Crowding Conditions Research Articles

Differentiated phenotypic plasticity in larvae of the cannibalistic salamander Hynobius retardatus

Alternative phenotypes in natural populations can arise from either genetic polymorphism or an environmentally induced phenotype, that is, polyphenism. Evolutionary models of polyphenism developed by theoretical studies predict that polyphenism is favored when there are environment-dependent fitness trade-offs between alternatives and that the threshold frequency for a facultative switch between alternative phenotypes is adjusted in accordance with different selection regimes. The broad-headed (alternative) larval morph of Hynobius retardatus, which is induced by crowding with conspecifics or heterospecific anuran (Rana pirica) larvae, is a representative example of cannibalistic polyphenism. Morph induction by such proximate factors must reflect evolutionary (conditional frequency-dependent) processes. To clarify the role of frequency-dependent processes in polyphenism, I investigated the occurrence rate of the broad-headed morph under experimental crowding conditions (low conspecific, high conspecific, and high heterospecific densities) using larvae from eight natural populations with different larval densities of conspecifics and heterospecifics, and found interpopulational differences in the expression of the morph. Thus, there is a larval density-dependent equilibrium frequency of the morph in each pond, suggesting that the local switch point for morph induction was modified by selection to produce evolved differences between ponds. The evolution of such interpond differences has three necessary conditions: (1) There are pond-dependent fitness trade-offs between alternatives, (2) The maintenance of the morph is costly, and (3) The presence of conspecific or, especially, heterospecific larvae provides a reliable cue to the receiver.

Structure and chiroptical properties of supramolecular flower pigments

Research over the last 30 years has shown that at physiological concentrations of ca. 5 x 10(-3) M, flower pigments composed of anthocyanins, either alone or complexed with flavone copigments, and frequently with metals, are self-assembled into non-covalent, chiral supramolecular complexes. This serves several biological functions including color stability, protection against UV radiation and provision for specific colors to attract insects for pollination. Self-association of the monomers takes place under conditions of molecular crowding by precise matching of the pi-pi stacking interactions of the aromatic chromophores and intermolecular hydrogen bonding between the attached sugars. The resulting handedness is controlled by the chiral information provided by the sugars joined glycosidically at certain positions around the periphery of the aromatic nuclei. This review gives an overview of (i) the physicochemical evidence including circular dichroism, (1)H NMR, and X-ray analysis for the structure and supramolecular chirality of these amphiphilic complexes, (ii) the role of the sugars on directing the chirality of the resulting supramolecules, (iii) the energetics of monomer association, and (iv) the possible influence of stacking chirality on insect pollination.

Influence of Osmolytes on Inactivation and Aggregation of Muscle Glycogen Phosphorylase b by Guanidine Hydrochloride. Stimulation of Protein Aggregation under Crowding Conditions

The effects of the osmolytes trimethylamine-N-oxide (TMAO), betaine, proline, and glycine on the kinetics of inactivation and aggregation of rabbit skeletal muscle glycogen phosphorylase b by guanidine hydrochloride (GuHCl) have been studied. It is shown that the osmolytes TMAO and betaine exhibit the highest protective efficacy against phosphorylase b inactivation. A test system for studying the effects of macromolecular crowding induced by osmolytes on aggregation of proteins is proposed. TMAO and glycine increase the rate of phosphorylase b aggregation induced by GuHCl.

Structure and stability of DNA quadruplexes under molecular crowding conditions

A living cell contains generally macromolecules occupying 20-40% of the total volume. To mimic the crowded cellular condition, we prepared solutions including various cosolutes and investigated the influence of the cosolute on the structure and stability of DNA G-quadruplex. As a result, Tetrahymena telomere sequences form well-ordered G-wires in the presence of cosolutes, whereas human telomere sequences remain as compact G-quadruplexes. Since these sequence motif differ by only one base, these results demonstrate that a single base difference in telomere sequences leads to drastically different structures under the molecular crowding conditions. The findings are useful for understanding G-quadruplex structures in cell-like conditions and for design of DNA nanomaterials.

