A noisy linear map underlies oscillations in cell size and gene expression in bacteria.

During bacterial growth, a cell approximately doubles in size before division, after which it splits into two daughter cells. This process is subjected to the inherent perturbations of cellular noise and thus requires regulation for cell-size homeostasis. The mechanisms underlying the control and dynamics of cell size remain poorly understood owing to the difficulty in sizing individual bacteria over long periods of time in a high-throughput manner. Here we measure and analyse long-term, single-cell growth and division across different Escherichia coli strains and growth conditions. We show that a subset of cells in a population exhibit transient oscillations in cell size with periods that stretch across several (more than ten) generations. Our analysis reveals that a simple law governing cell-size control-a noisy linear map-explains the origins of these cell-size oscillations across all strains. This noisy linear map implements a negative feedback on cell-size control: a cell with a larger initial size tends to divide earlier, whereas one with a smaller initial size tends to divide later. Combining simulations of cell growth and division with experimental data, we demonstrate that this noisy linear map generates transient oscillations, not just in cell size, but also in constitutive gene expression. Our work provides new insights into the dynamics of bacterial cell-size regulation with implications for the physiological processes involved.

Ets-1 p27: A novel Ets-1 isoform with dominant-negative effects on the transcriptional properties and the subcellular localization of Ets-1 p51

The transcription factor Ets-1 is implicated in various physiological processes and invasive pathologies. We identified a novel variant of ets-1, ets-1Delta(III-VI), resulting from the alternative splicing of exons III to VI. This variant encodes a 27 kDa isoform, named Ets-1 p27. Ets-1 p27 lacks the threonine-38 residue, the Pointed domain and the transactivation domain, all of which are required for the transactivation of Ets-1 target genes. Both inhibitory domains surrounding the DNA-binding domain are conserved, suggesting that Ets-1 p27, like the full-length Ets-1 p51 isoform, is autoinhibited for DNA binding. We showed that Ets-1 p27 binds DNA in the same way as Ets-1 p51 does and that it acts both at a transcriptional and a subcellular localization level, thereby constituting a dual-acting dominant negative of Ets-1 p51. Ets-1 p27 blocks Ets-1 p51-mediated transactivation of target genes and induces the translocation of Ets-1 p51 from the nucleus to the cytoplasm. Furthermore, Ets-1 p27 overexpression represses the tumor properties of MDA-MB-231 mammary carcinoma cells in correlation with the known implication of Ets-1 in various cellular mechanisms. Thus the dual-acting dominant-negative function of Ets-1 p27 gives to the Ets-1 p27/Ets-1 p51 ratio a determining effect on cell fate.

Coordinated Rhythms Of Digestion Absorption And Excretion In Mytilus-Edulis (Bivalvia Mollusca)

Sequential alternation of extracellular digestion in the stomach and intracellular digestion in the diverticula appears widespread among bivalves. The present study documents some physiological consequences of such processes in Mytilus edulis L. collected during 1981 from Whitsand Bay, Cornwall, England. Pronounced temporal fluctuations in faecal deposition are described that relate, in terms of amplitude and period, to both sinusoidal rhythmicity established for ammonia excretion and changes in the morphology of digestive tubules. Although at least partially synchronised among replicate groups of mussels, these cycles bore no consistent relationship with exogenous influences. Hourly fluctuation in the net absorption efficiency for nitrogen, as evidenced by the mean percentage ±2 SE, measured over 24 h sampling periods, was considerable (16.0±53.7, 49.3±10.9 and 52.8±6.6 for mussels acclimated in March, June and October, respectively). This variation in absorption derived from an inverse relationship between the percentage nitrogen within faeces and the rate of faecal egestion. Accordingly, peaks of faecal deposition presumably represented the pulsed remnants of intracellular digestion. Co-ordinated rhythms of digestion, absorption and excretion were thus evident in M. edulis. These processes displayed seasonally dependent periodicities of approximately 8, 3 and 4 h in March, June and October, respectively. It was concluded that, at least for M. edulis, this previously unquantified rhythmicity of physiological processes warrants careful consideration during assays commonly undertaken in the complication of nutrient and energy budgets.

