and promoter sequence. Here, we use time-lapse fluorescence microscopy to analyze 8,000 individual human genomic loci and find that at virtually all loci, episodic bursting--as opposed to constitutive expression--is the predominant mode of expression. Quantitative analysis of the expression dynamics at these 8,000 loci indicates that both the frequency and size of the transcriptional bursts varies equally across the human genome, independent of promoter sequence. Strikingly, weaker expression loci modulate burst frequency to increase activity, whereas stronger expression loci modulate burst size to increase activity. Transcriptional activators such as trichostatin A (TSA) and tumor necrosis factor α (TNF) only modulate burst size and frequency along a constrained trend line governed by the promoter. In summary, transcriptional bursting dominates across the human genome, both burst frequency and burst size vary by chromosomal location, and transcriptional activators alter burst frequency and burst size, depending on the expression level of the locus.
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http://dx.doi.org/10.1073/pnas.1213530109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3491463PMC
October 2012

Recommendations on bioanalytical method stability implications of co-administered and co-formulated drugs by Global CRO Council for Bioanalysis (GCC).

Bioanalysis 2012 Sep;4(17):2117-26

Advion Bioanalytical Laboratories, Quintiles, NY, USA.

An open letter written by the Global CRO Council for Bioanalysis (GCC) describing the GCC survey results on stability data from co-administered and co-formulated drugs was sent to multiple regulatory authorities on 14 December 2011. This letter and further discussions at different GCC meetings led to subsequent recommendations on this topic of widespread interest within the bioanalytical community over the past 2 years.
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http://dx.doi.org/10.4155/bio.12.192DOI Listing
September 2012

Two frizzled planar cell polarity signals in the Drosophila wing are differentially organized by the Fat/Dachsous pathway.

PLoS Genet 2011 Feb 17;7(2):e1001305. Epub 2011 Feb 17.

Department of Biological Sciences, Marshall University, Huntington, West Virginia, United States of America.

The regular array of distally pointing hairs on the mature Drosophila wing is evidence for the fine control of Planar Cell Polarity (PCP) during wing development. Normal wing PCP requires both the Frizzled (Fz) PCP pathway and the Fat/Dachsous (Ft/Ds) pathway, although the functional relationship between these pathways remains under debate. There is strong evidence that the Fz PCP pathway signals twice during wing development, and we have previously presented a Bidirectional-Biphasic Fz PCP signaling model which proposes that the Early and Late Fz PCP signals are in different directions and employ different isoforms of the Prickle protein. The goal of this study was to investigate the role of the Ft/Ds pathway in the context of our Fz PCP signaling model. Our results allow us to draw the following conclusions: (1) The Early Fz PCP signals are in opposing directions in the anterior and posterior wing and converge precisely at the site of the L3 wing vein. (2) Increased or decreased expression of Ft/Ds pathway genes can alter the direction of the Early Fz PCP signal without affecting the Late Fz PCP signal. (3) Lowfat, a Ft/Ds pathway regulator, is required for the normal orientation of the Early Fz PCP signal but not the Late Fz PCP signal. (4) At the time of the Early Fz PCP signal there are symmetric gradients of dachsous (ds) expression centered on the L3 wing vein, suggesting Ds activity gradients may orient the Fz signal. (5) Localized knockdown or over-expression of Ft/Ds pathway genes shows that boundaries/gradients of Ft/Ds pathway gene expression can redirect the Early Fz PCP signal specifically. (6) Altering the timing of ds knockdown during wing development can separate the role of the Ft/Ds pathway in wing morphogenesis from its role in Early Fz PCP signaling.
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http://dx.doi.org/10.1371/journal.pgen.1001305DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3040658PMC
February 2011

Using risk analysis to predict design flow exceedance of decentralized wastewater management systems.

Water Environ Res 2010 Dec;82(12):2357-61

AMEC Earth & Environmental, Nashville, Tennessee, USA.

In small communities, the number of residential units is a more stable indicator of wastewater volume than population. Large communities benefit from averaging because the likelihood of having concurrent large flows from all users is small. In contrast, a single septic system connected to a single residence must be designed to accommodate large flow variations. The objective of this study was to determine the risk of assigning various daily wastewater volumes to residential units. Risk was based on the probability of underestimating daily volume and, therefore, exceeding the capacity of the system. This study concluded that assigning a value of 950 L/d per residence (250 gpd per residence) is appropriate for communities with 15 or more residences, and that assigning a value of 850 L/d per residence (225 gpd per residence) is appropriate for communities with 30 or more residential connections.
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http://dx.doi.org/10.2175/106143010x12681059116374DOI Listing
December 2010

"My hip hurts": a collaboration of clinicians in the treatment of adhesive capsulitis to optimize patient care.

