Sequence-function relationships in E. coli transcriptional regulation

Understanding how transcriptional regulatory sequence maps to regulatory function remains a difficult problem in regulatory biology. Given a particular DNA sequence for a bacterial promoter region, we would like to be able to say which transcription factors bind there, how strongly they bind, and whether they interact with each other and/or RNA polymerase, with the ultimate objective of integrating knowledge of these parameters into a prediction of gene expression levels. The theoretical framework of statistical thermodynamics provides a useful framework for doing so, enabling us to predict how gene expression levels depend on transcription factor binding energies and concentrations. We used thermodynamic models, coupled with models of the sequence-dependent binding energies of transcription factors and RNAP, to construct a genotype to phenotype map for the level of repression exhibited by the lac promoter, and tested it experimentally using a set of promoter variants from E. coli strains isolated from different natural environments. For this work, we sought to ``reverse engineer'' naturally occurring promoter sequences to understand how variations in promoter sequence affects gene expression. The natural inverse of this approach is to ``forward engineer'' promoter sequences to obtain targeted levels of gene expression. We used a high precision model of RNAP-DNA sequence dependent binding energy, coupled with a thermodynamic model relating binding energy to gene expression, to predictively design and verify a suite of synthetic E. coli promoters whose expression varied over nearly three orders of magnitude.

However, although thermodynamic models enable predictions of mean levels of gene expression, it has become evident that cell-to-cell variability or ``noise'' in gene expression can also play a biologically important role. In order to address this aspect of gene regulation, we developed models based on the chemical master equation framework and used them to explore the noise properties of a number of common E. coli regulatory motifs; these properties included the dependence of the noise on parameters such as transcription factor binding strength and copy number. We then performed experiments in which these parameters were systematically varied and measured the level of variability using mRNA FISH. The results showed a clear dependence of the noise on these parameters, in accord with model predictions.

Finally, one shortcoming of the preceding modeling frameworks is that their applicability is largely limited to systems that are already well-characterized, such as the lac promoter. Motivated by this fact, we used a high throughput promoter mutagenesis assay called Sort-Seq to explore the completely uncharacterized transcriptional regulatory DNA of the E. coli mechanosensitive channel of large conductance (MscL). We identified several candidate transcription factor binding sites, and work is continuing to identify the associated proteins.

Measurement of the neutron (^3He) spin structure function at low Q^2 : a connection between the Bjorken and Drell-Hearn-Gerasimov sum rules

The spin dependent cross sections, σT_1/2 and σT_3/2 , and asymmetries, A_∥ and A_⊥ for ³He have been measured at the Jefferson Lab's Hall A facility. The inclusive scattering process ³He(e,e)X was performed for initial beam energies ranging from 0.86 to 5.1 GeV, at a scattering angle of 15.5°. Data includes measurements from the quasielastic peak, resonance region, and the deep inelastic regime. An approximation for the extended Gerasimov-Drell-Hearn integral is presented at a 4-momentum transfer Q² of 0.2-1.0 GeV².

Also presented are results on the performance of the polarized ³He target. Polarization of ³He was achieved by the process of spin-exchange collisions with optically pumped rubidium vapor. The ³He polarization was monitored using the NMR technique of adiabatic fast passage (AFP). The average target polarization was approximately 35% and was determined to have a systematic uncertainty of roughly ±4% relative.

Ecosystem study Altenwoerth: impacts of a hydroelectric power-station on the River Danube in Austria

The aim of this article is to briefly describe the effects of the Altenwoerth Barrage, on the River Danube, on some physical variables and their consequent effects on water chemistry and the biota of the river. The methods used for biological sampling are summarised, especially those used in the limnological part of the study, and the macroinvertebrate and fish fauna listed. Comparisons are then made between the impounded section of river immediately above the dam and two unimpounded free-flowing sections of the river. Further developments on the Danube are considered.

The changing ecosystem of Lake Victoria, East Africa

Dramatic changes are occurring in the Lake Victoria ecosystem. Two-thirds of the endemic haplochromine cichlid species, of international interest for studies of evolution, have disappeared, an event associated with the sudden population explosion of piscivorous Nile perch (Lates: order Perciformes, family Centropomidae) introduced to the lake some thirty years ago. The total fish yield has, however, increased 5-fold from 1970 to 1990, but this yield is now dominated by just three fish species: the introduced Nile perch (Lates niloticus), Nile tilapia (Oreochromis niloticus), and a small endemic pelagic cyprinid (Rastrineobola argentea); these three have replaced a multispecies fishery. Contemporaneously the lake is becoming increasingly eutrophic with associated deoxygenation of the bottom waters, thereby reducing fish habitats. Conditions appear to be unstable.

Ecosystem exploitation, sustainability and biodiversity: Are they compatible?

This articles offers a basis for describing sustainability and then seeks to place this concept on an energetic basis by reference to recent advances in the understanding of patterns and processes in (mainly pelagic) fresh waters. Finally, by relating these to terrestrial ecosystems, it is shown how their sustainability may be attained through encouraging healthy fresh waters. Features of population succession are taken from observations on phytoplankton ecology.

