Cestode regulation of inflammation and inflammatory diseases

Helminth parasites are masters of immune regulation; a likely prerequisite for long-term survival by circumventing their hosts’ attempt to eradicate them. From a translational perspective, knowledge of immune events as a response to infection with a helminth parasite could be used to reduce the intensity of unwanted inflammatory reactions. Substantial data have accumulated showing that inflammatory reactions that promote a variety of auto-inflammatory diseases are dampened as a consequence of infection with helminth parasites, via either the mobilization of an anti-worm spectrum of immune events or by the direct effect of secretory/excretory bioactive immunomodulatory molecules released from the parasite. However, many issues are outstanding in the definition of the mechanism(s) by which infection with helminth parasites can affect the outcome, positively or negatively, of concomitant disease. We focus on a subgroup of this complex group of metazoan parasites, the cestodes, summarizing studies from rodent models that illustrate if, and by what mechanisms, infection with tapeworms ameliorate or exaggerate disease in their host. The ability of infection with cestodes, or other classes of helminth, to worsen a disease course or confer susceptibility to intracellular pathogens should be carefully considered in the context of ‘helminth therapy’. In addition, poorly characterised cestode extracts can regulate murine and human immunocyte function, yet the impact of these in the context of autoimmune or allergic diseases is poorly understood. Thus, studies with cestodes, as representative helminths, have helped cement the concept that infection with parasitic helminths can inhibit concomitant disease; however, issues relating to long-term effects, potential side-effects, mixed pathogen infections and purification of immunomodulatory molecules from the parasite remain as challenges that need to be addressed in order to achieve the use of helminths as anti-inflammatory agents for human diseases.     

Limited Clinical Utility of Non-invasive Prenatal Testing for Subchromosomal Abnormalities

The use of massively parallel sequencing of maternal cfDNA for non-invasive prenatal testing (NIPT) of aneuploidy is widely available. Recently, the scope of testing has increased to include selected subchromosomal abnormalities, but the number of samples reported has been small. We developed a calling pipeline based on a segmentation algorithm for the detection of these rearrangements in maternal plasma. The same read depth used in our standard pipeline for aneuploidy NIPT detected 15/18 (83%) samples with pathogenic rearrangements > 6 Mb but only 2/10 samples with rearrangements < 6 Mb, unless they were maternally inherited. There were two false-positive calls in 534 samples with no known subchromosomal abnormalities (specificity 99.6%). Using higher read depths, we detected 29/31 fetal subchromosomal abnormalities, including the three samples with maternally inherited microduplications. We conclude that test sensitivity is a function of the fetal fraction, read depth, and size of the fetal CNV and that at least one of the two false negatives is due to a low fetal fraction. The lack of an independent method for determining fetal fraction, especially for female fetuses, leads to uncertainty in test sensitivity, which currently has implications for this technique’s future as a clinical diagnostic test. Furthermore, to be effective, NIPT must be able to detect chromosomal rearrangements across the whole genome for a very low false-positive rate. Because standard NIPT can only detect the majority of larger (>6 Mb) chromosomal rearrangements and requires knowledge of fetal fraction, we consider that it is not yet ready for routine clinical implementation.     

Transforming growth factor alpha.

Transforming growth factor alpha (TGFα) is a close relative of epidermal growth factor (EGF), the first polypeptide mitogen discovered in 1962 (Cohen, 1962). TGFα, like EGF, exerts its effect on cells through binding to the EGF-Receptor (EGF-R). Here we review the molecular and cell biology of TGFα before proceeding to describe our own work on signaling molecules induced in response to activation of the EGF-R.     

Control of Internode Length in Pisum sativum (Further Evidence for the Involvement of Indole-3-Acetic Acid)

The effects of altered endogenous indole-3-acetic (IAA) levels on elongation in garden pea (Pisum sativum L.) plants were investigated. The auxin transport inhibitors 2,3,5-triiodobenzoic acid (TIBA) and 9-hydroxyfluorene-9-carboxylic acid (HFCA) were applied to elongating internodes of wild-type and mutant lkb plants. The lkb mutant was included because elongating lkb internodes contained 2- to 3-fold less free IAA than those of the wild type. In the wild type, TIBA reduced both the IAA level and internode elongation below the site of application. Both TIBA and HFCA strongly promoted the elongation of lkb internodes and also raised IAA levels above the application site. The synthetic auxin 2,4-dichlorophenoxyacetic acid (2,4-D) also markedly increased internode elongation in lkb plants and virtually restored petioles and tendrils to their wild-type length. In contrast, treatment of wild-type plants with TIBA, HFCA, or 2,4-D caused little or no increase in elongation above the application site. The ethylene synthesis inhibitor aminoethoxyvinylglycine also increased stem elongation in lkb plants, and combined application of HFCA and aminoethoxy-vinylglycine restored lkb internodes to the wild-type length. It is concluded that the level of IAA in wild-type internodes is necessary for normal elongation, and that the reduced stature of lkb plants is at least partially attributable to a reduction in free IAA level in this mutant.     

Structural comparison suggests that thermolysin and related neutral proteases undergo hinge-bending motion during catalysis.