In vitro expression and characterization of native apomyoglobin under low molecular crowding conditions

The labile nature of membranes and organelles poses serious challenges to in situ biomolecule characterization in intact cells. Cell-free in vitro systems provide an alternative promising medium for the expression and characterization of protein conformation and function in a biochemical context that bears several similarities to the cellular environment. In addition, cell-free transcription–translation has recently emerged as a convenient method for protein selective isotope labeling, providing significant advantages for detailed NMR analysis. We report the cell-free expression of the model protein apomyoglobin (apoMb) in an Escherichia coli cell-free system and the effect of polyethylene glycol (PEG) on the expression yields. In contrast with in vivo protein production under control of the strong T7 promoter, apoMb is expressed in vitro in 100% soluble form. In-gel tryptic digestion followed by mass spectrometry were performed to confirm the protein identity. In order to probe the conformation of the newly expressed protein and investigate the feasibility of in situ structural analysis, high resolution protein characterization was carried out by 2D NMR spectroscopy. In vitro apoMb expression in a PEG-free environment is a convenient method for the production of soluble native-like protein under conditions amenable to selective isotopic labeling. Yields can be easily scaled-up by dialysis-assisted cell-free expression.

Drastic Effect of a Single Base Difference between Human and Tetrahymena Telomere Sequences on Their Structures under Molecular Crowding Conditions

Kleine Ursache, große Wirkung: Die Telomersequenzen von Tetrahymena bilden in Gegenwart von Cosoluten geordnete G-Drähte, die des Menschen dagegen bleiben kompakte G-Quadruplexe (siehe Bild). Eine einzige andere Base in der Telomersequenz kann somit die Telomerfunktion unter zellartigen Bedingungen beeinflussen. Die G-Draht-Struktur könnte für das Design von DNA-Nanomaterialien nützlich sein.

Drastic Effect of a Single Base Difference between Human and Tetrahymena Telomere Sequences on Their Structures under Molecular Crowding Conditions

Making a difference: Tetrahymena telomere sequences form well-ordered G-wires in the presence of cosolutes, whereas human telomere sequences remain as compact G-quadruplexes (see picture). A single base difference in telomere sequences therefore can affect telomere function under cell-like conditions. The G-wire structure may be useful in the design of DNA nanomaterials.

Social effects on crowding preferences of urban forest visitors

An image-based stated choice approach was used to investigate the conditions determining why visitors to an urban forest in Vienna feel crowded or not. Respondents ( N = 213 ) evaluated several sets of images depicting trail use scenarios with different levels of social crowding conditions and several types of social interferences. Forest users were segmented into three groups based on their global evaluations of use levels during weekends and work days, resulting in a crowding-averse, a crowding-tolerant, and a crowding-indifferent segment. Crowding-averse respondents reacted much more negatively to scenarios with high-use levels, heterogeneous trail use conditions, and violations of personal minimum spatial requirements caused by the presence of others. This user group felt overcrowded because social conditions experienced in the area interfered with their main visiting goals, especially to walk with their dog unleashed and to recreate. By contrast, crowding-tolerant respondents disliked very low-use and high-use situations, and preferred a certain amount of social stimulation in the form of some encounters, and more heterogeneous trail use conditions.

Triple helix-tetraplex equilibrium for G-rich oligonucleotide N3′→P5′ phosphoramidates: role of molecular concentration and counterions

The formation of a triple helix by a third N3′→P5′ phosphoramidate GT-rich strand 5′-d(T 9G 5) NP-3′ has been studied by FTIR, CD and UV absorbance spectroscopies. We show that in presence of sodium counterions, in conditions of molecular crowding induced by high DNA concentrations, the self-association of the phosphodiester analog into a tetrameric structure is favored and prevents the formation of the triple helix. The use of a phosphoramidate third strand in the same conditions allows to minimize self-association and to form the triple helix with a 5′-d(T 9G 5) NP-3′ third strand. Characteristic signatures of T*A·T and G*G·C base triplets have been obtained by FTIR spectroscopy. Formation of a triple helix with the unmodified GT third strand can be observed in dilute solution, in presence of lithium and divalent magnesium ions (1 M LiCl, 50 mM MgCl 2). In that case a biphasic UV melting profile ( T M at 40 and 60 °C) and a characteristic CD spectrum of a parallel stranded GT triple helix have been obtained.