Seasonal-Changes In The Physiology Reproductive Condition And Carbohydrate Content Of The Cockle Cardium (=Cerastoderma) Edule (Bivalvia Cardiidae)

Pronounced seasonal cycles in the rates of oxygen consumption and feeding were found for Cardium (=Cerastoderma) edule L. measured in the field under ambient conditions. The cockles had a maximum rate of oxygen consumption (0.89 ml O2 g-1 h-1) in April which declined to a minimum of 0.35 ml O2 g-1 h-1 in March. Their feeding rate was variable but had a maximum value (3.91 l g-1 h-1) in April and a minimum value (0.73 l g-1 h-1) in October. There was no apparent seasonal variation in absorption efficiency, with a mean value of 67.6%. Gametogenesis was initiated in January and the population reached a peak in reproductive condition in April/May, followed by a 3 month spawning period. Carbohydrate reserves were synthesised during spawning, and were then utilised during the winter and early spring. An adaptive function for a reduction in time spent feeding is postulated, and correlations between the rates of certain physiological processes and some exogenous and endogenous variables are discussed.

Assessing the environmental consequences of CO2 leakage from geological CCS: Generating evidence to support environmental risk assessment

At the start of the industrial revolution (circa 1750) the atmospheric concentration of carbon dioxide (CO2) was around 280 ppm. Since that time the burning of fossil fuel, together with other industrial processes such as cement manufacture and changing land use, has increased this value to 400 ppm, for the first time in over 3 million years. With CO2 being a potent greenhouse gas, the consequence of this rise for global temperatures has been dramatic, and not only for air temperatures. Global Sea Surface Temperature (SST) has warmed by 0.4–0.8 °C during the last century, although regional differences are evident (IPCC, 2007). This rise in atmospheric CO2 levels and the resulting global warming to some extent has been ameliorated by the oceanic uptake of around one quarter of the anthropogenic CO2 emissions (Sabine et al., 2004). Initially this was thought to be having little or no impact on ocean chemistry due to the capacity of the ocean’s carbonate buffering system to neutralise the acidity caused when CO2 dissolves in seawater. However, this assumption was challenged by Caldeira and Wickett (2005) who used model predictions to show that the rate at which carbonate buffering can act was far too slow to moderate significant changes to oceanic chemistry over the next few centuries. Their model predicted that since pre-industrial times, ocean surface water pH had fallen by 0.1 pH unit, indicating a 30% increase in the concentration of H+ ions. Their model also showed that the pH of surface waters could fall by up to 0.4 units before 2100, driven by continued and unabated utilisation of fossil fuels. Alongside increasing levels of dissolved CO2 and H+ (reduced pH) an increase in bicarbonate ions together with a decrease in carbonate ions occurs. These chemical changes are now collectively recognised as “ocean acidification”. Concern now stems from the knowledge that concentrations of H+, CO2, bicarbonate and carbonate ions impact upon many important physiological processes vital to maintaining health and function in marine organisms. Additionally, species have evolved under conditions where the carbonate system has remained relatively stable for millions of years, rendering them with potentially reduced capacity to adapt to this rapid change. Evidence suggests that, whilst the impact of ocean acidification is complex, when considered alongside ocean warming the net effect on the health and productivity of the oceans will be detrimental.

Environmental hypoxia but not minor shell damage affects scope for growth and body condition in the blue mussel Mytilus edulis (L.)

The effects of short-term (7 d) exposure to environmental hypoxia (2.11 mg O-2 L-1; control: 6.96 mg O-2 L-1) and varying degrees of shell damage (1 or 2, 1 mm diameter holes; control: no holes) on respiration rate, clearance rate, ammonia excretion rate, scope for growth (SFG) and body condition index were investigated in adult blue mussels (Mytilus edulis). There was a significant hypoxia-related reduction in SFG (>6.70 to 0.92J g(-1) h(-1)) primarily due to a reduction in energy acquisition as a result of reduced clearance rates during hypoxia. Shell damage had no significant affect on any of the physiological processes measured or the SFG calculated. Body condition was unaffected by hypoxia or shell damage. In conclusion, minor physical damage to mussels had no effect on physiological energetics but environmental hypoxia compromised growth, respiration and energy acquisition presumably by reducing feeding rates.

Fundamental shift in vitamin B12 eco-physiology of a model alga demonstrated by experimental evolution.