Authors:
Chris Cox

Int J Pharm Compd 2010 Nov-Dec;14(6):480-3

Yukon, Oklahoma.

In order to achieve a positive outcome for individuals who are diagnosed with a "frozen joint" or adhesive capsulitits, a collaborative relationship among clinicians provides support and communication. The relationship should include the physical therapist, physician, and the pharmacist. Capsulitis, or capsular restriction, is often treated with anti-inflammatory agents, localized steroidal injections, directed physical therapy, or orthopedic visual inspection. The professional judgments of coordinating healthcare practitioners provide for assessments and plans that result in favorable outcomes with optimized recovery time.
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February 2015

Legal responsibility and accountability.

Authors:
Chris Cox

Nurs Manag (Harrow) 2010 Jun;17(3):18-20

RCN.

Shifting boundaries in healthcare roles have led to anxiety among some nurses about their legal responsibilities and accountabilities. This is partly because of a lack of education about legal principles that underpin healthcare delivery. This article explains the law in terms of standards of care, duty of care, vicarious liability and indemnity insurance.
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http://dx.doi.org/10.7748/nm2010.06.17.3.18.c7797DOI Listing
June 2010

Transcriptional bursting from the HIV-1 promoter is a significant source of stochastic noise in HIV-1 gene expression.

Biophys J 2010 Apr;98(8):L32-4

Analysis of noise in gene expression has proven a powerful approach for analyzing gene regulatory architecture. To probe the regulatory mechanisms controlling expression of HIV-1, we analyze noise in gene-expression from HIV-1's long terminal repeat (LTR) promoter at different HIV-1 integration sites across the human genome. Flow cytometry analysis of GFP expression from the HIV-1 LTR shows high variability (noise) at each integration site. Notably, the measured noise levels are inconsistent with constitutive gene expression models. Instead, quantification of expression noise indicates that HIV-1 gene expression occurs through randomly timed bursts of activity from the LTR and that each burst generates an average of 2-10 mRNA transcripts before the promoter returns to an inactive state. These data indicate that transcriptional bursting can generate high variability in HIV-1 early gene products, which may critically influence the viral fate-decision between active replication and proviral latency.
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http://dx.doi.org/10.1016/j.bpj.2010.03.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2856162PMC
April 2010

Noise in biological circuits.

Wiley Interdiscip Rev Nanomed Nanobiotechnol 2009 Mar-Apr;1(2):214-25

Oak Ridge National Laboratory, Oak Ridge, TN, USA.

Noise biology focuses on the sources, processing, and biological consequences of the inherent stochastic fluctuations in molecular transitions or interactions that control cellular behavior. These fluctuations are especially pronounced in small systems where the magnitudes of the fluctuations approach or exceed the mean value of the molecular population. Noise biology is an essential component of nanomedicine where the communication of information is across a boundary that separates small synthetic and biological systems that are bound by their size to reside in environments of large fluctuations. Here we review the fundamentals of the computational, analytical, and experimental approaches to noise biology. We review results that show that the competition between the benefits of low noise and those of low population has resulted in the evolution of genetic system architectures that produce an uneven distribution of stochasticity across the molecular components of cells and, in some cases, use noise to drive biological function. We review the exact and approximate approaches to gene circuit noise analysis and simulation, and review many of the key experimental results obtained using flow cytometry and time-lapse fluorescent microscopy. In addition, we consider the probative value of noise with a discussion of using measured noise properties to elucidate the structure and function of the underlying gene circuit. We conclude with a discussion of the frontiers of and significant future challenges for noise biology.
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http://dx.doi.org/10.1002/wnan.22DOI Listing
March 2010

RCN is only prepared to take on pre-agenda for change claims.

Authors:
Chris Cox

Nurs Stand 2009 Aug;23(49):33

RCN.

The RCN takes issues of equal pay seriously. However, your article about the college supporting unfair pay claims is misleading (news August 5).
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http://dx.doi.org/10.7748/ns.23.49.33.s49DOI Listing
August 2009

Using noise to probe and characterize gene circuits.

