Trapping by Clusters of Channels,Receptors, and Transporters: Quantitative Description

Various membrane functional units such as receptors, transporters, and channels, whose action necessarily involves capturing diffusing molecules, are often organized into multimeric complexes forming clusters on the cell and organelle membranes. These functional units themselves are usually oligomers of several integral proteins, which have their own symmetry. Depending on the symmetry, they form clusters on different packing lattices. Moreover, local membrane inhomogeneities, e.g., the so-called membrane domains, rafts, stalks, etc., lead to different patterns even within the structures on the same packing lattice. Units in the cluster compete for diffusing molecules and screen each other. Here we propose a general approach that allows one to quantify the screening effects. The approach is used to derive simple approximate formulas giving the trapping rates of diffusing molecules by clusters of absorbers on lattices of different packing symmetries. The obtained results describe smooth variation of the trapping rate from the sum of the rates of individual absorbers forming the cluster to the effective collective rate. The latter shows how the trapping efficiency of an individual absorber decreases as the number of absorbers in the cluster increases and/or the inter-absorber distance decreases. Numerical tests demonstrate good agreement between the rates predicted by the theory and obtained from Brownian dynamics simulations for clusters of different shapes and sizes.     

Resistance to Bemisia tabaci biotype B of Solanum pimpinellifolium is associated with higher densities of type IV glandular trichomes and acylsugar accumulation

Advances in tomato breeding for pest resistance have been achieved via gene introgression from wild Solanum (section Lycopersicon) species (Solanaceae). Ninety‐nine F₃ families derived from an interspecific cross using as parental lines Solanum lycopersicum L. ‘LAM‐148' (susceptible standard) and Solanum pimpinellifolium L. ‘TO‐937‐15’ (multiple pest resistance accession with type IV glandular trichomes and acylsugar accumulation) were evaluated for their resistance against the whitefly Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) biotype B in free‐choice and no‐choice tests for oviposition and adult colonization. The parental lines and eight F₃ families with contrasting levels of resistance against the whitefly were selected and investigated in additional assays, which included the estimation of trichome densities and foliar acylsugar levels. The F₃ families BTR‐302 and BTR‐331 exhibited low amounts of eggs of whitefly and transgressive segregation for type IV glandular trichome density with values greater than that of TO‐937‐15 plants. However, the tested families did not surpass the total foliar acylsugar content found in TO‐937‐15. BTR‐331 exhibited low colonization in the free‐choice test and it was the least preferred F₃ family in the no‐choice test. The higher resistance levels of BTR‐331 were associated with a positive combination of higher type IV trichome density and higher acylsugar levels. Some F₃ families displayed reduced fruit set due to the presence of flowers with style exertion of the antheridial‐cone. Fruit weight at harvest stage of the selected families (from 4.9 to 14.5 g) was lower than that of LAM‐148 (139.5 g) but higher than that of TO‐937‐15 plants (1.3 g). Therefore, although difficult to reach due to the simultaneous segregation of many polygenic traits, the combination of high B. tabaci resistance levels with superior horticultural traits is feasible. These results confirm TO‐937‐15 as a source of biotype B resistance. From the breeding standpoint, the genetic similarity between S. lycopersicum and S. pimpinellifolium would allow a more efficient resistance introgression by facilitating recombination and minimizing the potentially undesirable linkage drag associated with this trait.     

Phenotypic integration of morphology and energetic performance under routine capacities: a study in the leaf-eared mouse Phyllotis darwini

A major goal of evolutionary physiology is to understand the intrinsic and the extrinsic factors that impose limitations on an animal’s energy budget. Although natural selection acts upon organismal traits such as performance (e.g., burst, sustained metabolic rates), from a mechanistic perspective, organismal performance results from the integrated functioning of different levels of biological organization. Hence, a better understanding of whole-animal performance must necessarily incorporate an explicit analysis of the integration between those different levels. Although this topic has been under intense scrutiny, overall there have been very few consistent patterns. Here, we explore the phenotypic integration between organ masses and the overall energy budget under routine capacities by statistically decomposing the covariance matrix (using path analysis and canonical correlation analysis) between organ masses and thermoregulatory burst and sustained metabolisms in cold acclimated individuals of Phyllotis darwini. Our results suggest that (a) central organs associated with the processing of food (cecum and liver), residuals (kidneys) and pumping of O₂ (heart) are tightly integrated to sustained expenditure and between themselves; (b) with the exception of the heart, central energy supplying organs are weakly related to burst expenditures; (c) sustained and burst metabolisms refer to complete different strategies and (d) basal metabolic rate is not related to any of the physiological or morphological traits considered in this study. Overall, our results support the hypothesis of an economic phenotype: animals maintain their excess capacities to face those critical extreme events, but their physiology and internal morphology are tightly integrated to function under routine needs.     