Crystal structures are known for three members of the bacterial neutral protease family: thermolysin from Bacillus thermoproteolyticus (TLN), the neutral protease from Bacillus cereus (NEU), and the elastase of Pseudomonas aeruginosa (PAE), both in free and ligand-bound forms. Each enzyme consists of an N-terminal and C-terminal domain with the active site formed at the junction of the two domains. Comparison of the different molecules reveals that the structure within each domain is well conserved, but there are substantial hinge-bending displacements (up to 16 degrees) of one domain relative to the other. These domain motions can be correlated with the presence or absence of bound inhibitor, as was previously observed in the specific example of PAE [Thayer, M.M., Flaherty, K.M., & McKay, D.B. (1991) J. Biol. Chem. 266, 2864-2871]. The binding of inhibitor appears to be associated with a reduction of the domain hinge-bending angle by 6-14 degrees and a closure of the "jaws" of the active site cleft by about 2 A. Crystallographic refinement of the structure of thermolysin suggests that electron density seen in the active site of the enzyme in the original structure determination probably corresponds to a bound dipeptide. Thus, the crystal structure appears to correspond to an enzyme-inhibitor or enzyme-product complex, rather than the free enzyme, as has previously been assumed.     

Malignant transformation of murine fibroblasts by a human c-Ha-ras-1 oncogene does not require a functional epidermal growth factor receptor.

Although mutations in ras genes are thought to be important for the development of about 20% of human tumors, almost nothing is known about the way in which these mutations lead to cellular transformation. The known biochemical properties of the 21-kilodalton ras proteins suggest that they may behave as G proteins, regulating the proliferation of cells in response to growth factor stimulation of a receptor. Although the putative receptor(s) has not been identified, several lines of evidence, in particular the fact that rodent cell lines containing ras oncogenes produce transforming growth factor alpha, have suggested that the epidermal growth factor (EGF) receptor is involved in ras transformation. Here we show that murine fibroblasts with no EGF receptors can be transformed to a completely malignant phenotype with a mutated ras gene. It appears, therefore, that the EGF receptor is not required for ras-mediated transformation of these cells.     

Optimization of muscle activity for task-level goals predicts complex changes in limb forces across biomechanical contexts

Optimality principles have been proposed as a general framework for understanding motor control in animals and humans largely based on their ability to predict general features movement in idealized motor tasks. However, generalizing these concepts past proof-of-principle to understand the neuromechanical transformation from task-level control to detailed execution-level muscle activity and forces during behaviorally-relevant motor tasks has proved difficult. In an unrestrained balance task in cats, we demonstrate that achieving task-level constraints center of mass forces and moments while minimizing control effort predicts detailed patterns of muscle activity and ground reaction forces in an anatomically-realistic musculoskeletal model. Whereas optimization is typically used to resolve redundancy at a single level of the motor hierarchy, we simultaneously resolved redundancy across both muscles and limbs and directly compared predictions to experimental measures across multiple perturbation directions that elicit different intra- and interlimb coordination patterns. Further, although some candidate task-level variables and cost functions generated indistinguishable predictions in a single biomechanical context, we identified a common optimization framework that could predict up to 48 experimental conditions per animal (n = 3) across both perturbation directions and different biomechanical contexts created by altering animals' postural configuration. Predictions were further improved by imposing experimentally-derived muscle synergy constraints, suggesting additional task variables or costs that may be relevant to the neural control of balance. These results suggested that reduced-dimension neural control mechanisms such as muscle synergies can achieve similar kinetics to the optimal solution, but with increased control effort (≈2×) compared to individual muscle control. Our results are consistent with the idea that hierarchical, task-level neural control mechanisms previously associated with voluntary tasks may also be used in automatic brainstem-mediated pathways for balance.     

Toward routine,DNA-based detection methods for marine pests

Marine pest incursions can cause significant ongoing damage to aquaculture, biodiversity, fisheries habitat, infrastructure and social amenity. They represent a significant and ongoing economic burden. Marine pests can be introduced by several vectors including aquaculture, aquarium trading, commercial shipping, fishing, floating debris, mining activities and recreational boating. Despite the inherent risks, there is currently relatively little routine surveillance of marine pest species conducted in the majority of countries worldwide. Accurate and rapid identification of marine pest species is central to early detection and management. Traditional techniques (e.g. physical sampling and sorting), have limitations, which has motivated some progress towards the development of molecular diagnostic tools. This review provides a brief account of the techniques traditionally used for detection and describes developments in molecular-based methods for the detection and surveillance of marine pest species. Recent advances provide a platform for the development of practical, specific, sensitive and rapid diagnosis and surveillance tools for marine pests for use in effective prevention and control strategies.     

Crystals of an ATPase fragment of bovine clathrin uncoating ATPase

A 44,000 Mr amino-terminal, clathrin-independent ATPase fragment of the bovine clathrin uncoating ATPase has been crystallized in a form suitable for X-ray diffraction studies. The crystals are orthorhombic, space group P2(1)2(1)2(1), a = 145.3 A, b = 65.0 A, c = 46.9 A, with one protein molecule per asymmetric unit (1 A = 0.1 nm).