A widespread and complex distribution of vitamin requirements exists over the entire tree of life, with many species having evolved vitamin dependence, both within and between different lineages. Vitamin availability has been proposed to drive selection for vitamin dependence, in a process that links an organism's metabolism to the environment, but this has never been demonstrated directly. Moreover, understanding the physiological processes and evolutionary dynamics that influence metabolic demand for these important micronutrients has significant implications in terms of nutrient acquisition and, in microbial organisms, can affect community composition and metabolic exchange between coexisting species. Here we investigate the origins of vitamin dependence, using an experimental evolution approach with the vitamin B(12)-independent model green alga Chlamydomonas reinhardtii. In fewer than 500 generations of growth in the presence of vitamin B(12), we observe the evolution of a B(12)-dependent clone that rapidly displaces its ancestor. Genetic characterization of this line reveals a type-II Gulliver-related transposable element integrated into the B(12)-independent methionine synthase gene (METE), knocking out gene function and fundamentally altering the physiology of the alga.

miR-7 and miR-214 are specifically expressed during neuroblastoma differentiation, cortical development and embryonic stem cells differentiation, and control neurite outgrowth in vitro

The mammalian nervous system exerts essential control on many physiological processes in the organism and is itself controlled extensively by a variety of genetic regulatory mechanisms. microRNA (miR), an abundant class of small non-coding RNA, are emerging as important post-transcriptional regulators of gene expression in the brain. Increasing evidence indicates that miR regulate both the development and function of the nervous system. Moreover, deficiency in miR function has also been implicated in a number of neurological disorders. Expression profile analysis of miR is necessary to understand their complex role in the regulation of gene expression during the development and differentiation of cells. Here we present a comparative study of miR expression profiles in neuroblastoma, in cortical development, and in neuronal differentiation of embryonic stem (ES) cells. By microarray profiling in combination with real time PCR we show that miR-7 and miR-214 are modulated in neuronal differentiation (as compared to miR-1, -16 and -133a), and control neurite outgrowth in vitro. These findings provide an important step toward further elucidation of miR function and miR-related gene regulatory networks in the mammalian central nervous system. (C) 2010 Elsevier Inc. All rights reserved.

Unraveling the Structure and Function of G Protein-Coupled Receptors Through NMR Spectroscopy

G protein-coupled receptors (GPCRs) are a large superfamily of signaling proteins expressed on the plasma membrane. They are involved in a wide range of physiological processes and, therefore, are exploited as drug targets in a multitude of therapeutic areas. In this extent, knowledge of structural and functional properties of GPCRs may greatly facilitate rational design of modulator compounds. Solution and solid-state nuclear magnetic resonance (NMR) spectroscopy represents a powerful method to gather atomistic insights into protein structure and dynamics. In spite of the difficulties inherent the solution of the structure of membrane proteins through NMR, these methods have been successfully applied, sometimes in combination with molecular modeling, to the determination of the structure of GPCR fragments, the mapping of receptor-ligand interactions, and the study of the conformational changes associated with the activation of the receptors. In this review, we provide a summary of the NMR contributions to the study of the structure and function of GPCRs, also in light of the published crystal structures.

Force-induced activation of covalent bonds in mechanoresponsive polymeric materials

Mechanochemical transduction enables an extraordinary range of physiological processes such as the sense of touch, hearing, balance, muscle contraction, and the growth and remodelling of tissue and
bone1–6. Although biology is replete with materials systems that actively and functionally respond to mechanical stimuli, the default mechanochemical reaction of bulk polymers to large external stress is the unselective scission of covalent bonds, resulting in damage or failure7. An alternative to this degradation process is the rational molecular design of synthetic materials such that mechanical stress
favourably altersmaterial properties. A few mechanosensitive polymers with this property have been developed8–14; but their active response is mediated through non-covalent processes, which may
limit the extent to which properties can be modified and the longterm stability in structural materials. Previously, we have shown with dissolved polymer strands incorporating mechanically sensitive chemical groups—so-called mechanophores—that the directional nature of mechanical forces can selectively break and re-form covalent bonds15,16. We now demonstrate that such forceinduced covalent-bond activation can also be realized with mechanophore-linked elastomeric and glassy polymers, by using a mechanophore that changes colour as it undergoes a reversible electrocyclic ring-opening reaction under tensile stress and thus allows us to directly and locally visualize the mechanochemical reaction. We find that pronounced changes in colour and fluorescence emerge with the accumulation of plastic deformation, indicating that in these polymeric materials the transduction of mechanical force into the ring-opening reaction is an activated process. We anticipate that force activation of covalent bonds can serve as a general strategy for the development of new mechanophore building blocks that impart polymeric materials with desirable functionalities ranging from damage sensing to fully regenerative self-healing.

