Insights into early mycobacterial pathogenesis from the zebrafish

Here we discuss the application of the zebrafish as a relatively new model host for the study of mycobacterial pathogenesis. Recent advances in our understanding of host-mycobacteria interactions from the zebrafish include insights into the role of the innate immune system in both controlling and facilitating infection. Analysis in the zebrafish has revealed that innate macrophages restrict initial bacterial growth, but also convey infecting bacteria into the granuloma, which serves as a place for bacterial growth and spread. Bacterial virulence determinants interact with these processes at different steps in pathogenesis, which can be dissected in these living see-through hosts. As these studies uncover new facets of the bacteria-host interactions in tuberculosis they raise even more questions for future investigation.     

The Early Stone Age lithic assemblages of Gadeb (Ethiopia) and the Developed Oldowan/early Acheulean in East Africa

The Early Stone Age sites of Gadeb (Ethiopian South-East Plateau) were excavated under the direction of Desmond Clark in the 1970s. Dated to between 1.45 and 0.7 Ma, Gadeb proved that humans had already occupied high altitude areas in the Lower Pleistocene. Despite the importance of the Gadeb sites, their lithic assemblages were never published in detail, and no review of the stone tools has ever been reported since the original 1970s study. This paper updates the information available on Gadeb by presenting a systematic review of the lithic technology of several assemblages. The objectives are to evaluate the technological skills of Gadeb knappers and to contextualize them into the current discussion of the origins of the Acheulean and its possible coexistence with the so-called Developed Oldowan in East Africa.     

Landscape-scale variation in hominin tool use: Evidence from the Developed Oldowan

The relationship between artifact manufacture, use, and discard in the Developed Oldowan is complex. Here we use digital-image-analysis techniques to investigate the intensity of reduction in single-platform cores of the Developed Oldowan of the Okote Member, Koobi Fora Formation. Data suggest that this method provides a more accurate measure of reduction intensity than previous applications of a unifacial-scraper model. Assemblages of single-platform cores excavated from extensive lateral exposures of the Okote Member provide insights into the relationship between raw-material availability and discard patterns. Variation in reduction intensity suggests that tools are not always discarded in patterns that would be predicted by the availability of raw material. Further, it appears that hominin transport decisions involved an assessment of the potential use-life of certain forms. Many aspects of Developed Oldowan technology conform to previously developed models of curated technologies.     

Palynology of OGS-6a and OGS-7, two new 2.6 Ma archaeological sites from Gona, Afar, Ethiopia: Insights on aspects of Late Pliocene habitats and the beginnings of stone-tool use

Fossil pollen grains extracted from sediments sampled from OGS-6a and OGS-7, two newly excavated Late Pliocene sites of Gona, in the Afar Depression of Ethiopia, indicate a high percentage of afromontane forest and highland taxa (36.8-43.3%), among which Podocarpus cf. gracilor is dominant (19.6-24.2%). Forest taxa typical of the afromontane phytogeographic region have also been identified in the samples. The overall pollen spectrum suggests a mosaic of open and closed habitats, with a considerable wooded environment present during the time associated with the beginnings of early hominid use of flaked stones at Gona. Here we provide details of the results from the pollen analyses. The two sites lie 4-8 m directly below a volcanic tuff dated by ⁴⁰Ar/³⁹Ar to 2.53 ± 0.15 million years (Ma). The materials were recovered within fine-grained sediments located just above a geomagnetic polarity transition identified as the Gauss-Matuyama boundary dated close to 2.6 Ma, also corroborating the earlier age reported for the East Gona artifacts from EG-10 and EG-12. Thus, the stone artifacts, fossilized broken fauna and pollen grains from OGS-6a and OGS-7 represent archaeological materials from the world's oldest securely dated sites, and offer insights on aspects of the palaeoenvironments around the time of the beginnings of ancestral hominid use of flaked stones, ca. 2.6 Ma.     

Tool-using strategies by early hominids at bed II, Olduvai Gorge, Tanzania

This study examines the current prevailing model of Oldowan technology-the opportunistic, least-effort strategy of stone tool making and using by early hominids. The sample includes the MNK chert factory site and three contemporaneous assemblages from Olduvai Gorge, all dated between 1.65 and 1.53 m.y.a. The analysis suggests that early hominids at Olduvai may have been selective, applying distinctive strategies in making and using tools depending on the different types of raw materials available to them. The preponderance of lava cores and near absence of flakes associated with the cores suggest that lava cores at Olduvai did not provide a source of flakes. They were primarily heavy-duty core tools, despite the fact that the majority of Olduvai lava is of excellent quality for flaking. Contrary to this pattern, the abundance of chert flakes and the lack of large chert cores suggest that the production of flakes was the most important strategy applied to chert. Original forms and flaking mechanics of the raw materials may have been important factors in the simultaneous application of the different, complementary strategies. The Oldowan tool-using strategy was dynamic and flexible, in response to changes in raw material availability. The use of chert between 1.65 and 1.53 m.y.a. was apparently related to the drastic decrease in flake production in lava and quartz. Finally, lack of initial reduction episodes of lava material challenges the idea of the stone cache strategy at Olduvai between 1.65 to 1.53 m.y.a.     