Proc Natl Acad Sci U S A 2008 Aug 31;105(31):10809-14. Epub 2008 Jul 31.

Department of Civil and Environmental Engineering, University of Tennessee, Knoxville, TN 37996-2010, USA.

Stochastic fluctuations (or "noise") in the single-cell populations of molecular species are shaped by the structure and biokinetic rates of the underlying gene circuit. The structure of the noise is summarized by its autocorrelation function. In this article, we introduce the noise regulatory vector as a generalized framework for making inferences concerning the structure and biokinetic rates of a gene circuit from its noise autocorrelation function. Although most previous studies have focused primarily on the magnitude component of the noise (given by the zero-lag autocorrelation function), our approach also considers the correlation component, which encodes additional information concerning the circuit. Theoretical analyses and simulations of various gene circuits show that the noise regulatory vector is characteristic of the composition of the circuit. Although a particular noise regulatory vector does not map uniquely to a single underlying circuit, it does suggest possible candidate circuits, while excluding others, thereby demonstrating the probative value of noise in gene circuit analysis.
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http://dx.doi.org/10.1073/pnas.0804829105DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2504843PMC
August 2008

Bound to care.

Authors:
Chris Cox

Nurs Stand 2006 Sep 20-26;21(2):16-8

Healthcare practitioners must be better educated in the legal and ethical principles underpinning the delivery of health care.
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http://dx.doi.org/10.7748/ns.21.2.16.s24DOI Listing
October 2006

Direct quantification of N-(3-oxo-hexanoyl)-L-homoserine lactone in culture supernatant using a whole-cell bioreporter.

J Microbiol Methods 2007 Jan 17;68(1):40-5. Epub 2006 Aug 17.

Center for Environmental Biotechnology, Department of Civil and Environmental Engineering, University of Tennessee, Knoxville, TN 37996-2010, USA.

The autoinducer N-(3-oxo-hexanoyl)-L-homoserine lactone (3-oxo-C6-HSL) plays a significant role in the quorum-sensing system of the marine bacterium Vibrio fischeri. Upon forming a transcriptional activation complex with LuxR, 3-oxo-C6-HSL induces transcription of the luxICDABEG operon, leading to the increased production of both the 3-oxo-C6-HSL synthase (LuxI) and the bioluminescent proteins. In order to quantitatively analyze this regulatory mechanism, a novel approach was developed to measure 3-oxo-C6-HSL concentrations in V. fischeri cell culture supernatant. A bioluminescent strain of Escherichia coli that responds to 3-oxo-C6-HSL was used as a bioreporter. Although a linear response of the bioreporter to exogenously added synthetic 3-oxo-C6-HSL was found over several orders of magnitude, we show that bioreporter performance was dramatically impacted by variations in the supernatants using samples from a V. fischeri LuxI- strain. However, when maintained in the same supernatant background, the normalized peak bioluminescence maintained a linear response to 3-oxo-C6-HSL concentrations. Therefore, a standard additions technique was developed in which a known concentration of 3-oxo-C6-HSL was added to supernatant samples from wild-type V. fischeri cultures, and the incremental increase of the normalized peak bioluminescence relative to the untreated sample was determined. The concentration of 3-oxo-C6-HSL in the supernatant of the unknown sample was then quantified from the slope of the response between the normalized bioluminescent peaks with and without the addition of 3-oxo-C6-HSL. Advantages of this method are that it is rapid, does not require concentration or extraction, uses a small sample volume (ca. 2 ml), and accounts for effects caused by the composition of the supernatant. Furthermore, the findings can be broadly applicable to other bioreporter systems involving variable background conditions.
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http://dx.doi.org/10.1016/j.mimet.2006.06.002DOI Listing
January 2007

Frequency domain analysis of noise in simple gene circuits.

Chaos 2006 Jun;16(2):026102

Department of Civil and Environmental Engineering, University of Tennessee, Knoxville, Tennessee 37996, USA.