Histone deacetylase 3 unconventional splicing mediates endothelial-to- mesenchymal transition through transforming growth factor β2

Histone deacetylase 3 (HDAC3) plays a critical role in the maintenance of endothelial integrity and other physiological processes. In this study, we demonstrated that HDAC3 undergoes unconventional splicing during stem cell differentiation. Four different splicing variants have been identified, designated as HD3α, -β, -γ, and -Δ, respectively. HD3α was confirmed in stem cell differentiation by specific antibody against the sequences from intron 12. Immunofluorescence staining indicated that the HD3α isoform co-localized with CD31-positive or α-smooth muscle actin-positive cells at different developmental stages of mouse embryos. Overexpression of HD3α reprogrammed human aortic endothelial cells into mesenchymal cells featuring an endothelial-to-mesenchymal transition (EndMT) phenotype. HD3α directly interacts with HDAC3 and Akt1 and selectively activates transforming growth factor β2 (TGFβ2) secretion and cleavage. TGFβ2 functioned as an autocrine and/or paracrine EndMT factor. The HD3α-induced EndMT was both PI3K/Akt- and TGFβ2-dependent. This study provides the first evidence of the role of HDAC3 splicing in the maintenance of endothelial integrity.

The influence of water motion on the growth rate of the kelp Laminaria hyperborea

The kelp Laminaria hyperborea is a dominant component of the subtidal nearshore ecosystem and is subjected to a heterogeneous wave and current climate. Water motion is known to influence physiological processes in macroalgae such as photosynthesis and nutrient uptake attributed to mass-transfer limitation. The study attempts to establish the effect of water motion on the growth rates of blades and elongation rates of the stipes of L. hyperborea at adjacent wave-exposed and wave-sheltered locations over a 12month period from field observations. The observations were supported by detailed physical and chemical measurements (light, temperature, seawater nutrient concentrations and hydrodynamics) and of tissue carbon and nitrogen concentrations together with δ¹³carbon. Despite a 30% difference in the root mean square of the velocity (Vel_rms) between the two survey locations, there was no evidence to suggest that water motion had any direct influence on the growth rates of either the blades or elongation of stipes of L. hyperborea. No significant differences were observed between either environmental or plant physiological variables between the sheltered and exposed locations. Using an integral velocity parameter (Vel_rms) the present study also highlighted the importance of the tidally induced current component of water flow in the subtidal zone. 

Multidrug transporter proteins of the major facilitator superfamily: not only for drug efflux

Multidrug resistance in prokaryotes is due primarily to efflux of offending antimicrobials from the cell by representatives of several different families of integral membrane transporter proteins. Clearly, in evolutionary terms, these proteins did not arise specifically to pump human-made antimicrobials out of the cell and thereby confer resistance. Despite this, often only their role in antibiotic resistance is characterised and highlighted.
In recent years, however, a transition from the traditional anthropocentric perception of antibiotic resistance mechanisms in microorganisms has occurred, with naturally produced antimicrobials now generally regarded as physiologically important signalling molecules or sources of nutrition for bacteria rather than antimicrobial agents, and bacterial multidrug efflux proteins not merely as a defensive response to antimicrobials but as important players in fundamental physiological processes such as cellular homeostasis.
This emerging perspective supports the notion that a better understanding of the complexities of infection and multidrug resistance in bacteria can be achieved via a more detailed understanding of those physiological processes. In this chapter, we review the ‘true’ physiological roles of multidrug efflux proteins of the largest non-ATP-hydrolysing family of membrane transporters, the major facilitator superfamily, and explore the evidence for their function in processes such as pH and metal homeostasis, import and export of metabolites and biofilm formation

In vitro and in vivo methodologies for studying the Sigma 54-dependent transcription.

Here we describe approaches and methods to assaying in vitro the major variant bacterial sigma factor, Sigma 54 (σ54), in a purified system. We include the complete transcription system, binding interactions between σ54 and its activators, as well as the self-assembly and the critical ATPase activity of the cognate activators which serve to remodel the closed promoter complexes. We also present in vivo methodologies that are used to study the impact of physiological processes, metabolic states, global signalling networks, and cellular architecture on the control of σ54-dependent gene expression.

