Biochemical complexity of serum HLA class I molecules

Human serum was found to contain a variety of class I-like molecules by Western blotting with anti-class I heavy chain reagents: major bands usually are observed around M _r 44 000, 40 000, and 35 000–37 000. HLA-A24-positive individuals are distinguished by higher serum levels of M _r 44 000 and 40 000 class I-like molecules than those found in HLA-A24-negative individuals. The M _r 44 000 serum molecules are probably intact class I molecules that have been shed from the cell membrane, because they contain both a transmembrane segment (TM), as deduced from detergent-binding experiments, and a cytoplasmic tail (CT), as inferred from reactivity with an antipeptide serum specific for the cytoplasmic domain of class I antigens (RaCT). The M _r 35 000 and 37 000 molecules contain neither a TM nor a CT region and therefore are probably proteolytic breakdown products of cellular and/or serum M _r 44 000 molecules, although the existence of Q10-like molecules in man cannot be ruled out. The M _r 40 000 molecules do not contain a TM region. M _r 40 000 molecules reactive with the RaCT serum were found in the minority (2/13) of sera tested. We conclude that alternative splicing resulting in a precise excision of the TM exon plays a minor role in the generation of serum HLA class I antigens.Abbreviations used in this paper B2m beta-2 microglobulin - BSA bovine serum albumin - EBV-BLCL Epstein-Barr virus-transformed B lymphoblastoid cell line - mAb monoclonal antibody - SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis - TCA trichloroacetic acid     

Polymorphism and complexity of HLA-DR: evidence for intra-HLA-DR region crossing-over events

HLA-DR molecules were isolated from HLA-DR3, –5, and –w6 positive homozygous B-cell lines by immunoprecipitation with monoclonal antibodies and analyzed by gel electrophoretic techniques. DNA isolated from the same cell lines was digested with the restriction enzyme Taq I and hybridized with a DR beta full-length cDNA probe. We demonstrated that certain DR I alleles are found in combination with different DR III alleles as defined by Southern blotting, protein chemistry, a functional assay using purified protein derivative-specific T-cell lines, and, in one case, also alloreactive T-cell reagents. Our results indicate that within the family of HLA-DRw52-associated haplotypes DR beta chain genes may have been transferred from one haplotype to another. The implications of these findings are discussed.     

Possible functional roles of cortical depsides and medullary depsidones in the foliose lichen Hypogymnia physodes

Secondary compounds were quantified in 75 thalli of the foliose lichen Hypogymnia physodes collected in habitats along a natural shade-sun gradient from dark spruce forests to sun-exposed sea cliffs. The irradiance in all the 75 lichen sites was estimated from hemispherical photographs. The content of lichen compounds per thallus area correlated positively with irradiance level (r²=0.73), and the mean concentration increased from 6.7% in the spruce forest to 14.4% on sea cliffs. The medullary depsidones, physodic, physodalic and protocetraric acids, constituted >95% of the total pool of extractable secondary compounds, the cortical depsides, atranorin and chloratranorin, represented <5%. Both cortical compounds correlated well with direct and with diffuse radiation, whereas the three medullary compounds correlated better with diffuse than with direct radiation. Mentioned trends are consistent with a solar radiation screening hypothesis of both groups of these colourless compounds occuring as tiny crystals outside fungal hyphae. However, the UV-B protective hypothesis of the compounds was further tested in a lab experiment. Unnaturally high UV-B doses were required to significantly reduce the PSII efficiency (F_V/F_M) of symbiotic algae. Removal of the major pool of secondary compounds with acetone did not increase photobiont susceptibility to UV-B. Therefore, the main functional role of the UV-B absorbing secondary compounds in H. physodes is hardly UV-B screening. Other roles such as PAR-screening and defence against herbivores and pathogenic organisms are discussed.     

Mollusc assemblages of the Pontian and Dacian deposits from the Topolog-Arge? area (southern Carpathian foredeep - Romania)

A better understanding of Mediterranean-Paratethys water-exchange during the Messinian Salinity Crisis has since long been hampered by the absence of a reliable time frame for the Paratethys. High-resolution magnetostratigraphic studies on the sedimentary sequences of the eastern and southern Carpathian foredeep recently resulted in an accurate chronology for the Mio-Pliocene deposits of the Dacic Basin of Romania. This allowed a straightforward correlation of the Pontian and Dacian stages to the geological time scale, which revived earlier discussions on Mediterranean-Paratethys connectivity. Here, we present Pontian and Dacian mollusc assemblages of the Getic Depression (Topolog-Arge? area, southern Carpathians) of Romania, which are incorporated in a magnetostratigraphic time frame. They indicate that a hiatus - comprising the Early Pontian - is present in the stratigraphic successions, which could be related to a base level drop of the Paratethys water column or to more local tectonic processes. The mollusc assemblages furthermore show a gradual transition at the Pontian/Dacian boundary, which is magnetostratigraphically dated at ∼4.9 Ma. This is significantly later (by more than 400 kyrs) than the Mio-Pliocene boundary in the Mediterranean sequences.     