The role of ecotoxicology for monitoring ecosystem health

The proposed EC Water Framework Directive (WFD)incorporates some new concepts in the field of water protection. Most of these concepts rely on the use of applied ecology of water systems. The expected improvement of environmental management is very new in this context. The new WFD will allow the checking of the eco-epidemiological results of several human impacts on aquatic ecosystems, such as toxic pollution and habitat modification. This paper intends to show some consequences of the WFD in the field of ecotoxicology.

Climate change and its risk reduction by mangrove ecosystem of Bangladesh

Climate change is amongst the most dreaded problems of the new millennium. Bangladesh is a coastal country bounded by Bay of Bengal on its southern part and here natural disasters are an ongoing part of human life. This paper discusses about the possible impact of climate change through tropical cyclones, storm surges, coastal erosion and sea level rise in the coastal community of Bangladesh and how they cope with these extreme events by the help of mangrove ecosystem. Both qualitative and quantitative discussions are made by collected data from different research work those are conducted in Bangladesh. Mangrove ecosystem provides both goods and services for coastal community, helps to improve livelihood options and protect them from natural disaster by providing variety of environmental support

The Semiglades: the collision of restoration, social values, and the ecosystem concept

Opinion article

MicroRNA-132 is a physiological regulator of hematopoietic stem cell function and B-cell development

MicroRNAs are a class of small non-coding RNAs that negatively regulate gene expression. Several microRNAs have been implicated in altering hematopoietic cell fate decisions. Importantly, deregulation of many microRNAs can lead to deleterious consequences in the hematopoietic system, including the onset of cancer, autoimmunity, or a failure to respond effectively to infection. As such, microRNAs fine-tune the balance between normal hematopoietic output and pathologic consequences. In this work, we explore the role of two microRNAs, miR-132 and miR-125b, in regulating hematopoietic stem cell (HSC) function and B cell development. In particular, we uncover the role of miR-132 in maintaining the appropriate balance between self-renewal, differentiation, and survival in aging HSCs by buffering the expression of a critical transcription factor, FOXO3. By maintain this balance, miR-132 may play a critical role in preventing aging-associated hematopoietic conditions such as autoimmune disease and cancer. We also find that miR-132 plays a critical role in B cell development by targeting a key transcription factor, Sox4, that is responsible for the differentiation of pro-B cells into pre-B cells. We find that miR-132 regulates B cell apoptosis, and by delivering miR-132 to mice that are predisposed to developing B cell cancers, we can inhibit the formation of these cancers and improve the survival of these mice. In addition to miR-132, we uncovered the role of another critical microRNA, miR-125b, that potentiates hematopoietic stem cell function. We found that enforced expression of miR-125b causes an aggressive myeloid leukemia by downregulation of its target Lin28a. Importantly, miR-125b also plays a critical role in inhibiting the formation of pro-B cells. Thus, we have discovered two microRNAs with important roles in regulating normal hematopoiesis, and whose dregulation can lead to deleterious consequences such as cancer in the aging hematopoietic system. Both miR-132 and miR-125b may therefore be targeted for therapeutics to inhibit age-related immune diseases associated with the loss of HSC function and cancer progression.

Investigations into the structure and function of the exocrine pancreas of lampreys. [Translation from: Morph.Jb. 110 245-269, 1967.]

As representatives of the most primitive of recent vertebrate groups, lampreys show fundamental differences in different features of organisation to the species of the remaining classes of vertebrates. The topical distinction between exocrine and endocrine pancreas is also considered among the morphological peculiarities of Petromyzontida. This study aims to contribute to a further explanation of this phenomenon. 50 brook lampreys were histologically examined.

Structure-Function Studies of Nicotinic Acetylcholine Receptors Using Selective Agonists and Positive Allosteric Modulators

This dissertation primarily describes chemical-scale studies of nicotinic acetylcholine receptors (nAChRs) in order to better understand ligand-receptor selectivity and allosteric modulation influences during receptor activation. Electrophysiology coupled with canonical and non-canonical amino acids mutagenesis is used to probe subtle changes in receptor function.

The first half of this dissertation focuses on differential agonist selectivity of α4β2-containing nAChRs. The α4β2 nAChR can assemble in alternative stoichiometries as well as assemble with other accessory subunits. Chapter 2 identifies key structural residues that dictate binding and activation of three stoichiometry-dependent α4β2 receptor ligands: sazetidine-A, cytisine, and NS9283. These do not follow previously suggested hydrogen-bonding patterns of selectivity. Instead, three residues on the complementary subunit strongly influence binding ability of a ligand and receptor activation. Chapter 3 involves isolation of a α5α4β2 receptor-enriched population to test for a potential alternative agonist binding location at the α5 α4 interface. Results strongly suggest that agonist occupation of this site is not necessary for receptor activation and that the α5 subunit only incorporates at the accessory subunit location.

The second half of this dissertation seeks to identify residue interactions with positive allosteric modulators (PAMs) of the α7 nAChR. Chapter 4 focuses on methods development to study loss of potentiation of Type I PAMs, which indicate residues vital to propagation of PAM effects and/or binding. Chapter 5 investigates α7 receptor modulation by a Type II PAM (PNU 120596). These results show that PNU 120596 does not alter the agonist binding site, thus is relegated to influencing only the gating component of activation. From this, we were able to map a potential network of residues from the agonist binding site to the proposed PNU 120596 binding site that are essential for receptor potentiation.