Influence of speciation in the bioavailability of cadmium to an isopod

Devido às actividades antropogénicas várias substâncias químicas têm sido introduzidas no meio ambiente em concentrações que de outro modo não ocorreriam de forma tão elevada naturalmente. Assim, o conhecimento acerca das características de um químico, tais como, o potencial para se acumular em diferentes níveis tróficos, a sua mobilidade dentro do ecossistema, a toxicidade específica e a bioacumulação, é fundamental para compreender os seus efeitos nos ecossistemas. Esta tese investiga a influência de especiação, na biodisponibilidade do cádmio (Cd) para o isópode Porcellio dilatatus, incluindo os efeitos de especiação do metal: (i) na assimilação do Cd, (ii) no modo como o Cd se distribui internamente no organismo, e (iii) como a sobrevivência e a reprodução são afectadas em isópodes terrestres. Num primeiro ensaio laboratorial avaliou-se a importância da transferência trófica na assimilação do Cd em P. dilatatus. Para tal analisou-se a eficiência de assimilação (EA) do Cd em isópodes, adicionado superficialmente ao alimento (alface) na forma de Cd(NO3)2 e contaminando o meio de crescimento da alface. A hipótese era de que a alface contaminada biologicamente através do cultivo em meio hidropónico contaminado teria uma maior proporção de complexos com proteína ou conjugado na forma de Cd (ex. Cd cisteína). A EA de Cd foi maior entre os isópodes que foram alimentados com o sal (71%, SE = 7%), do que entre os isópodes que se alimentaram de alface contaminada biologicamente (52%, SE = 5%), demonstrando-se assim num teste laboratorial que é provável que a especiação do Cd influencie a taxa de assimilação e acumulação do Cd. Na experiência alimentar que se seguiu, estudou-se em detalhe a especiação do metal comparando as EA do Cd conjugado com cisteína (Cd(Cys)2) e na forma de Cd(NO3)2, com os quais se contaminou gelatina com alface. A utilização de Cd-cisteína, proporcionou uma forma experimental para explorar a biodisponibilidade do Cd complexado dentro do tecido biológico. Como esperado, a EA de Cd em isópodes alimentados com nitrato de Cd (64%, SE = 5%) foi maior do que no caso de isópodes alimentados com o conjugado de cisteína (20%, SE = 3%). De seguida estudou-se a distribuição subcelular das espécies de Cd assimilado através de um processo de fraccionamento. Supunha-se que as diferenças de especiação de Cd reflectiria diferentes estratégias de compartimentalização, com consequências ao nível da detoxificação, armazenamento celular e distribuição subcelular do metal. O “sequestro” na forma de metal biologicamente detoxificado (BDM = proteínas estáveis ao calor - HSP e grânulos ricos em metal - RMG) foi maior nos isópodes alimentados com Cd(NO3)2, sugerindo que são mais eficientes na detoxificação de Cd (22%) do que quando alimentados com Cd(Cys)2 (15%). Foi também demonstrado que os isópodes alimentados com Cd(Cys)2 possuíam níveis de armazenamento de Cd superior nas fracções sensíveis ao metal (MSF = organelos e proteínas desnaturadas pelo calor - HDP) consideradas fracções potencialmente vulneráveis e afectando os isópodes em termos de toxicidade. As diferentes distribuições internas que se seguiram à assimilação e detoxificação das diferentes espécies de Cd foram finalmente avaliadas em termos da sobrevivência e reprodução dos isópodes. O tratamento com Cd(Cys)2 teve maior mortalidade, provavelmente devido à maior disponibilidade de Cd ingerido com implicações ao nível dos processos fisiológicos. Os isópodes alimentados com Cd(NO3)2 armazenaram o Cd nos MRG, como estratégia de detoxificação, sendo mais eficientes a detoxificar o Cd ainda que aumentando a concentração total do metal que se tornou menos tóxico para o isópode. Desta forma, o Cd nos grânulos não estava disponível para os processos fisiológicos e deixou de ser tóxico. Isso poderia estar relacionado com a resistência e tolerância aos metais devido à capacidade dos isópodes compartimentalizarem o Cd no hepatopâncreas, que actua como um mecanismo de detoxificação e contribui para a tolerância a altos níveis de cádmio. Em termos de parâmetros reprodutivos, observou-se uma redução de gestações e duração da gestação na presença de ambas as espécies de metal, mas no caso do Cd(Cys)2 as gravidezes não se concluíram. O número de jovens produzido por fêmeas alimentadas com Cd(NO3)2 foi menor do que no controlo, mas os pesos dos juvenis foram superiores. Finalmente sugere-se assim que esta abordagem seja considerada em estudos do movimento trófico de metais nas cadeias alimentares dado que se espera que a especiação de metais implique diferentes fluxos, dentro de uma dada cadeia trófica.