Executive summary of the 12th HHT international scientific conference

Hereditary hemorrhagic telangiectasia is an autosomal dominant trait affecting approximately 1 in 5000 people. A pathogenic DNA sequence variant in the ENG, ACVRL1 or SMAD4 genes, can be found in the majority of patients. The 12th International Scientific HHT Conference was held on June 8–11, 2017 in Dubrovnik, Croatia to present and discuss the latest scientific achievements, and was attended by over 200 scientific and clinical researchers. In total 174 abstracts were accepted of which 58 were selected for oral presentations. This article covers the basic science and clinical talks, and discussions from three theme-based workshops. We focus on significant emergent themes and unanswered questions. Understanding these topics and answering these questions will help to define the future of HHT research and therapeutics, and ultimately bring us closer to a cure

The catalytic subunit of the system L1 amino acid transporter (S<i>lc7a5</i>) facilitates nutrient signalling in mouse skeletal muscle

The System L1-type amino acid transporter mediates transport of large neutral amino acids (LNAA) in many mammalian cell-types. LNAA such as leucine are required for full activation of the mTOR-S6K signalling pathway promoting protein synthesis and cell growth. The SLC7A5 (LAT1) catalytic subunit of high-affinity System L1 functions as a glycoprotein-associated heterodimer with the multifunctional protein SLC3A2 (CD98). We generated a floxed Slc7a5 mouse strain which, when crossed with mice expressing Cre driven by a global promoter, produced Slc7a5 heterozygous knockout (Slc7a5+/-) animals with no overt phenotype, although homozygous global knockout of Slc7a5 was embryonically lethal. Muscle-specific (MCK Cre-mediated) Slc7a5 knockout (MS-Slc7a5-KO) mice were used to study the role of intracellular LNAA delivery by the SLC7A5 transporter for mTOR-S6K pathway activation in skeletal muscle. Activation of muscle mTOR-S6K (Thr389 phosphorylation) in vivo by intraperitoneal leucine injection was blunted in homozygous MS-Slc7a5-KO mice relative to wild-type animals. Dietary intake and growth rate were similar for MS-Slc7a5-KO mice and wild-type littermates fed for 10 weeks (to age 120 days) with diets containing 10%, 20% or 30% of protein. In MS-Slc7a5-KO mice, Leu and Ile concentrations in gastrocnemius muscle were reduced by ∼40% as dietary protein content was reduced from 30 to 10%. These changes were associated with >50% decrease in S6K Thr389 phosphorylation in muscles from MS-Slc7a5-KO mice, indicating reduced mTOR-S6K pathway activation, despite no significant differences in lean tissue mass between groups on the same diet. MS-Slc7a5-KO mice on 30% protein diet exhibited mild insulin resistance (e.g. reduced glucose clearance, larger gonadal adipose depots) relative to control animals. Thus, SLC7A5 modulates LNAA-dependent muscle mTOR-S6K signalling in mice, although it appears non-essential (or is sufficiently compensated by e.g. SLC7A8 (LAT2)) for maintenance of normal muscle mass

Ancient Ethiopian genome reveals extensive Eurasian admixture throughout the African continent.

