scholarly journals THE ROLE OF A ZINC-BASED, SERUM-BORNE SULPHIDE-BINDING COMPONENT IN THE UPTAKE AND TRANSPORT OF DISSOLVED SULPHIDE BY THE CHEMOAUTOTROPHIC SYMBIONT-CONTAINING CLAM CALYPTOGENA ELONGATA

1993 ◽  
Vol 179 (1) ◽  
pp. 131-158
Author(s):  
J. J. Childress ◽  
C. R. Fisher ◽  
J. A. Favuzzi ◽  
A. J. Arp ◽  
D. R. Oros
Keyword(s):  
Isotope Composition ◽  
Stable Carbon Isotope ◽  
Binding Activity ◽  
Santa Barbara Channel ◽  
Ambient Oxygen ◽  
Sill Depth ◽  
Chemoautotrophic Symbiosis ◽  
Toxic Concentrations ◽  
Binding Component

Calyptogena elongata is a small (about 7 cm maximum length) species of vesicomyid clam which lives at depths of 494–503 m, near the sill depth, in the Santa Barbara Channel in mildly reducing muds at low ambient oxygen concentrations. This species has abundant autotrophic sulphur- oxidizing bacteria in bacteriocytes in its gills. The stable carbon isotope composition values of its gills and other tissues range from −36 to −38 “, supporting the suggestion that the primary carbon source for this symbiosis is inorganic carbon fixed by the endosymbionts. This species of clam concentrates sulphide into its blood serum by using a sulphide-binding component and into the gills by using an unknown sulphide-binding activity. In both tissues, total H2S concentrations within the clam can greatly exceed those outside. This apparently enables the clam to concentrate sufficient sulphide from the mildly reducing muds to support the needs of its endosymbionts. Both of these binding activities are reversible in vivo as shown by the rapid declines in blood and gill sulphide levels when the clams are deprived of sulphide and the rapid concentration of sulphide into the blood and gills when it is provided. For example, within minutes of exposure to 65 micromolar H2S, gill and blood total H2S concentrations in individual C. elongata exceed the external concentration; within 2 h they reach maximum concentrations of about 2 mmol l-1. When such experiments are carried out under anoxic conditions, the blood and gill total H2S concentrations approach saturation (10–20 mmol l-1), indicating that under oxic conditions the oxidation of sulphide by the clam and its endosymbionts holds the binding components below saturation and enables them to protect the animal tissues and endosymbionts from toxic concentrations of sulphide. In contrast to these results for C. elongata, our experiments show that the host of another chemoautotrophic symbiosis, Solemya reidi, does not concentrate sulphide from the medium into either its blood or its gills. Data are presented which indicate that the serum sulphide-binding component is a large molecular mass molecule with Zn2+ at the active site. This study strongly supports the model of vesicomyid functioning in which the blood- borne sulphide-binding component concentrates sulphide from the reducing environment around the clam's foot and transports this sulphide to the symbionts in the gills. Data are also presented which indicate that the clam oxidizes some sulphide to thiosulphate and transports this to the gills as well. Thus, individual C. elongata, like the previously studied C. magnifica, appear to bridge the reducing and oxidizing zones of their habitats to provide needed substrates to their endosymbionts. Examination of these two species and the anatomy of other vesicomyid species suggest that vesicomyid clams are functionally quite conservative.

scholarly journals Centromere function on minichromosomes isolated from budding yeast.

1993 ◽  
Vol 4 (8) ◽  
pp. 859-870 ◽  
Author(s):  
J Kingsbury ◽  
D Koshland
Keyword(s):  
Budding Yeast ◽  
Binding Activity ◽  
Chromosomal Dna ◽  
Microtubule Binding ◽  
Specific Factors ◽  
Stable Core ◽  
Insight Into ◽  
Binding Component

Centromeres are a complex of centromere DNA (CEN DNA) and specific factors that help mediate microtubule-dependent movement of chromosomes during mitosis. Minichromosomes can be isolated from budding yeast in a way that their centromeres retain the ability to bind microtubules in vitro. Here, we use the binding of these minichromosomes to microtubules to gain insight into the properties of centromeres assembled in vivo. Our results suggest that neither chromosomal DNA topology nor proximity of telomeres influence the cell's ability to assemble centromeres with microtubule-binding activity. The microtubule-binding activity of the minichromosome's centromere is stable in the presence of competitor CEN DNA, suggesting that the complex between the minichromosome CEN DNA and proteins directly bound to it is very stable. The efficiency of centromere binding to microtubules is dependent upon the concentration of microtubule polymer and is inhibited by ATP. These properties are similar to those exhibited by mechanochemical motors. The binding of minichromosomes to microtubules can be inactivated by the presence of 0.2 M NaCl and then reactivated by restoring NaCl to 0.1 M. In 0.2 M NaCl, some centromere factor(s) bind to microtubules, whereas other(s) apparently remain bound to the minichromosome's CEN DNA. Therefore, the yeast centromere appears to consist of two domains: the first consists of a stable core containing CEN DNA and CEN DNA-binding proteins; the second contains a microtubule-binding component(s). The molecular functions of this second domain are discussed.