SVM2Motif—Reconstructing Overlapping DNA Sequence Motifs by Mimicking an SVM Predictor

Identifying discriminative motifs underlying the functionality and evolution of organisms is a major challenge in computational biology. Machine learning approaches such as support vector machines (SVMs) achieve state-of-the-art performances in genomic discrimination tasks, but—due to its black-box character—motifs underlying its decision function are largely unknown. As a remedy, positional oligomer importance matrices (POIMs) allow us to visualize the significance of position-specific subsequences. Although being a major step towards the explanation of trained SVM models, they suffer from the fact that their size grows exponentially in the length of the motif, which renders their manual inspection feasible only for comparably small motif sizes, typically k ≤ 5. In this work, we extend the work on positional oligomer importance matrices, by presenting a new machine-learning methodology, entitled motifPOIM, to extract the truly relevant motifs—regardless of their length and complexity—underlying the predictions of a trained SVM model. Our framework thereby considers the motifs as free parameters in a probabilistic model, a task which can be phrased as a non-convex optimization problem. The exponential dependence of the POIM size on the oligomer length poses a major numerical challenge, which we address by an efficient optimization framework that allows us to find possibly overlapping motifs consisting of up to hundreds of nucleotides. We demonstrate the efficacy of our approach on a synthetic data set as well as a real-world human splice site data set.     

Gaze in Visual Search Is Guided More Efficiently by Positive Cues than by Negative Cues

Visual search can be accelerated when properties of the target are known. Such knowledge allows the searcher to direct attention to items sharing these properties. Recent work indicates that information about properties of non-targets (i.e., negative cues) can also guide search. In the present study, we examine whether negative cues lead to different search behavior compared to positive cues. We asked observers to search for a target defined by a certain shape singleton (broken line among solid lines). Each line was embedded in a colored disk. In “positive cue” blocks, participants were informed about possible colors of the target item. In “negative cue” blocks, the participants were informed about colors that could not contain the target. Search displays were designed such that with both the positive and negative cues, the same number of items could potentially contain the broken line (“relevant items”). Thus, both cues were equally informative. We measured response times and eye movements. Participants exhibited longer response times when provided with negative cues compared to positive cues. Although negative cues did guide the eyes to relevant items, there were marked differences in eye movements. Negative cues resulted in smaller proportions of fixations on relevant items, longer duration of fixations and in higher rates of fixations per item as compared to positive cues. The effectiveness of both cue types, as measured by fixations on relevant items, increased over the course of each search. In sum, a negative color cue can guide attention to relevant items, but it is less efficient than a positive cue of the same informational value.     

Study of the reactivity of quinohemoprotein alcohol dehydrogenase with heterocycle-pentacyanoferrate(III) complexes and the electron transfer path calculations

The reactivity of alcohol dehydrogenase IIG (ADH IIG) from Pseudomonas putida HK5 with new heterocycle-pentacyanoferrate(III) complexes and hexacyanoferrate(III) was determined at pH 7.2. The pentacyanoferrate(III) complexes contained imidazole, pyrazole, pyridine, their derivatives and 2-aminobenzothiazole as the sixth ligand. The largest reactivity of the complexes with ADH IIG was estimated for the complex containing pyridine. An apparent bimolecular constant (k _ox ) for this complex was 8.7 × 10⁵ M⁻¹s⁻¹. The lowest value of k _ox was estimated for the complex with benzotriazole (k _ox = 3.1 × 10⁴ M⁻¹s⁻¹). The investigation of the hexacyanoferrate(III) enzymatic reduction rate at different ionic strength gave a single negative charge of reduced ADH IIG. Docking calculations revealed two binding sites of the complexes in ADH-IIG structure. The first one is located at the entrance to the PQQ pocket, and the second is at the site of cytochrome domain. The calculations of electron transfer (ET) path indicated that the most effective ET takes place from heme to the complex docked at the entrance to the PQQ pocket. This shortest path is constructed of amino acids Ser607 and Cys606.     

Heat stress inhibits skeletal muscle hypertrophy

Heat shock proteins (Hsps) are molecular chaperones that aid in protein synthesis and trafficking and have been shown to protect cells/tissues from various protein damaging stressors. To determine the extent to which a single heat stress and the concurrent accumulation of Hsps influences the early events of skeletal muscle hypertrophy, Sprague-Dawley rats were heat stressed (42 degrees C, 15 minutes) 24 hours prior to overloading 1 plantaris muscle by surgical removal of the gastrocnemius muscle. The contralateral plantaris muscles served as controls. Heat-stressed and/or overloaded plantaris muscles were assessed for muscle mass, total muscle protein, muscle protein concentration, Type I myosin heavy chain (Type I MHC) content, as well as Hsp72 and Hsp25 content over the course of 7 days following removal of the gastrocnemius muscle. As expected, in non-heat-stressed animals, muscle mass, total muscle protein and MHC I content were significantly increased (P < 0.05) following overload. In addition, Hsp25 and Hsp72 increased significantly after 2 and 3 days of overload, respectively. A prior heat stress-elevated Hsp25 content to levels similar to those measured following overload alone, but heat stress-induced Hsp72 content was increased significantly greater than was elicited by overload alone. Moreover, overloaded muscles from animals that experienced a prior heat stress showed a lower muscle mass increase at 5 and 7 days; a reduced total muscle protein elevation at 3, 5, and 7 days; reduced protein concentration; and a diminished Type I MHC content accumulation at 3, 5, and 7 days relative to nonheat-stressed animals. These data suggest that a prior heat stress and/or the consequent accumulation of Hsps may inhibit increases in muscle mass, total muscle protein content, and Type I MHC in muscles undergoing hypertrophy.