Characterizing genetic diversity in Africa is a crucial step for most analyses reconstructing the evolutionary history of anatomically modern humans. However, historic migrations from Eurasia into Africa have affected many contemporary populations, confounding inferences. Here, we present a 12.5× coverage ancient genome of an Ethiopian male ("Mota") who lived approximately 4500 years ago. We use this genome to demonstrate that the Eurasian backflow into Africa came from a population closely related to Early Neolithic farmers, who had colonized Europe 4000 years earlier. The extent of this backflow was much greater than previously reported, reaching all the way to Central, West, and Southern Africa, affecting even populations such as Yoruba and Mbuti, previously thought to be relatively unadmixed, who harbor 6 to 7% Eurasian ancestry.A.M. was supported by European Research Council (ERC) Consolidator Grant 647787 “LocalAdaptation”; R.P by ERC Starting Grant 263441, “ADNABIOARC”; M.H. by ERC Consolidator Grant 310763 “GeneFlow”; J.B. by the 2014 Research Fund (1.140113.01, 1.140064.01) of UNIST (Ulsan National Institute of Science and Technology) and Geromics internal research funding; J.T.S. by ERC Consolidator Grant 617627 “ADaPt”; K.W.A. by NSF award 1027607; D.G.B. by ERC Investigator Grant 295729-CodeX; V.S. by a scholarship from the Gates Cambridge Trust; and M.G.L. by a Biotechnology and Biological Sciences Research Council (BBSRC) DTP studentship. Permission for the archaeology was given by the Ethiopian Authority for Research and Conservation of Cultural Heritage and offices of the Ministry of Culture and Tourism for the Southern Nations, Nationalities, and Peoples Region. Raw reads from Mota are available for download through the National Center for Biotechnology Information, BioProject ID PRJNA295861, and the corresponding BAM and VCF files are available at africangenome.orgThis is the author accepted manuscript. The final version is available from AAAS via http://dx.doi.org/10.1126/science.aad287

The Na(+)–H(+ )exchanger-1 induces cytoskeletal changes involving reciprocal RhoA and Rac1 signaling, resulting in motility and invasion in MDA-MB-435 cells

INTRODUCTION: An increasing body of evidence shows that the tumour microenvironment is essential in driving neoplastic progression. The low serum component of this microenvironment stimulates motility/invasion in human breast cancer cells via activation of the Na(+)–H(+ )exchanger (NHE) isoform 1, but the signal transduction systems that underlie this process are still poorly understood. We undertook the present study to elucidate the role and pattern of regulation by the Rho GTPases of this serum deprivation-dependent activation of both NHE1 and subsequent invasive characteristics, such as pseudopodia and invadiopodia protrusion, directed cell motility and penetration of normal tissues. METHODS: The present study was performed in a well characterized human mammary epithelial cell line representing late stage metastatic progression, MDA-MB-435. The activity of RhoA and Rac1 was modified using their dominant negative and constitutively active mutants and the activity of NHE1, cell motility/invasion, F-actin content and cell shape were measured. RESULTS: We show for the first time that serum deprivation induces NHE1-dependent morphological and cytoskeletal changes in metastatic cells via a reciprocal interaction of RhoA and Rac1, resulting in increased chemotaxis and invasion. Deprivation changed cell shape by reducing the amount of F-actin and inducing the formation of leading edge pseudopodia. Serum deprivation inhibited RhoA activity and stimulated Rac1 activity. Rac1 and RhoA were antagonistic regulators of both basal and stimulated tumour cell NHE1 activity. The regulation of NHE1 activity by RhoA and Rac1 in both conditions was mediated by an alteration in intracellular proton affinity of the exchanger. Interestingly, the role of each of these G-proteins was reversed during serum deprivation; basal NHE1 activity was regulated positively by RhoA and negatively by Rac1, whereas RhoA negatively and Rac1 positively directed the stimulation of NHE1 during serum deprivation. Importantly, the same pattern of RhoA and Rac1 regulation found for NHE1 activity was observed in both basal and serum deprivation dependent increases in motility, invasion and actin cytoskeletal organization. CONCLUSION: Our findings suggest that the reported antagonistic roles of RhoA and Rac1 in cell motility/invasion and cytoskeletal organization may be due, in part, to their concerted action on NHE1 activity as a convergence point

P50, the Small Subunit of DNA Polymerase Delta, Is Required for Mediation of the Interaction of Polymerase Delta Subassemblies with PCNA