When animals are not quite what they eat: diet digestibility influences 13C-incorporation rates and apparent discrimination in a mixed-feeding herbivore

2011 ◽  
Vol 89 (6) ◽  
pp. 453-465 ◽  
Author(s):  
Daryl Codron ◽  
Jacqui Codron ◽  
Matt Sponheimer ◽  
Stefano M. Bernasconi ◽  
Marcus Clauss
Keyword(s):  
Isotope Composition ◽  
Stable Carbon Isotope ◽  
Food Sources ◽  
Capra Hircus ◽  
Stable Carbon ◽  
Animal Tissues ◽  
Themeda Triandra ◽  
Low Protein ◽  
Medicago Sativa L ◽  
Discrimination Factors

The stable carbon isotope composition of animal tissues represents the weighted sum of the variety of food sources eaten. If sources differ in digestibility, tissues may overrepresent intake of more digestible items and faeces may overrepresent less digestible items. We tested this idea using whole blood and faeces of goats ( Capra hircus L., 1758) fed different food mixtures of C3 lucerne ( Medicago sativa L.) and C4 grass ( Themeda triandra Forssk.). Although blood and faecal δ13C values were broadly consistent with diet, results indicate mismatch between consumer and diet isotope compositions: both materials overrepresented the C3 (lucerne) component of diets. Lucerne had lower fibre digestibility than T. triandra, which explains the results for faeces, whereas underrepresentation of dietary C4 in blood is consistent with low protein content of the grass hay. A diet switch experiment revealed an important difference in 13C-incorporation rates across diets, which were slower for grass than lucerne diets, and in fact equilibrium states were not reached for all diets. Although more research is needed to link digestive kinetics with isotope incorporation, these results provide evidence for nonlinear relationships between consumers and their diets, invoking concerns about the conceptual value of “discrimination factors” as the prime currency for contemporary isotope ecology.

scholarly journals Novel DNA Motif Binding Activity Observed In Vivo With an Estrogen Receptor α Mutant Mouse

2014 ◽  
Vol 28 (6) ◽  
pp. 899-911 ◽  
Author(s):  
Sylvia C. Hewitt ◽  
Leping Li ◽  
Sara A. Grimm ◽  
Wipawee Winuthayanon ◽  
Katherine J. Hamilton ◽  
...  
Keyword(s):  
Dna Binding ◽  
Target Gene ◽  
Mutant Mouse ◽  
Estrogen Receptor Α ◽  
Binding Activity ◽  
Estrogen Response Element ◽  
Dna Binding Domain ◽  
Estrogen Response ◽  
Uterine Growth

Abstract Estrogen receptor α (ERα) interacts with DNA directly or indirectly via other transcription factors, referred to as “tethering.” Evidence for tethering is based on in vitro studies and a widely used “KIKO” mouse model containing mutations that prevent direct estrogen response element DNA- binding. KIKO mice are infertile, due in part to the inability of estradiol (E2) to induce uterine epithelial proliferation. To elucidate the molecular events that prevent KIKO uterine growth, regulation of the pro-proliferative E2 target gene Klf4 and of Klf15, a progesterone (P4) target gene that opposes the pro-proliferative activity of KLF4, was evaluated. Klf4 induction was impaired in KIKO uteri; however, Klf15 was induced by E2 rather than by P4. Whole uterine chromatin immunoprecipitation-sequencing revealed enrichment of KIKO ERα binding to hormone response elements (HREs) motifs. KIKO binding to HRE motifs was verified using reporter gene and DNA-binding assays. Because the KIKO ERα has HRE DNA-binding activity, we evaluated the “EAAE” ERα, which has more severe DNA-binding domain mutations, and demonstrated a lack of estrogen response element or HRE reporter gene induction or DNA-binding. The EAAE mouse has an ERα null–like phenotype, with impaired uterine growth and transcriptional activity. Our findings demonstrate that the KIKO mouse model, which has been used by numerous investigators, cannot be used to establish biological functions for ERα tethering, because KIKO ERα effectively stimulates transcription using HRE motifs. The EAAE-ERα DNA-binding domain mutant mouse demonstrates that ERα DNA-binding is crucial for biological and transcriptional processes in reproductive tissues and that ERα tethering may not contribute to estrogen responsiveness in vivo.

