Rooting and dating maples (Acer) with an uncorrelated-rates molecular clock: implications for north American/Asian disjunctions

Simulations suggest that molecular clock analyses can correctly identify the root of a tree even when the clock assumption is severely violated. Clock-based rooting of phylogenies may be particularly useful when outgroup rooting is problematic. Here, we explore relaxed-clock rooting in the Acer/Dipteronia clade of Sapindaceae, which comprises genera of highly uneven species richness and problematic mutual monophyly. Using an approach that does not presuppose rate autocorrelation between ancestral and descendant branches and hence does not require a rooted a priori topology, we analyzed data from up to seven chloroplast loci for some 50 ingroup species. For comparison, we used midpoint and outgroup rooting and dating methods that rely on rooted input trees, namely penalized likelihood, a Bayesian autocorrelated-rates model, and a strict clock. The chloroplast sequences used here reject a single global substitution rate, and the assumption of autocorrelated rates was also rejected. The root was placed between Acer and Dipteronia by all three rooting methods, albeit with low statistical support. Analyses of Acer diversification with a lineage-through-time plot and different survival models, although sensitive to missing data, suggest a gradual decrease in the average diversification rate. The nine North American species of Acer diverged from their nearest relatives at widely different times: eastern American Acer diverged in the Oligocene and Late Miocene; western American species in the Late Eocene and Mid Miocene; and the Acer core clade, including A. saccharum, dates to the Miocene. Recent diversification in North America is strikingly rare compared to diversification in eastern Asia.     

Homologous recombination in real time: DNA strand exchange by RecA

Homologous recombination, the exchange of strands between different DNA molecules, is essential for proper maintenance and accurate duplication of the genome. Using magnetic tweezers, we monitor RecA-driven homologous recombination of individual DNA molecules in real time. We resolve several key aspects of DNA structure during and after strand exchange. Changes in DNA length and twist yield helical parameters for the protein-bound three-stranded structure in conditions in which ATP was not hydrolyzed. When strand exchange was completed under ATP hydrolysis conditions that allow protein dissociation, a "D wrap" structure formed. During homologous recombination, strand invasion at one end and RecA dissociation at the other end occurred at the same rate, and our single-molecule analysis indicated that a region of only about 80 bp is actively involved in the synapsis at any time during the entire reaction involving a long ( approximately 1 kb) region of homology.     

The alternative sigma factor sigma B of Staphylococcus aureus modulates virulence in experimental central venous catheter-related infections

The impact of the alternative sigma factor sigma B (SigB) on pathogenesis of Staphylococcus aureus is not conclusively clarified. In this study, a central venous catheter (CVC) related model of multiorgan infection was used to investigate the role of SigB for the pathogenesis of S. aureus infections and biofilm formation in vivo. Analysis of two SigB-positive wild-type strains and their isogenic mutants revealed uniformly that the wild-type was significantly more virulent than the SigB-deficient mutant. The observed difference in virulence was apparently not linked to the capability of the strains to form biofilms in vivo since wild-type and mutant strains were able to produce biofilm layers inside of the catheter. The data strongly indicate that the alternative sigma factor SigB plays a role in CVC-associated infections caused by S. aureus.     

Expressed sequence tag-derived microsatellite markers of perennial ryegrass (Lolium perenne L.)

An expressed sequence tag (EST) library of the key grassland species perennial ryegrass (Lolium perenne L.) has been exploited as a resource for microsatellite marker development. Out of 955 simple sequence repeat (SSR) containing ESTs, 744 were used for primer design. Primer amplification was tested in eight genotypes of L. perenne and L. multiflorum representing (grand-) parents of four mapping populations and resulted in 464 successfully amplified EST-SSRs. Three hundred and six primer pairs successfully amplified products in the mapping population VrnA derived from two of the eight genotypes included in the original screening and revealed SSR polymorphisms for 143 ESTs. Here, we report on 464 EST-derived SSR primer sequences of perennial ryegrass established in laboratory assays, providing a dedicated tool for marker assisted breeding and comparative mapping within and among forage and turf grasses. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Isoaspartate-containing amyloid precursor protein-derived peptides alter efficacy and specificity of potential beta-secretases