Casein Proteins as Molecular Chaperones

Under conditions of stress, such as elevated temperature, molecular chaperones stabilize proteins from unfolding, aggregating, and precipitating. We have investigated the chaperone activity of the major milk proteins alpha(S)-, beta-, and kappa-casein with reduced insulin and the milk whey proteins, alpha-lactalbumin and beta-lactoglobulin, and compared it with that of the mammalian small heat shock protein (sHsp), alpha-crystallin, and clusterin. alpha(S)-Casein exhibited different chaperone behavior under reduction and heat stresses, i.e., chaperone activity increased with increasing temperature (as observed with alpha-crystallin), but under reduction stress, its chaperone activity increased at lower temperatures. beta- and kappa-casein had comparable chaperone ability with each other but were less effective than alpha(S)-casein. Under molecular crowding conditions, precipitation of stressed protein was accelerated, and alpha(S)-casein was a poorer chaperone. Furthermore, at slightly alkaline pH values, alpha(S)-casein was a less effective chaperone than at neutral pH. Detailed fluorescence, size exclusion chromatography, and real-time NMR studies studies indicated that the casein proteins underwent conformational changes and stabilized the partially unfolded whey proteins prior to formation of high molecular weight soluble complexes. These results are consistent with casein proteins acting as molecular chaperones in a manner similar to sHsps and clusterin.

Stabilization of a DNA duplex under molecular crowding conditions of PEG

A living cell generally contains macromolecules occupying 20-40% of the total volume. To mimic the crowded cellular condition, we prepared solutions including poly(ethylene glycol) (PEG) as a cosolute and investigated the influence of the cosolute on the DNA duplex stability. In the presence of PEG 200 or PEG 8000, the Tm (melting temperature) of a self-complementary duplex of 5'-dATGCGCAT-3' decreased by 11.8 degrees C in the presence of 20 wt% PEG200 and by 1.5 degrees C in the presence of 20 wt% PEG 8000. The dln K(obs) vs. dln a(w) plots for PEG 200 and PEG 8000 were linear with a negative slope, suggesting the association of water molecules upon the duplex formation. Interestingly, when the NaCl concentration decreased from 1 M to 400 mM, the Tm increased in the presence of PEGs. Our results imply that the nucleic acid stabilities in a living cell may be different from those in in vitro conditions.

Emergency department crowding and thrombolysis delays in acute myocardial infarction

We estimate the effect of emergency department (ED) crowding on door-to-needle time for patients given intravenous thrombolysis for suspected acute myocardial infarction. This was a retrospective observational study of patients thrombolyzed in the ED for suspected acute myocardial infarction in 1998 to 2000 in 25 community and teaching hospital EDs in Ontario. EDs located close together and sharing a common ambulance diversion system were grouped into networks consisting of 2 to 5 hospitals each. At patient registration in an ED, the ambulance diversion status of all EDs in the network was determined. Network crowding was calculated as the percentage of EDs that were diverting ambulances on patient registration, categorized as none (0%), moderate (<60%), and high (> or =60%). Door-to-needle time was defined as time from ED registration to drug administration. Multivariable quantile regression and logistic regression were carried out; covariates included age, sex, ECG characteristics, previous acute myocardial infarction, vital signs, time of presentation, and hospital type. A total of 3,452 thrombolysis patients were included: mean age was 62.9 years, and 73% were male patients. Overall median door-to-needle time was 43 minutes (interquartile ratio 27 to 80). Median door-to-needle time was 40, 45, and 47 minutes in conditions of none, moderate, and high network crowding, respectively ( P <.001). The adjusted odds ratios for door-to-needle time delay (>30 minutes) and major delay (>60 minutes) were 1.32 (95% confidence interval [CI] 0.98 to 1.79) and 1.40 (95% CI 1.12 to 1.75), respectively, for high network crowding compared with none, and 1.21 (95% CI 0.89 to 1.63) and 1.06 (95% CI 0.86 to 1.29), respectively, for moderate crowding compared with none. In multivariate analyses, moderate and high crowding conditions were associated with increased median door-to-needle time (3.0 minutes [95% CI 0.1 to 6.0] and 5.8 minutes [95% CI 2.7 to 9.0], respectively). ED crowding is associated with increased door-to-needle times for patients with suspected acute myocardial infarction and may represent a barrier to improving cardiac care in EDs.

Compact Modeling of the Noise of a Bipolar Transistor Under DC and AC Current Crowding Conditions

The effect of current crowding on dc, on ac, and in particular on the noise characteristic of bipolar transistors, is studied. An equivalent circuit able to model these effects is presented. General formulations to calculate current crowding in arbitrary geometries are derived. Both rectangular and circular geometries are discussed in detail.

