Pep1, a Secreted Effector Protein of Ustilago maydis, Is Required for Successful Invasion of Plant Cells

The basidiomycete Ustilago maydis causes smut disease in maize. Colonization of the host plant is initiated by direct penetration of cuticle and cell wall of maize epidermis cells. The invading hyphae are surrounded by the plant plasma membrane and proliferate within the plant tissue. We identified a novel secreted protein, termed Pep1, that is essential for penetration. Disruption mutants of pep1 are not affected in saprophytic growth and develop normal infection structures. However, Δpep1 mutants arrest during penetration of the epidermal cell and elicit a strong plant defense response. Using Affymetrix maize arrays, we identified 116 plant genes which are differentially regulated in Δpep1 compared to wild type infections. Most of these genes are related to plant defense. By in vivo immunolocalization, live-cell imaging and plasmolysis approaches, we detected Pep1 in the apoplastic space as well as its accumulation at sites of cell-to-cell passages. Site-directed mutagenesis identified two of the four cysteine residues in Pep1 as essential for function, suggesting that the formation of disulfide bridges is crucial for proper protein folding. The barley covered smut fungus Ustilago hordei contains an ortholog of pep1 which is needed for penetration of barley and which is able to complement the U. maydis Δpep1 mutant. Based on these results, we conclude that Pep1 has a conserved function essential for establishing compatibility that is not restricted to the U. maydis / maize interaction

Global patterns of declining temperature variability from the Last Glacial Maximum to the Holocene

Changes in climate variability are as important for society to address as are changes in mean climate1. Contrasting temperature variability during the Last Glacial Maximum and the Holocene can provide insights into the relationship between the mean state of the climate and its variability2,3. However, although glacial–interglacial changes in variability have been quantified for Greenland2, a global view remains elusive. Here we use a network of marine and terrestrial temperature proxies to show that temperature variability decreased globally by a factor of four as the climate warmed by 3–8 degrees Celsius from the Last Glacial Maximum (around 21,000 years ago) to the Holocene epoch (the past 11,500 years). This decrease had a clear zonal pattern, with little change in the tropics (by a factor of only 1.6–2.8) and greater change in the mid-latitudes of both hemispheres (by a factor of 3.3–14). By contrast, Greenland ice-core records show a reduction in temperature variability by a factor of 73, suggesting influences beyond local temperature or a decoupling of atmospheric and global surface temperature variability for Greenland. The overall pattern of reduced variability can be explained by changes in the meridional temperature gradient, a mechanism that points to further decreases in temperature variability in a warmer future

In the Laboratory and during Free-Flight: Old Honey Bees Reveal Learning and Extinction Deficits that Mirror Mammalian Functional Decline

Loss of brain function is one of the most negative and feared aspects of aging. Studies of invertebrates have taught us much about the physiology of aging and how this progression may be slowed. Yet, how aging affects complex brain functions, e.g., the ability to acquire new memory when previous experience is no longer valid, is an almost exclusive question of studies in humans and mammalian models. In these systems, age related cognitive disorders are assessed through composite paradigms that test different performance tasks in the same individual. Such studies could demonstrate that afflicted individuals show the loss of several and often-diverse memory faculties, and that performance usually varies more between aged individuals, as compared to conspecifics from younger groups. No comparable composite surveying approaches are established yet for invertebrate models in aging research. Here we test whether an insect can share patterns of decline similar to those that are commonly observed during mammalian brain aging. Using honey bees, we combine restrained learning with free-flight assays. We demonstrate that reduced olfactory learning performance correlates with a reduced ability to extinguish the spatial memory of an abandoned nest location (spatial memory extinction). Adding to this, we show that learning performance is more variable in old honey bees. Taken together, our findings point to generic features of brain aging and provide the prerequisites to model individual aspects of learning dysfunction with insect models

Thermodynamic Selection of Steric Zipper Patterns in the Amyloid Cross-β Spine

At the core of amyloid fibrils is the cross-β spine, a long tape of β-sheets formed by the constituent proteins. Recent high-resolution x-ray studies show that the unit of this filamentous structure is a β-sheet bilayer with side chains within the bilayer forming a tightly interdigitating “steric zipper” interface. However, for a given peptide, different bilayer patterns are possible, and no quantitative explanation exists regarding which pattern is selected or under what condition there can be more than one pattern observed, exhibiting molecular polymorphism. We address the structural selection mechanism by performing molecular dynamics simulations to calculate the free energy of incorporating a peptide monomer into a β-sheet bilayer. We test filaments formed by several types of peptides including GNNQQNY, NNQQ, VEALYL, KLVFFAE and STVIIE, and find that the patterns with the lowest binding free energy correspond to available atomistic structures with high accuracy. Molecular polymorphism, as exhibited by NNQQ, is likely because there are more than one most stable structures whose binding free energies differ by less than the thermal energy. Detailed analysis of individual energy terms reveals that these short peptides are not strained nor do they lose much conformational entropy upon incorporating into a β-sheet bilayer. The selection of a bilayer pattern is determined mainly by the van der Waals and hydrophobic forces as a quantitative measure of shape complementarity among side chains between the β-sheets. The requirement for self-complementary steric zipper formation supports that amyloid fibrils form more easily among similar or same sequences, and it also makes parallel β-sheets generally preferred over anti-parallel ones. But the presence of charged side chains appears to kinetically drive anti-parallel β-sheets to form at early stages of assembly, after which the bilayer formation is likely driven by energetics

Genomic Characterization of Haemophilus parasuis SH0165, a Highly Virulent Strain of Serovar 5 Prevalent in China

Haemophilus parasuis can be either a commensal bacterium of the porcine respiratory tract or an opportunistic pathogen causing Glässer's disease, a severe systemic disease that has led to significant economical losses in the pig industry worldwide. We determined the complete genomic sequence of H. parasuis SH0165, a highly virulent strain of serovar 5, which was isolated from a hog pen in North China. The single circular chromosome was 2,269,156 base pairs in length and contained 2,031 protein-coding genes. Together with the full spectrum of genes detected by the analysis of metabolic pathways, we confirmed that H. parasuis generates ATP via both fermentation and respiration, and possesses an intact TCA cycle for anabolism. In addition to possessing the complete pathway essential for the biosynthesis of heme, this pathogen was also found to be well-equipped with different iron acquisition systems, such as the TonB system and ABC-type transport complexes, to overcome iron limitation during infection and persistence. We identified a number of genes encoding potential virulence factors, such as type IV fimbriae and surface polysaccharides. Analysis of the genome confirmed that H. parasuis is naturally competent, as genes related to DNA uptake are present. A nine-mer DNA uptake signal sequence (ACAAGCGGT), identical to that found in Actinobacillus pleuropneumoniae and Mannheimia haemolytica, followed by similar downstream motifs, was identified in the SH0165 genome. Genomic and phylogenetic comparisons with other Pasteurellaceae species further indicated that H. parasuis was closely related to another swine pathogenic bacteria A. pleuropneumoniae. The comprehensive genetic analysis presented here provides a foundation for future research on the metabolism, natural competence and virulence of H. parasuis

Innate immunity against HIV: a priority target for HIV prevention research

This review summarizes recent advances and current gaps in understanding of innate immunity to human immunodeficiency virus (HIV) infection, and identifies key scientific priorities to enable application of this knowledge to the development of novel prevention strategies (vaccines and microbicides). It builds on productive discussion and new data arising out of a workshop on innate immunity against HIV held at the European Commission in Brussels, together with recent observations from the literature

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)1.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field