Improved secretion of the cancer-testis antigen SSX2 in Pichia pastoris by deletion of its nuclear localization signal

The cancer-testis (CT) antigen synovial sarcoma X break point 2 (SSX2) was expressed in Pichia pastoris as a means to produce a delayed-type hypersensitivity skin test reagent for monitoring SSX2-specific anti-cancer immune responses. SSX2 was detected intracellularly in P. pastoris despite the addition of the Saccharomyces cerevisiae alpha-mating factor secretion signal. Increasing the SSX2 gene copy number did not improve its secretion but did enhance intracellular SSX2 levels. SSX2 with its C-terminal nuclear localization signal (NLS) deleted (SSX2NORD), however, was secreted. Indirect immunofluorescence indicated that SSX2 containing the NLS did not translocate to the nucleus but accumulated in the endoplasmic reticulum (ER). Experimental results further suggested that SSX2 containing the NLS was misfolded in the ER, while deletion of the NLS facilitated correct folding of SSX2 inside the ER and improved its secretion. Production of SSX2NORD was scaled-up to a 2-L fermentor using a fed-batch protocol to maintain methanol at a concentration of 1 g L⁻¹. Decreasing the cultivation temperature from 25 °C to 16 °C improved protein stability in the culture supernatant. In this process, after 120 h cultivation, the wet cell weight of P. pastoris reached 280 mg mL⁻¹, and the yield of SSX2NORD was 21.6 mg L⁻¹.     

Experimental Evidence that Sexual Displays are Costly for Nest Survival

Nest predation is one of the most significant limitations for successful breeding of tropical passerines. Thus, parental strategies may include choosing appropriate nest sites and behaving in ways that minimize predation. Habitat characteristics that may influence nest success include degree of nest concealment, proximity to habitat edge, plant architecture as well as several others cited in the literature. However, few studies have examined display behavior as a factor that could also influence nest survival. We experimentally tested whether sexual motor displays served as a cue for visually oriented predators to locate artificial nests in a population of blue‐black grassquits Volatinia jacarina, a Neotropical passerine that exhibits a complex sexual display and is subjected to elevated rates of nest predation. We also evaluated the effect of nest substrate on survival. Predation rate was higher for nests within territories of displaying males relative to areas without displaying males and for nests placed in shrubs relative to grasses. Predation increased sharply in the third experimental replicate, at the end of the breeding season, which suggests that predators may develop a search image for nests or may become more abundant during specific periods of the season. Avian predators appear to be the most important nest predators. Results suggest that there may be a trade‐off between the increase in fitness derived from sexual displays of males to attract potential mates and the decrease owing to predation of active nests within their territories.     

Detection of Helicobacter pylori in saliva and esophagus

The route of Helicobacter pylori transmission remains unclear and the currently suggested route is person-to-person transfer by faecal-oral and oral-oral mode. The aim of this study was to verify the presence of H. pylori in esophagus and saliva of humans. Saliva samples, mucosal biopsies from esophagus, gastric antrum and fundus were collected from 19 patients with positive Urea Breath Test (UBT). Gastric biopsies were used for H. pylori colture and antimicrobial susceptibility tests whereas saliva samples were collected to detect H. pylori with a Nested-PCR targeting 16S rRNA gene as well as esophagus biopsies which were also investigated with immunohistochemical staining. Helicobacter pylori was isolated in 18 patients both in gastric antrum and fundus. The molecular analysis, confirmed by comparative sequences evaluation, gave positive results in all saliva and esophageal samples whereas the immunohistochemistry revealed the presence of H. pylori in 15.8% (3/19) of the esophagus samples. Our data suggest that saliva and esophagus may be considered reservoirs for H. pylori in humans and emphasize the need to use more susceptible techniques for H. pylori detection, in particular in over-crowded sites. Identification of the transmission route of H. pylori is crucial in developing an effective plan of surveillance by finding new means of disease management.     