Consequences of early adverse rearing experience (EARE) on development: insights from non-human primate studies

Early rearing experiences are important in one's whole life,whereas early adverse rearing experience (EARE) is usually related to various physical and mental disorders in later life.Although there were many studies on human and animals,regarding the effect of EARE on brain development,neuroendocrine systems,as well as the consequential mental disorders and behavioral abnormalities,the underlying mechanisms remain unclear.Due to the close genetic relationship and similarity in social organizations with humans,non-human primate (NHP) studies were performed for over 60 years.Various EARE models were developed to disrupt the early normal interactions between infants and mothers or peers.Those studies provided important insights of EARE induced effects on the physiological and behavioral systems of NHPs across life span,such as social behaviors (including disturbance behavior,social deficiency,sexual behavior,etc),learning and memory ability,brain structural and functional developments (including influences on neurons and glia cells,neuroendocrine systems,e.g.,hypothalamic-pituitary-adrenal (HPA) axis,etc).In this review,the effects of EARE and the underlying epigenetic mechanisms were comprehensively summarized and the possibility of rehabilitation was discussed.     

Structural Insights into Serine-rich Fimbriae from Gram-positive Bacteria

The serine-rich repeat family of fimbriae play important roles in the pathogenesis of streptococci and staphylococci. Despite recent attention, their finer structural details and precise adhesion mechanisms have yet to be determined. Fap1 (Fimbriae-associated protein 1) is the major structural subunit of serine-rich repeat fimbriae from Streptococcus parasanguinis and plays an essential role in fimbrial biogenesis, adhesion, and the early stages of dental plaque formation. Combining multidisciplinary, high resolution structural studies with biological assays, we provide new structural insight into adhesion by Fap1. We propose a model in which the serine-rich repeats of Fap1 subunits form an extended structure that projects the N-terminal globular domains away from the bacterial surface for adhesion to the salivary pellicle. We also uncover a novel pH-dependent conformational change that modulates adhesion and likely plays a role in survival in acidic environments.     

Insights into neural stem cell biology from flies

Drosophila neuroblasts are similar to mammalian neural stem cells in their ability to self-renew and to produce many different types of neurons and glial cells. In the past two decades, great advances have been made in understanding the molecular mechanisms underlying embryonic neuroblast formation, the establishment of cell polarity and the temporal regulation of cell fate. It is now a challenge to connect, at the molecular level, the different cell biological events underlying the transition from neural stem cell maintenance to differentiation. Progress has also been made in understanding the later stages of development, when neuroblasts become mitotically inactive, or quiescent, and are then reactivated postembryonically to generate the neurons that make up the adult nervous system. The ability to manipulate the steps leading from quiescence to proliferation and from proliferation to differentiation will have a major impact on the treatment of neurological injury and neurodegenerative disease.     

Insights into gene expression and packaging from computer simulations

Within the nucleus of each cell lies DNA—an unfathomably long, twisted, and intricately coiled molecule—segments of which make up the genes that provide the instructions that a cell needs to operate. As we near the 60th anniversary of the discovery of the DNA double helix, crucial questions remain about how the physical arrangement of the DNA in cells affects how genes work. For example, how a cell stores the genetic information inside the nucleus is complicated by the necessity of maintaining accessibility to DNA for genetic processing. In order to gain insight into the roles played by various proteins in reading and compacting the genome, we have developed new methodologies to simulate the dynamic, three-dimensional structures of long, fluctuating, protein-decorated strands of DNA. Our a priori approach to the problem allows us to determine the effects of individual proteins and their chemical modifications on overall DNA structure and function. Here, we present our recent treatment of the communication between regulatory proteins attached to precisely constructed stretches of chromatin. Our simulations account for the enhancement in communication detected experimentally on chromatin compared to protein-free DNA of the same chain length, as well as the critical roles played by the cationic ‘tails’ of the histone proteins in this signaling. The states of chromatin captured in the simulations offer new insights into the ways that the DNA, histones, and regulatory proteins contribute to long-range communication along the genome.     

Insights into ALS pathomechanisms: from flies to humans

Amyotrophic Lateral Sclerosis (ALS) is a devastating neurodegenerative disease causing the death of motor neurons with consequent muscle atrophy and paralysis. Several neurodegenerative diseases have been modeled in Drosophila and genetic studies on this model organism led to the elucidation of crucial aspects of disease mechanisms. ALS, however, has lagged somewhat behind possibly because of the lack of a suitable genetic model. We were the first to develop a fly model for ALS and over the last few years, we have implemented and used this model for a large scale, unbiased modifier screen. We also report an extensive bioinformatic analysis of the genetic modifiers and we show that most of them are associated in a network of interacting genes controlling known as well as novel cellular processes involved in ALS pathogenesis. A similar analysis for the human homologues of the Drosophila modifiers and the validation of a subset of them in human tissues confirm and expand the significance of the data for the human disease. Finally, we analyze a possible application of the model in the process of therapeutic discovery in ALS and we discuss the importance of novel “non-obvious” models for the disease.     