Neuritic plaques of Alzheimer patients are composed of multiple protein components. Among them, the amyloid beta-peptides (Abeta) 1-40/42 and further N- and C-terminally modified fragments of Abeta are highly abundant. Most prominent are the isoaspartate (isoAsp)-Abeta peptides and pyroglutamyl (pGlu)-Abeta. While pGlu-Abeta can only be formed from an N-terminal glutamate by glutaminyl cyclase, spontaneous isoAsp-isomerization cannot occur at an N-terminal aspartate of peptides. This means that isoAsp-Abeta formation must precede proteolysis of the amyloid precursor protein (APP). Abeta generation from APP by beta- and gamma-secretases initiates the amyloid peptide aggregation and deposition process. Two aspartate proteases have been identified as secretases: BACE-1 (beta-site amyloid precursor protein cleaving enzyme) and the intramembrane gamma-secretase multiprotein complex. However, recent evidence supports more than one beta-secretase initiating this cascade. Formation of Abeta1-40/42 was predominantly studied by expression of mutated human APP sequences in cell culture and transgenic animals, generating Abeta fragments that did not contain such multiple posttranslational modifications as in Alzheimer's disease. This prompted us to investigate the catalytic turnover of Asp- or isoAsp-containing APP-derived peptide sequences by BACE-1 and cathepsin B, another potential beta-secretase. While cathepsin B is more effective than BACE-1 in processing the Asp-containing peptide derivatives, only cathepsin B can cleave the isoAsp-containing peptides, which occurs with high catalytic efficiency.     

Non-regulatory CD8+CD45RO+CD25+ T-lymphocytes may compensate for the loss of antigen-inexperienced CD8+CD45RA+ T-cells in old age

The age-related decline in immune system functions is responsible for the increased prevalence of infectious diseases and the low efficacy of vaccination in elderly individuals. In particular, the number of peripheral naive T-cells declines throughout life and they exhibit severe functional defects at advanced age. However, we have recently identified a non-regulatory CD8+CD45RO+ CD25+ T-cell subset that occurs in a subgroup of healthy elderly individuals, who still exhibit an intact humoral immune response following influenza vaccination. Here, we demonstrate that CD8+CD45RO+CD25+ T-cells share phenotypic and functional characteristics with naive CD8+CD45RA+CD28+ T-cells from young individuals, despite their expression of CD45RO. CD8+CD45RO+ CD25+ T-cells also have long telomeres and upon antigenic challenge, they efficiently expand in vitro and differentiate into functional effector cells. The expanded population also maintains a diverse T-cell receptor repertoire. In conclusion, CD8+CD45RO+CD25+ T-cells from elderly individuals compensate for the loss of functional naive T-cells and may therefore be used as a marker of immunological competence in old age.     

Effects of regular and irregular temporal patterns of disturbance on biomass accrual and species composition of a subtidal hard-bottom assemblage

Assessing patterns of species distribution and abundance is important to understand the driving processes of, and predict future changes in, biodiversity. To this date, ecological studies have been mainly designed to investigate the effects of the mean magnitude of predictor variables, although ecological factors naturally vary in space and time. In a nine month long field experiment, we tested the effects of different temporal patterns (regular, lowly and highly irregular) in biomass removal (=disturbance event) on the diversity, species composition, and biomass accrual of macrobenthic assemblages grown on 15 × 15 cm² PVC-panels. For each pattern of disturbance, disturbance events were timed at three sequences to control for possible confounding effects with recruitment patterns. Disturbance intensity was kept identical among treatments. Assemblages developed in the absence of disturbance for 3 months prior to a 150-day manipulation period, during which the biomass from 20% of the panel area was removed at each of ten disturbance events. Additional undisturbed settlement panels were deployed in the field to assess monthly recruitment rates and species succession over a one year period. Disturbance (i) reduced biomass and total species cover, (ii) changed species composition during the first half of the manipulation period significantly, and (iii) was without effect on species richness and evenness. Irregular disturbance regimes enhanced the abundance of the ascidian Ciona intestinalis, biomass accrual, and total species cover of assemblages relative to the regular disturbance regime, but had either no or only transient effects on diversity and species composition, respectively. Neither the degree of irregularity in disturbance nor the sequence of disturbance events affected any of the response variables significantly. Recruitment of species was strongly seasonal with almost only diatoms recruiting during winter, while recruitment was most intense during summer. Our results suggest that the temporal patterns of predictor variables might be of low explanatory power for the variance of responses in communities with seasonal recruitment patterns that are exposed to a high level of disturbance. Thus the need to include temporal patterns of predictor variables in experimental designs may depend on community dynamics and the characteristics of the process under investigation. S. Wollgast and M. Molis contributed equally to this work.     