Site fidelity and movements of humpback whales (Megaptera novaeangliae) on the Brazilian breeding ground,southwestern Atlantic

Site fidelity and movements were studied for humpback whales photo-identified from 1989 to 2006 in the Abrolhos Bank, southwestern Atlantic, Brazil. A total of 2,612 individuals were identified, 374 of which were observed on more than one occasion. The cumulative number of identified whales has increased since 1989. Recapture rate was low and varied among different years. A total of 33 whales was observed using the Abrolhos Bank for longer than 10 yr, up to a maximum of 16 yr. Our data suggest that different whales show distinct movement rates. Some whales used a large extent of the Abrolhos Bank region. Opportunistic photo-identification data (on the scale of the Brazilian coast from 4° to 23°S) revealed important information about stock identity. The longest distance between within-season resightings was over 600 km, while one whale was observed in two locations separated by more than 1,400 km in different years. Long-range movements within and between seasons support the single stock hypothesis for humpback whales wintering off the Brazilian coast.     

Segregation distortion and linkage analysis in eggplant (Solanum melongena L.)

An anther-derived doubled haploid (DH) population and an F2 mapping population were developed from an intraspecific hybrid between the eggplant breeding lines 305E40 and 67/3. The former incorporates an introgressed segment from Solanum aethiopicum Gilo Group carrying the gene Rfo-sa1, which confers resistance to Fusarium oxysporum; the latter is a selection from an intraspecific cross involving two conventional eggplant varieties and lacks Rfo-sa1. Initially, 28 AFLP primer combinations (PCs) were applied to a sample of 93 F2 individuals and 93 DH individuals, from which 170 polymorphic AFLP fragments were identified. In the DH population, the segregation of 117 of these AFLPs as well as markers closely linked to Rfo-sa1 was substantially distorted, while in the F2 population, segregation distortion was restricted to just 10 markers, and thus the latter was chosen for map development. A set of 141 F2 individuals was genotyped with 73 AFLP PCs (generating 406 informative markers), 32 SSRs, 4 tomato RFLPs, and 3 CAPS markers linked to Rfo-sa1. This resulted in the assignment of 348 markers to 12 major linkage groups. The framework map covered 718.7?cM, comprising 238 markers (212 AFLPs, 22 SSRs, 1 RFLP, and the Rfo-sa1 CAPS). Marker order and inter-marker distances in this eggplant map were largely consistent with those reported in a recently published SSR-based map. From an eggplant breeding perspective, DH populations produced by anther culture appear to be subject to massive segregation distortion and thus may not be very efficient in capturing the full range of genetic variation present in the parental lines.     

Unraveling the molecular basis for ligand binding in truncated hemoglobins: The trHbO Bacillus subtilis case

Truncated hemoglobins (trHbs) are heme proteins present in bacteria, unicellular eukaryotes, and higher plants. Their tertiary structure consists in a 2‐over‐2 helical sandwich, which display typically an inner tunnel/cavity system for ligand migration and/or storage. The microorganism Bacillus subtilis contains a peculiar trHb, which does not show an evident tunnel/cavity system connecting the protein active site with the solvent, and exhibits anyway a very high oxygen association rate. Moreover, resonant Raman results of CO bound protein, showed that a complex hydrogen bond network exists in the distal cavity, making it difficult to assign unambiguously the residues involved in the stabilization of the bound ligand. To understand these experimental results with atomistic detail, we performed classical molecular dynamics simulations of the oxy, carboxy, and deoxy proteins. The free energy profiles for ligand migration suggest that there is a key residue, GlnE11, that presents an alternate conformation, in which a wide ligand migration tunnel is formed, consistently with the kinetic data. This tunnel is topologically related to the one found in group I trHbs. On the other hand, the results for the CO and O₂ bound protein show that GlnE11 is directly involved in the stabilization of the cordinated ligand, playing a similar role as TyrB10 and TrpG8 in other trHbs. Our results not only reconcile the structural data with the kinetic information, but also provide additional insight into the general behaviour of trHbs. Proteins 2010. © 2009 Wiley‐Liss, Inc.