Mammalian DNA polymerase δ (pol δ), a four-subunit enzyme, plays a crucial and versatile role in DNA replication and various DNA repair processes. Its function as a chromosomal DNA polymerase is dependent on the association with proliferating cell nuclear antigen (PCNA) which functions as a molecular sliding clamp. All four of the pol δ subunits (p125, p50, p68, and p12) have been reported to bind to PCNA. However, the identity of the subunit of pol δ that directly interacts with PCNA and is therefore primarily responsible for the processivity of the enzyme still remains controversial. Previous model for the network of protein-protein interactions of the pol δ-PCNA complex showed that pol δ might be able to interact with a single molecule of PCNA homotrimer through its three subunits, p125, p68, and p12 in which the p50 was not included in. Here, we have confirmed that the small subunit p50 of human pol δ truthfully interacts with PCNA by the use of far-Western analysis, quantitative ELISA assay, and subcellular co-localization. P50 is required for mediation of the interaction between pol δ subassemblies and PCNA homotrimer. Thus, pol δ interacts with PCNA via its four subunits

Stable Isotope Tracking of Endangered Sea Turtles: Validation with Satellite Telemetry and δ15N Analysis of Amino Acids

Effective conservation strategies for highly migratory species must incorporate information about long-distance movements and locations of high-use foraging areas. However, the inherent challenges of directly monitoring these factors call for creative research approaches and innovative application of existing tools. Highly migratory marine species, such as marine turtles, regularly travel hundreds or thousands of kilometers between breeding and feeding areas, but identification of migratory routes and habitat use patterns remains elusive. Here we use satellite telemetry in combination with compound-specific isotope analysis of amino acids to confirm that insights from bulk tissue stable isotope analysis can reveal divergent migratory strategies and within-population segregation of foraging groups of critically endangered leatherback sea turtles (Dermochelys coriacea) across the Pacific Ocean. Among the 78 turtles studied, we found a distinct dichotomy in δ15N values of bulk skin, with distinct “low δ15N” and “high δ15N” groups. δ15N analysis of amino acids confirmed that this disparity resulted from isotopic differences at the base of the food chain and not from differences in trophic position between the two groups. Satellite tracking of 13 individuals indicated that their bulk skin δ15N value was linked to the particular foraging region of each turtle. These findings confirm that prevailing marine isoscapes of foraging areas can be reflected in the isotopic compositions of marine turtle body tissues sampled at nesting beaches. We use a Bayesian mixture model to show that between 82 and 100% of the 78 skin-sampled turtles could be assigned with confidence to either the eastern Pacific or western Pacific, with 33 to 66% of all turtles foraging in the eastern Pacific. Our forensic approach validates the use of stable isotopes to depict leatherback turtle movements over broad spatial ranges and is timely for establishing wise conservation efforts in light of this species’ imminent risk of extinction in the Pacific

Piezo1 integration of vascular architecture with physiological force

The mechanisms by which physical forces regulate endothelial cells to determine the complexities of vascular structure and function are enigmatic¹⁻⁵. Studies of sensory neurons have suggested Piezo proteins as subunits of Ca²⁺-permeable non-selective cationic channels for detection of noxious mechanical impact⁶⁻⁸. Here we show Piezo1 (Fam38a) channels as sensors of frictional force (shear stress) and determinants of vascular structure in both development and adult physiology. Global or endothelial-specific disruption of mouse Piezo1 profoundly disturbed the developing vasculature and was embryonic lethal within days of the heart beating. Haploinsufficiency was not lethal but endothelial abnormality was detected in mature vessels. The importance of Piezo1 channels as sensors of blood flow was shown by Piezo1 dependence of shear-stress-evoked ionic current and calcium influx in endothelial cells and the ability of exogenous Piezo1 to confer sensitivity to shear stress on otherwise resistant cells. Downstream of this calcium influx there was protease activation and spatial reorganization of endothelial cells to the polarity of the applied force. The data suggest that Piezo1 channels function as pivotal integrators in vascular biology