scholarly journals C-Terminal Deletions Can Suppress Temperature-Sensitive Mutations and Change Dominance in the Phage Mu Repressor

Genetics ◽  
1996 ◽  
Vol 142 (3) ◽  
pp. 661-672 ◽  
Author(s):  
Jodi L Vogel ◽  
Vincent Geuskens ◽  
Lucie Desmet ◽  
N Patrick Higgins ◽  
Ariane Toussaint
Keyword(s):  
Binding Activity ◽  
Temperature Sensitive ◽  
Dna Binding Activity ◽  
Heat Stable ◽  
C Terminus ◽  
Acid Domain ◽  
Mutant Forms ◽  
Amino Acid Domain

Abstract Mutations in an N-terminal 70-amino acid domain of bacteriophage Mu's repressor cause temperature-sensitive DNA-binding activity. Surprisingly, amber mutations can conditionally correct the heat-sensitive defect in three mutant forms of the repressor gene, cts25 (D43-G), cts62 (R47-Q and cts71 (M28-I), and in the appropriate bacterial host produce a heat-stable Sts phenotype (for survival of temperature shifts). Sts repressor mutants are heat sensitive when in supE or supF hosts and heat resistant when in Sup° hosts. Mutants with an Sts phenotype have amber mutations at one of three codons, Q179, Q187, or Q190. The Sts phenotype relates to the repressor size: in Sup° hosts sts repressors are shorter by seven, 10, or 18 amino acids compared to repressors in supE or supF hosts. The truncated form of the sts62-1 repressor, which lacks 18 residues (Q179–V196), binds Mu operator DNA more stably at 42° in vitro compared to its full-length counterpart (cts62 repressor). In addition to influencing temperature sensitivity, the C-terminus appears to control the susceptibility to in vivo Clp proteolysis by influencing the multimeric structure of repressor.

scholarly journals Ethanol inhibits monocyte chemotactic protein-1 expression in interleukin-1β-activated human endothelial cells

2005 ◽  
Vol 289 (4) ◽  
pp. H1669-H1675 ◽  
Author(s):  
John P. Cullen ◽  
Shariq Sayeed ◽  
Ying Jin ◽  
Nicholas G. Theodorakis ◽  
James V. Sitzmann ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Umbilical Vein ◽  
Binding Activity ◽  
Protective Effects ◽  
Monocyte Adhesion ◽  
Monocyte Chemotactic Protein ◽  
Chemotactic Protein ◽  
Subendothelial Space ◽  
Umbilical Vein Endothelial Cells

The aim of this study was to determine the effect of ethanol (EtOH) on endothelial monocyte chemotactic protein-1 (MCP-1) expression. IL-1β increased the production of MCP-1 by human umbilical vein endothelial cells from undetectable levels to ∼900 pg/ml at 24 h. EtOH dose-dependently inhibited IL-1β-stimulated MCP-1 secretion as determined by ELISA: 25 ± 1%, 35 ± 7%, and 65 ± 5% inhibition for 1, 10, and 100 mM EtOH, respectively, concomitant with inhibition of monocyte adhesion to activated endothelial cells. Similarly, EtOH dose-dependently inhibited IL-1β-stimulated MCP-1 mRNA expression. Experiments with actinomycin D demonstrated that EtOH decreased the stability of MCP-1 mRNA. In addition, EtOH significantly reduced NF-κB and AP-1 binding activity induced by IL-1β and inhibited MCP-1 gene transcription. Binding of 125I-labeled MCP-1 to its receptor (CCR2) on THP-1 human monocytic cells was not affected by EtOH treatment. Modulation of the expression of MCP-1 represents a mechanism whereby EtOH could inhibit atherogenesis by blocking the crucial early step of monocyte adhesion and subsequent recruitment to the subendothelial space. These actions of EtOH may underlie, in part, its cardiovascular protective effects in vivo.

Differential use of SCL/TAL-1 DNA-binding domain in developmental hematopoiesis

Blood ◽  
2008 ◽  
Vol 112 (4) ◽  
pp. 1056-1067 ◽  
Author(s):  
Mira T. Kassouf ◽  
Hedia Chagraoui ◽  
Paresh Vyas ◽  
Catherine Porcher
Keyword(s):  
Dna Binding ◽  
Molecular Mechanisms ◽  
Binding Activity ◽  
Hematopoietic Stem ◽  
Helix Loop Helix ◽  
Experimental Systems ◽  
Dna Binding Activity ◽  
Independent Functions