Recent advances in single cell methods have spurred progress in quantifying and analyzing stochastic fluctuations, or noise, in genetic networks. Many of these studies have focused on identifying the sources of noise and quantifying its magnitude, and at the same time, paying less attention to the frequency content of the noise. We have developed a frequency domain approach to extract the information contained in the frequency content of the noise. In this article we review our work in this area and extend it to explicitly consider sources of extrinsic and intrinsic noise. First we review applications of the frequency domain approach to several simple circuits, including a constitutively expressed gene, a gene regulated by transitions in its operator state, and a negatively autoregulated gene. We then review our recent experimental study, in which time-lapse microscopy was used to measure noise in the expression of green fluorescent protein in individual cells. The results demonstrate how changes in rate constants within the gene circuit are reflected in the spectral content of the noise in a manner consistent with the predictions derived through frequency domain analysis. The experimental results confirm our earlier theoretical prediction that negative autoregulation not only reduces the magnitude of the noise but shifts its content out to higher frequency. Finally, we develop a frequency domain model of gene expression that explicitly accounts for extrinsic noise at the transcriptional and translational levels. We apply the model to interpret a shift in the autocorrelation function of green fluorescent protein induced by perturbations of the translational process as a shift in the frequency spectrum of extrinsic noise and a decrease in its weighting relative to intrinsic noise.
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http://dx.doi.org/10.1063/1.2204354DOI Listing
June 2006

The sorting direct method for stochastic simulation of biochemical systems with varying reaction execution behavior.

Comput Biol Chem 2006 Feb;30(1):39-49

Computational Biology Institute, Oak Ridge National Laboratory, P.O. Box 2008 MS6164, Oak Ridge, TN 37831, USA.

A key to advancing the understanding of molecular biology in the post-genomic age is the development of accurate predictive models for genetic regulation, protein interaction, metabolism, and other biochemical processes. To facilitate model development, simulation algorithms must provide an accurate representation of the system, while performing the simulation in a reasonable amount of time. Gillespie's stochastic simulation algorithm (SSA) accurately depicts spatially homogeneous models with small populations of chemical species and properly represents noise, but it is often abandoned when modeling larger systems because of its computational complexity. In this work, we examine the performance of different versions of the SSA when applied to several biochemical models. Through our analysis, we discover that transient changes in reaction execution frequencies, which are typical of biochemical models with gene induction and repression, can dramatically affect simulator performance. To account for these shifts, we propose a new algorithm called the sorting direct method that maintains a loosely sorted order of the reactions as the simulation executes. Our measurements show that the sorting direct method performs favorably when compared to other well-known exact stochastic simulation algorithms.
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http://dx.doi.org/10.1016/j.compbiolchem.2005.10.007DOI Listing
February 2006

Statistical distributions of uncertainty and variability in activated sludge model parameters.

Authors:
Chris D Cox

Water Environ Res 2004 Nov-Dec;76(7):2672-85

Department of Civil and Environmental Engineering, Center for Environmental Biotechnology, University of Tennessee, Knoxville, 27996, USA.

All models used in activated sludge design and analysis use parameters to characterize process performance. The values of these parameters are often assumed based on default values recommended in the literature, but to date, no quantitative estimates of the parameter uncertainties have been published. Similarly, little attention has been given to quantifying site-specific parameter variability, even though its occurrence has been observed several times in the literature. In this paper, universal uncertainty distributions of the model parameters from Activated Sludge Model No. 1 are developed from a database of parameter values reported in the literature using Bayesian statistics. Site-specific distributions of parameter variability were developed using the same techniques. All parameter distributions developed demonstrated that significant uncertainty and variability exist, which could lead to overdesign or plant failure if not considered during the design process.
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August 2005

Frequency domain chemical Langevin analysis of stochasticity in gene transcriptional regulation.

J Theor Biol 2004 Aug;229(3):383-94

Molecular Scale Engineering and Nanoscale Technologies Research Group, Oak Ridge National Laboratory, Oak Ridge, TN 37381-6006, USA.

We present a frequency domain Langevin approach for stochastic analysis that remains valid for many important gene circuit elements even as molecular populations approach zero. We begin by considering the case of low-rate transcription and show that the previously reported shot noise representation is exact at all mRNA population levels for a constant transcription rate. Next, we consider transcriptional control through protein-DNA interactions at an operator site within the gene promoter region. This analysis results in expressions for the dynamics and noise behavior of this important gene sub-circuit, including the spectral density of the intrinsic operator noise and the processing of extrinsic noise by this transcriptional regulation system. This analysis shows that mRNA synthesis noise is composed of wideband shot noise and band-limited operator binding generated noise components. We find that the bandwidth of operator noise and its ultimate effect on total mRNA and protein noise is controlled by operator binding and unbinding dynamics. The most substantial impact of the operator noise is seen at transcription rates just above basal expression. This analysis captures the full behavior of this transcriptional regulation system, and points to potentially serious flaws in simplified mathematical relationships often used to model transcriptional regulation.
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http://dx.doi.org/10.1016/j.jtbi.2004.04.017DOI Listing
August 2004

Emerging foundations: nano-engineering and bio-microelectronics for environmental biotechnology.