The type I TGF-beta receptor engages TRAF6 to activate TAK1 in a receptor kinase-independent manner

Transforming growth factor-beta (TGF-beta) is a multifunctional cytokine that regulates embryonic development and tissue homeostasis; however, aberrations of its activity occur in cancer. TGF-beta signals through its Type II and Type I receptors (TbetaRII and TbetaRI) causing phosphorylation of Smad proteins. TGF-beta-associated kinase 1 (TAK1), a member of the mitogen-activated protein kinase kinase kinase (MAPKKK) family, was originally identified as an effector of TGF-beta-induced p38 activation. However, the molecular mechanisms for its activation are unknown. Here we report that the ubiquitin ligase (E3) TRAF6 interacts with a consensus motif present in TbetaRI. The TbetaRI-TRAF6 interaction is required for TGF-beta-induced autoubiquitylation of TRAF6 and subsequent activation of the TAK1-p38/JNK pathway, which leads to apoptosis. TbetaRI kinase activity is required for activation of the canonical Smad pathway, whereas E3 activity of TRAF6 regulates the activation of TAK1 in a receptor kinase-independent manner. Intriguingly, TGF-beta-induced TRAF6-mediated Lys 63-linked polyubiquitylation of TAK1 Lys 34 correlates with TAK1 activation. Our data show that TGF-beta specifically activates TAK1 through interaction of TbetaRI with TRAF6, whereas activation of Smad2 is not dependent on TRAF6.     

Helminths and allergy: the example of tropomyosin

Parasitic worms contain potent allergens, but epidemiological and experimental studies suggest that infections with certain helminths are negatively associated with the prevalence of allergic diseases. This seeming contradiction can be addressed by using filarial tropomyosin as an example. This protein shares structural features and crossreacting B-cell epitopes with other highly allergenic invertebrate tropomyosins. Nevertheless, it usually does not provoke allergic disease in infected individuals. In addition, it is one of the most prominent candidates for an anti-nematode vaccine. Recent data suggest mechanisms that might prevent hosts from developing allergic reactions against allergens of their parasites, such as filarial tropomyosin.     

High Functional Diversity in Mycobacterium tuberculosis Driven by Genetic Drift and Human Demography

Mycobacterium tuberculosis infects one third of the human world population and kills someone every 15 seconds. For more than a century, scientists and clinicians have been distinguishing between the human- and animal-adapted members of the M. tuberculosis complex (MTBC). However, all human-adapted strains of MTBC have traditionally been considered to be essentially identical. We surveyed sequence diversity within a global collection of strains belonging to MTBC using seven megabase pairs of DNA sequence data. We show that the members of MTBC affecting humans are more genetically diverse than generally assumed, and that this diversity can be linked to human demographic and migratory events. We further demonstrate that these organisms are under extremely reduced purifying selection and that, as a result of increased genetic drift, much of this genetic diversity is likely to have functional consequences. Our findings suggest that the current increases in human population, urbanization, and global travel, combined with the population genetic characteristics of M. tuberculosis described here, could contribute to the emergence and spread of drug-resistant tuberculosis.