Effect of molecular crowding on self-association of phosphorylase kinase and its interaction with phosphorylase b and glycogen.

Self-association of phosphorylase kinase (PhK) and its interaction with glycogen (M=5500 kDa) and phosphorylase b (Phb) has been studied using analytical ultracentrifugation and turbidimetry under the conditions of molecular crowding arising from the presence of high concentrations of osmolytes. In accordance with the predictions of the molecular crowding theory, trimethylamine N-oxide (TMAO) and betaine greatly favor self-association of PhK induced by Mg2+ and Ca2+ and PhK interaction with glycogen. In contrast, proline suppresses these processes, probably, due to its specific interaction with PhK. All osmolytes tested prevented the complex formation between PhK and its physiological substrate, Phb. The specific interactions of PhK and Phb with glycogen, in the living cell, presumably is a factor allowing the negative effect of crowding on the recognition of Phb by PhK to be overcome.

The effect of macromolecular crowding on protein aggregation and amyloid fibril formation.

Macromolecular crowding is expected to have several significant effects on protein aggregation; the major effects will be those due to excluded volume and increased viscosity. In this report we summarize data demonstrating that macromolecular crowding may lead to a dramatic acceleration in the rate of protein aggregation and formation of amyloid fibrils, using the protein alpha-synuclein. The aggregation of alpha-synuclein has been implicated as a critical factor in development of Parkinson's disease. Various types of polymers, from neutral polyethylene glycols and polysaccharides (Ficolls, dextrans) to inert proteins, are shown to accelerate alpha-synuclein fibrillation. The stimulation of fibrillation increases with increasing length of polymer, as well as increasing polymer concentration. At lower polymer concentrations (typically up to approximately 100 mg/ml) the major effect is ascribed to excluded volume, whereas at higher polymer concentrations evidence of opposing viscosity effects become apparent. Pesticides and metals, which are linked to increased risk of Parkinson's disease by epidemiological studies, are shown to accelerate alpha-synuclein fibrillation under conditions of molecular crowding.

Personal control and coping with retail crowding

To provide high quality services under conditions of crowding, it is important to understand the relationships between crowding and personal control. Indeed, in recent years, there has been growing belief that personal control is significant in coping with crowding. However, most studies have been of limited theoretical and practical value because they did not provide an integrated conceptualization of crowding. The results of a field study demonstrate that the personal control‐crowding relationships depend on the individual's crowding experience and the nature of personal control.

Apical sensory neurones mediate developmental retardation induced by conspecific environmental stimuli in freshwater pulmonate snails.

Freshwater pond snails Helisoma trivolvis and Lymnaea stagnalis undergo larval development and metamorphosis inside egg capsules. We report that their development is permanently under slight tonic inhibitory influence of the anterior sensory monoaminergic neurones, which are the remnants of the apical sensory organ. Conspecific juvenile snails, when reared under conditions of starvation and crowding, release chemical signals that are detected by these neurones in encapsulated larvae and reversibly suppress larval development, thus providing a link between environmental signals and developmental regulation. Induced retardation starts from the trochophore stage and results in up to twofold prolongation of the larval lifespan. Upon stimulation with the signal, the neurones increase synthesis and release of monoamines [serotonin (5-HT) in Helisoma and dopamine in Lymnaea] that inhibit larval development acting via ergometrine-sensitive internal receptors. Thus, the novel regulatory mechanism in larval development of molluscs is suggested and compared with the phenomenon of dauer larvae formation in the nematode Caenorhabditis elegans.

Polymerization of nucleotide-free, GDP- and GTP-bound cell division protein FtsZ: GDP makes the difference

Stable, more than 98% nucleotide-free apo-FtsZ was prepared from purified Methanococcus jannaschhi FtsZ. This facilitates the study of the functional mechanisms of this FtsZ, an assembling GTPase, which shares a common fold with eukaryotic tubulin. Apo-FtsZ underwent cooperative magnesium-induced polymerization with a similar critical concentration and morphology related to that of reconstituted GTP-bound FtsZ, suggesting that the binding of GTP contributes insignificantly to the stability of the FtsZ polymers. On the other hand, reconstituted GDP-FtsZ polymerized with a larger critical concentration than GTP-FtsZ, indicating that GDP binding destabilizes FtsZ polymers. Upon GTP hydrolysis by FtsZ polymers, in the absence of a continued GTP supply and under macromolecular crowding conditions enhancing FtsZ polymerization, the straight GTP polymers disappeared and were replaced by characteristic helically curved GDP-bound polymers. These results suggest that the roles of GTP binding and hydrolysis by this archaeal FtsZ are simply to facilitate disassembly. In a physiological situation in GTP excess, GDP-bound FtsZ subunits could again bind GTP, or trigger disassembly, or be recognized by FtsZ filament depolymerizing proteins, allowing the Z-ring dynamics during prokaryotic cell division.