Curr Opin Microbiol 2004 Jun;7(3):267-73

The Center for Environmental Biotechnology, Department of Microbiology, Molecular-Scale Engineering and Nanoscale Technologies Research Group, Oak Ridge National Laboratory, Knoxville, Tennessee 37996, USA.

The growth of nanotechnology, the emergence of 'nanobiotechnology', and the incorporation of living organisms in biomicroelectronic devices are revolutionizing the interdisciplinary opportunities for microbiologists to participate in understanding, developing and exploiting microbial processes in and from the environment.
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http://dx.doi.org/10.1016/j.mib.2004.04.003DOI Listing
June 2004

Analysis of noise in quorum sensing.

OMICS 2003 ;7(3):317-34

Department of Civil and Environmental Engineering, University of Tennessee, Knoxville, USA.

Noise may play a pivotal role in gene circuit functionality, as demonstrated for the genetic switch in the bacterial phage lambda. Like the lambda switch, bacterial quorum sensing (QS) systems operate within a population and contain a bistable switching element, making it likely that noise plays a functional role in QS circuit operation. Therefore, a detailed analysis of the noise behavior of QS systems is needed. We have developed a set of tools generally applicable to the analysis of gene circuits, with an emphasis on investigations in the frequency domain (FD), that we apply here to the QS system in the marine bacterium Vibrio fischeri. We demonstrate that a tight coupling between exact stochastic simulation and FD analysis provides insights into the structure/function relationships in the QS circuit. Furthermore, we argue that a noise analysis is incomplete without consideration of the power spectral densities (PSDs) of the important molecular output signals. As an example we consider reversible reactions in the QS circuit, and show through analysis and exact stochastic simulation that these circuits make significant and dynamic modifications to the noise spectra. In particular, we demonstrate a "whitening" effect, which occurs as the noise is processed through these reversible reactions.
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http://dx.doi.org/10.1089/153623103322452422DOI Listing
May 2004

Ignorance: don't let it leave you feeling ill.

Authors:
Chris Cox

Health Serv J 2003 May;113(5854):30-1

Royal College of Nursing.

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May 2003

Frequency domain analysis of noise in autoregulated gene circuits.

Proc Natl Acad Sci U S A 2003 Apr 1;100(8):4551-6. Epub 2003 Apr 1.

Molecular Scale Engineering and Nanoscale Technologies Research Group, Oak Ridge National Laboratory, P.O. Box 2008, MS 6006, Oak Ridge, TN 37831-6006, USA.

We describe a frequency domain technique for the analysis of intrinsic noise within negatively autoregulated gene circuits. This approach is based on the transfer function around the feedback loop (loop transmission) and the equivalent noise bandwidth of the system. The loop transmission, T, is shown to be a determining factor of the dynamics and the noise behavior of autoregulated gene circuits, and this T-based technique provides a simple and flexible method for the analysis of noise arising from any source within the gene circuit. We show that negative feedback not only reduces the variance of the noise in the protein concentration, but also shifts this noise to higher frequencies where it may have a negligible effect on the noise behavior of following gene circuits within a cascade. This predicted effect is demonstrated through the exact stochastic simulation of a two-gene cascade. The analysis elucidates important aspects of gene circuit structure that control functionality, and may provide some insights into selective pressures leading to this structure. The resulting analytical relationships have a simple form, making them especially useful as synthetic gene circuit design equations. With the exception of the linearization of Hill kinetics, this technique is general and may be applied to the analysis or design of networks of higher complexity. This utility is demonstrated through the exact stochastic simulation of an autoregulated two-gene cascade operating near instability.
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http://dx.doi.org/10.1073/pnas.0736140100DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC404696PMC
April 2003

Can my employer dredge up allegations against me?

Authors:
Chris Cox

Nurs Times 2002 Jun 11-17;98(24):18

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September 2002