Genetic Diversity among Enterococcus faecalis

Enterococcus faecalis, a ubiquitous member of mammalian gastrointestinal flora, is a leading cause of nosocomial infections and a growing public health concern. The enterococci responsible for these infections are often resistant to multiple antibiotics and have become notorious for their ability to acquire and disseminate antibiotic resistances. In the current study, we examined genetic relationships among 106 strains of E. faecalis isolated over the past 100 years, including strains identified for their diversity and used historically for serotyping, strains that have been adapted for laboratory use, and isolates from previously described E. faecalis infection outbreaks. This collection also includes isolates first characterized as having novel plasmids, virulence traits, antibiotic resistances, and pathogenicity island (PAI) components. We evaluated variation in factors contributing to pathogenicity, including toxin production, antibiotic resistance, polymorphism in the capsule (cps) operon, pathogenicity island (PAI) gene content, and other accessory factors. This information was correlated with multi-locus sequence typing (MLST) data, which was used to define genetic lineages. Our findings show that virulence and antibiotic resistance traits can be found within many diverse lineages of E. faecalis. However, lineages have emerged that have caused infection outbreaks globally, in which several new antibiotic resistances have entered the species, and in which virulence traits have converged. Comparing genomic hybridization profiles, using a microarray, of strains identified by MLST as spanning the diversity of the species, allowed us to identify the core E. faecalis genome as consisting of an estimated 2057 unique genes

Intestinal strongyloidiasis and hyperinfection syndrome

In spite of recent advances with experiments on animal models, strongyloidiasis, an infection caused by the nematode parasite Strongyloides stercoralis, has still been an elusive disease. Though endemic in some developing countries, strongyloidiasis still poses a threat to the developed world. Due to the peculiar but characteristic features of autoinfection, hyperinfection syndrome involving only pulmonary and gastrointestinal systems, and disseminated infection with involvement of other organs, strongyloidiasis needs special attention by the physician, especially one serving patients in areas endemic for strongyloidiasis. Strongyloidiasis can occur without any symptoms, or as a potentially fatal hyperinfection or disseminated infection. Th(2 )cell-mediated immunity, humoral immunity and mucosal immunity have been shown to have protective effects against this parasitic infection especially in animal models. Any factors that suppress these mechanisms (such as intercurrent immune suppression or glucocorticoid therapy) could potentially trigger hyperinfection or disseminated infection which could be fatal. Even with the recent advances in laboratory tests, strongyloidiasis is still difficult to diagnose. But once diagnosed, the disease can be treated effectively with antihelminthic drugs like Ivermectin. This review article summarizes a case of strongyloidiasis and various aspects of strongyloidiasis, with emphasis on epidemiology, life cycle of Strongyloides stercoralis, clinical manifestations of the disease, corticosteroids and strongyloidiasis, diagnostic aspects of the disease, various host defense pathways against strongyloidiasis, and available treatment options

Immunization with Single-Cycle SIV Significantly Reduces Viral Loads After an Intravenous Challenge with SIVmac239

Strains of simian immunodeficiency virus (SIV) that are limited to a single cycle of infection were evaluated for the ability to elicit protective immunity against wild-type SIVmac239 infection of rhesus macaques by two different vaccine regimens. Six animals were inoculated at 8-week intervals with 6 identical doses consisting of a mixture of three different envelope variants of single-cycle SIV (scSIV). Six additional animals were primed with a mixture of cytoplasmic domain-truncated envelope variants of scSIV and boosted with two doses of vesicular stomatitis virus glycoprotein (VSV G) trans-complemented scSIV. While both regimens elicited detectable virus-specific T cell responses, SIV-specific T cell frequencies were more than 10-fold higher after boosting with VSV G trans-complemented scSIV (VSV G scSIV). Broad T cell recognition of multiple viral antigens and Gag-specific CD4+ T cell responses were also observed after boosting with VSV G scSIV. With the exception of a single animal in the repeated immunization group, all of the animals became infected following an intravenous challenge with SIVmac239. However, significantly lower viral loads and higher memory CD4+ T cell counts were observed in both immunized groups relative to an unvaccinated control group. Indeed, both scSIV immunization regimens resulted in containment of SIVmac239 replication after challenge that was as good as, if not better than, what has been achieved by other non-persisting vaccine vectors that have been evaluated in this challenge model. Nevertheless, the extent of protection afforded by scSIV was not as good as typically conferred by persistent infection with live, attenuated SIV. These observations have potentially important implications to the design of an effective AIDS vaccine, since they suggest that ongoing stimulation of virus-specific immune responses may be essential to achieving the degree of protection afforded by live, attenuated SIV