Insights into transport mechanism from LeuT engineered to transport tryptophan

LeuT is a bacterial homologue of the neurotransmitter:sodium symporter (NSS) family and, being the only NSS member to have been structurally characterized by X-ray crystallography, is a model protein for studying transporter structure and mechanism. Transport activity in LeuT was hypothesized to require structural transitions between open-to-out and occluded conformations dependent upon protein:ligand binding complementarity. Here, using crystallographic and functional analysis, we show that binding site modification produces changes in both structure and activity that are consistent with complementarity-dependent structural transitions to the occluded state. The mutation I359Q converts the activity of tryptophan from inhibitor to transportable substrate. This mutation changes the local environment of the binding site, inducing the bound tryptophan to adopt a different conformer than in the wild-type complex. Instead of trapping the transporter open, tryptophan binding now allows the formation of an occluded state. Thus, transport activity is correlated to the ability of the ligand to promote the structural transition to the occluded state, a step in the transport cycle that is dependent on protein:ligand complementarity in the central binding site.     

Insights into plant metabolic networks from steady-state metabolic flux analysis

Steady-state metabolic flux analysis (MFA) is an experimental approach that allows the measurement of multiple fluxes in the core network of primary carbon metabolism. It is based on isotopic labelling experiments, and although well established in the analysis of micro-organisms, and some mammalian systems, the extension of the method to plant cells has been challenging because of the extensive subcellular compartmentation of the metabolic network. Despite this difficulty there has been substantial progress in developing robust protocols for the analysis of heterotrophic plant metabolism by steady-state MFA, and flux maps have now been published that reflect the metabolic phenotypes of excised root tips, developing embryos and cotyledons, hairy root cultures, and cell suspensions under a variety of physiological conditions. There has been a steady improvement in the quality, extent and statistical reliability of these analyses, and new information is emerging on the performance of the plant metabolic network and the contributions of specific pathways.     

Insights into fruit function from the proteome of the hypanthium

Apple (Malus×domestica Borkh.) was used as a model to studying essential biological processes occurring in mature fruit hypanthium, commonly referred to as the fruit flesh or pulp, a highly active tissue where numerous metabolic processes such as carbohydrate metabolism and signal transduction occur. To understand the complex biological processes occurring in the hypanthium, a proteomics approach was used to analyze the proteome from freshly harvested ripe apple fruits. A total of 290 well-resolved spots were detected using two-dimensional gel electrophoresis (2-DE). Out of these, 216 proteins were identified representing 116 non-redundant proteins using matrix-assisted laser-desorption ionisation time-of-flight mass spectrometry (MALDI-TOF MS) and either the MASCOT or ProteinProspector engine for peptide mass fingerprinting (PMF) database searching. Identified proteins were classified into 13 major functional categories. Among these, the energy metabolism class was the most represented and included 50% of proteins homologous to Arabidopsis proteins that are involved in the response to biotic and abiotic stresses, suggesting a dual role for these proteins in addition to energy metabolism. We also identified dynein heavy chain in the hypanthium although this protein has been proposed as absent from angiosperms and thus suggest that the lack of dyneins in higher plants studied to date may not be a general characteristic to angiosperm genomic organisation. We therefore conclude that the detection and elucidation of the apple hypanthium proteome is an indispensable step towards the comprehension of fruit metabolism, the integration of genomic, proteomic and metabolomic data to agronomic trait information and thus fruit quality improvements.     

Insights into Coupled Folding and Binding Mechanisms from Kinetic Studies

Intrinsically disordered proteins (IDPs) are characterized by a lack of persistent structure. Since their identification more than a decade ago, many questions regarding their functional relevance and interaction mechanisms remain unanswered. Although most experiments have taken equilibrium and structural perspectives, fewer studies have investigated the kinetics of their interactions. Here we review and highlight the type of information that can be gained from kinetic studies. In particular, we show how kinetic studies of coupled folding and binding reactions, an important class of signaling event, are needed to determine mechanisms.     

Insights into buforin II membrane translocation from molecular dynamics simulations

Buforin II is a histone-derived antimicrobial peptide that readily translocates across lipid membranes without causing significant membrane permeabilization. Previous studies showed that mutating the sole proline of buforin II dramatically decreases its translocation. As well, researchers have proposed that the peptide crosses membranes in a cooperative manner by forming transient toroidal pores. This paper reports molecular dynamics simulations designed to investigate the structure of buforin II upon membrane entry and evaluate whether the peptide is able to form toroidal pore structures. These simulations showed a relationship between protein–lipid interactions and increased structural deformations of the buforin N-terminal region promoted by proline. Moreover, simulations with multiple peptides show how buforin II can embed deeply into membranes and potentially form toroidal pores. Together, these simulations provide structural insight into the translocation process for buforin II in addition to providing more general insight into the role proline can play in antimicrobial peptides.