Abstract Dissecting the molecular mechanisms used by developmental regulators is essential to understand tissue specification/differentiation. SCL/TAL-1 is a basic helix-loop-helix transcription factor absolutely critical for hematopoietic stem/progenitor cell specification and lineage maturation. Using in vitro and forced expression experimental systems, we previously suggested that SCL might have DNA-binding–independent functions. Here, to assess the requirements for SCL DNA-binding activity in vivo, we examined hematopoietic development in mice carrying a germline DNA-binding mutation. Remarkably, in contrast to complete absence of hematopoiesis and early lethality in scl-null embryos, specification of hematopoietic cells occurred in homozygous mutant embryos, indicating that direct DNA binding is dispensable for this process. Lethality was forestalled to later in development, although some mice survived to adulthood. Anemia was documented throughout development and in adulthood. Cellular and molecular studies showed requirements for SCL direct DNA binding in red cell maturation and indicated that scl expression is positively autoregulated in terminally differentiating erythroid cells. Thus, different mechanisms of SCL's action predominate depending on the developmental/cellular context: indirect DNA binding activities and/or sequestration of other nuclear regulators are sufficient in specification processes, whereas direct DNA binding functions with transcriptional autoregulation are critically required in terminal maturation processes.

scholarly journals Cell Cycle-Dependent Binding of Yeast Heat Shock Factor to Nucleosomes

2000 ◽  
Vol 20 (17) ◽  
pp. 6435-6448 ◽  
Author(s):  
Christina Bourgeois Venturi ◽  
Alexander M. Erkine ◽  
David S. Gross
Keyword(s):  
Heat Shock ◽  
Nuclear Dna ◽  
Heat Shock Factor ◽  
Binding Activity ◽  
S Phase ◽  
Regulatory Sequences ◽  
Prevailing Paradigm ◽  
Cycle Dependent ◽  
Gain Access

ABSTRACT In the nucleus, transcription factors must contend with the presence of chromatin in order to gain access to their cognate regulatory sequences. As most nuclear DNA is assembled into nucleosomes, activators must either invade a stable, preassembled nucleosome or preempt the formation of nucleosomes on newly replicated DNA, which is transiently free of histones. We have investigated the mechanism by which heat shock factor (HSF) binds to target nucleosomal heat shock elements (HSEs), using as our model a dinucleosomal heat shock promoter (hsp82-ΔHSE1). We find that activated HSF cannot bind a stable, sequence-positioned nucleosome in G1-arrested cells. It can do so readily, however, following release from G1 arrest or after the imposition of either an early S- or late G2-phase arrest. Surprisingly, despite the S-phase requirement, HSF nucleosomal binding activity is restored in the absence of hsp82 replication. These results contrast with the prevailing paradigm for activator-nucleosome interactions and implicate a nonreplicative, S-phase-specific event as a prerequisite for HSF binding to nucleosomal sites in vivo.

scholarly journals Gastric Hyperplasia and Increased Proliferative Responses of Lymphocytes in Mice Lacking the COOH-terminal Ankyrin Domain of NF-κB2

1997 ◽  
Vol 186 (7) ◽  
pp. 999-1014 ◽  
Author(s):  
Hideaki Ishikawa ◽  
Daniel Carrasco ◽  
Estefania Claudio ◽  
Rolf-Peter Ryseck ◽  
Rodrigo Bravo
Keyword(s):  
T Cells ◽  
Lymphocyte Proliferation ◽  
Cytokine Production ◽  
Cell Types ◽  
Homozygous Deletion ◽  
Binding Activity ◽  
Activated T Cells ◽  
Physiological Relevance

The nfkb2 gene encodes the p100 precursor which produces the p52 protein after proteolytic cleavage of its COOH-terminal domain. Although the p52 product can act as an alternative subunit of NF-κB, the p100 precursor is believed to function as an inhibitor of Rel/NF-κB activity by cytoplasmic retention of Rel/NF-κB complexes, like other members of the IκB family. However, the physiological relevance of the p100 precursor as an IκB molecule has not been understood. To assess the role of the precursor in vivo, we generated, by gene targeting, mice lacking p100 but still containing a functional p52 protein. Mice with a homozygous deletion of the COOH-terminal ankyrin repeats of NF-κB2 (p100−/−) had marked gastric hyperplasia, resulting in early postnatal death. p100−/− animals also presented histopathological alterations of hematopoietic tissues, enlarged lymph nodes, increased lymphocyte proliferation in response to several stimuli, and enhanced cytokine production in activated T cells. Dramatic induction of nuclear κB–binding activity composed of p52-containing complexes was found in all tissues examined and also in stimulated lymphocytes. Thus, the p100 precursor is essential for the proper regulation of p52-containing Rel/NF-κB complexes in various cell types and its absence cannot be efficiently compensated for by other IκB proteins.

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