Immobilization and cold stress affect sympatho–adrenomedullary system and pituitary–adrenocortical axis of rats exposed to long-term isolation and crowding

Changes in plasma levels of noradrenaline (NA), adrenaline (A), adrenocorticotropic hormone (ACTH) and corticosterone (CORT), as well as in cytosol glucocorticoid receptor (GR) and heat shock protein 70 (Hsp 70) in hippocampus of adult rat males exposed to two long-term types of psychosocial stress, both under basal conditions and in response to immobilization and cold as heterotypic additional stressor were studied. Long-term isolation produced a significant elevation of basal plasma ACTH and CORT levels, but did not affect that of NA and A, while long-term crowding conditions did not elevate the basal plasma levels of these hormones. Long-term isolation of rats exposed to 2 h of immobilization or cold led to a significant elevation of plasma NA, A and CORT in comparison with the controls. Long-term crowding conditions and exposure of animals to immobilization or cold also resulted in an increased plasma NA, A and CORT levels, but to a lesser extent in comparison with the long-term isolation. At the same time, plasma ACTH was significantly more elevated in long-term crowded than in long-term isolated rats. Both kinds of long-term psychosocial stresses (isolation and crowding) had similar but less pronounced effects on cytosol GR and Hsp 70 concentrations in hippocampus comparing to acute immobilization and cold stress. It seems that long-term psychosocial stresses attenuate the effects of an additional stress on hippocampal GR and Hsp 70 concentrations. These data suggest that individual housing of rats appear to act as a stronger stressor than crowding conditions. When the animals suffering a long-term isolation were exposed to either acute immobilization or cold, a stronger activation of the sympatho–adrenomedullary system (SAS) was recorded in comparison with that found in the long-term crowded group subjected to short-term immobilization or cold. No significant differences in the activity of hypotalamo–pituitary–adrenal (HPA) axis were observed between long-term isolated and long-term crowded rats.

Tuberculosis in Bissau: incidence and risk factors in an urban community in sub-Saharan Africa.

Despite the long history of tuberculosis (TB) research, population-based studies from developing countries are rare. In a prospective community study in Bissau, the capital of Guinea-Bissau, we assessed the impact of demographic, socioeconomic and cultural risk factors on active TB. A surveillance system in four districts of the capital identified 247 adult (>or=15 years) cases of intrathoracic TB between May 1996 and June 1998. Risk factors were evaluated comparing cases with the 25,189 adults living in the area in May 1997. The incidence of intrathoracic TB in the adult population was 471 per 100 000 person-years. Significant risk factors in a multivariate analysis were increasing age (P < 0.0001), male sex (odds ratio [OR] = 2.58, 95% CI: 1.85, 3.60), ethnic group other than the largest group (Pepel) (OR = 1.64, 95% CI: 1.20, 2.22), adult crowding (OR = 1.68, 95% CI: 1.18, 2.39 for >2 adults in household), and poor quality of housing (OR = 1.66, 95% CI: 1.24, 2.22). Household type was important; adults living alone or with adults of their own sex only, had a higher risk of developing TB than households with husband and wife present, the adjusted OR being 1.76 (95% CI: 1.11, 2.78) for male households and 3.80 (95% CI: 1.69, 8.56) for female households. In a multivariate analysis excluding household type, child crowding was a protective factor, the OR being 0.68 (95% CI: 0.51, 0.90) for households with >2 children per household. Bissau has a very high incidence of intrathoracic TB. Human immunodeficiency virus (HIV), increasing age, male sex, ethnicity, adult crowding, family structure, and poor housing conditions were independent risk factors for TB. Apart from HIV prevention, TB control programmes need to emphasize risk factors such as socioeconomic inequality, ethnic differences, crowding, and gender.