Defining functional interactions during biogenesis of epithelial junctions

In spite of extensive recent progress, a comprehensive understanding of how actin cytoskeleton remodelling supports stable junctions remains to be established. Here we design a platform that integrates actin functions with optimized phenotypic clustering and identify new cytoskeletal proteins, their functional hierarchy and pathways that modulate E-cadherin adhesion. Depletion of EEF1A, an actin bundling protein, increases E-cadherin levels at junctions without a corresponding reinforcement of cell-cell contacts. This unexpected result reflects a more dynamic and mobile junctional actin in EEF1A-depleted cells. A partner for EEF1A in cadherin contact maintenance is the formin DIAPH2, which interacts with EEF1A. In contrast, depletion of either the endocytic regulator TRIP10 or the Rho GTPase activator VAV2 reduces E-cadherin levels at junctions. TRIP10 binds to and requires VAV2 function for its junctional localization. Overall, we present new conceptual insights on junction stabilization, which integrate known and novel pathways with impact for epithelial morphogenesis, homeostasis and diseases

E-cadherin and cell adhesion: a role in architecture and function in the pancreatic islet

Background/Aims: The efficient secretion of insulin from beta-cells requires extensive intra-islet communication. The cell surface adhesion protein epithelial (E)-cadherin (ECAD) establishes and maintains epithelial tissues such as the islets of Langerhans. In this study, the role of ECAD in regulating insulin secretion from pseudoislets was investigated. Methods: The effect of an immuno-neutralising ECAD on gross morphology, cytosolic calcium signalling, direct cell-to-cell communication and insulin secretion was assessed by fura-2 microfluorimetry, Lucifer Yellow dye injection and insulin ELISA in an insulin-secreting model system. Results: Antibody blockade of ECAD reduces glucose-evoked changes in [Ca2+](i) and insulin secretion. Neutralisation of ECAD causes a breakdown in the glucose-stimulated synchronicity of calcium oscillations between discrete regions within the pseudoislet, and the transfer of dye from an individual cell within a cell cluster is attenuated in the absence of ECAD ligation, demonstrating that gap junction communication is disrupted. The functional consequence of neutralising ECAD is a significant reduction in insulin secretion. Conclusion: Cell adhesion via ECAD has distinct roles in the regulation of intercellular communication between beta-cells within islets, with potential repercussions for insulin secretion. Copyright (C) 2007 S. Karger AG, Basel

miR-17-5p regulates endocytic trafficking through targeting TBC1D2/ Armus

miRNA cluster miR-17-92 is known as oncomir-1 due to its potent oncogenic function. miR-17-92 is a polycistronic cluster that encodes 6 miRNAs, and can both facilitate and inhibit cell proliferation. Known targets of miRNAs encoded by this cluster are largely regulators of cell cycle progression and apoptosis. Here, we show that miRNAs encoded by this cluster and sharing the seed sequence of miR-17 exert their influence on one of the most essential cellular processes – endocytic trafficking. By mRNA expression analysis we identified that regulation of endocytic trafficking by miR-17 can potentially be achieved by targeting of a number of trafficking regulators. We have thoroughly validated TBC1D2/Armus, a GAP of Rab7 GTPase, as a novel target of miR-17. Our study reveals regulation of endocytic trafficking as a novel function of miR-17, which might act cooperatively with other functions of miR-17 and related miRNAs in health and disease

Vps34 regulates Rab7 and late endocytic trafficking through recruitment of the GTPase-activating protein Armus

The class III phosphoinositide 3-kinase (PI3K) Vps34 (also known as PIK3C3 in mammals) produces phosphatidylinositol 3-phosphate [PI(3)P] on both early and late endosome membranes to control membrane dynamics. We used Vps34-deficient cells to delineate whether Vps34 has additional roles in endocytic trafficking. In Vps34−/− mouse embryonic fibroblasts (MEFs), transferrin recycling and EEA1 membrane localization were unaffected despite elevated Rab5-GTP levels. Strikingly, a large increase in Rab7-GTP levels, an accumulation of enlarged late endosomes, and decreased EGFR degradation were observed in Vps34-deficient cells. The hyperactivation of Rab7 in Vps34-deficient cells stemmed from the failure to recruit the Rab7 GTPase-activating protein (GAP) Armus (also known as TBC1D2), which binds to PI(3)P, to late endosomes. Protein–lipid overlay and liposome-binding assays reveal that the putative pleckstrin homology (PH) domain in Armus can directly bind to PI(3)P. Elevated Rab7-GTP led to the failure of intraluminal vesicle (ILV) formation and lysosomal maturation. Rab7 silencing and Armus overexpression alleviated the vacuolization seen in Vps34-deficient cells. Taken together, these results demonstrate that Vps34 has a previously unknown role in regulating Rab7 activity and late endosomal trafficking

Rare Exonic Minisatellite Alleles in MUC2 Influence Susceptibility to Gastric Carcinoma

BACKGROUND: Mucins are the major components of mucus and their genes share a common, centrally-located region of sequence that encodes tandem repeats. Mucins are well known genes with respect to their specific expression levels; however, their genomic levels are unclear because of complex genomic properties. In this study, we identified eight novel minisatellites from the entire MUC2 region and investigated how allelic variation in these minisatellites may affect susceptibility to gastrointestinal cancer. METHODOLOGY/PRINCIPLE FINDINGS: We analyzed genomic DNA from the blood of normal healthy individuals and multi-generational family groups. Six of the eight minisatellites exhibited polymorphism and were transmitted meiotically in seven families, following Mendelian inheritance. Furthermore, a case-control study was performed that compared genomic DNA from 457 cancer-free controls with DNA from individuals with gastric (455), colon (192) and rectal (271) cancers. A statistically significant association was identified between rare exonic MUC2-MS6 alleles and the occurrence of gastric cancer: odds ratio (OR), 2.56; 95% confidence interval (CI), 1.31-5.04; and p = 0.0047. We focused on an association between rare alleles and gastric cancer. Rare alleles were divided into short (40, 43 and 44) and long (47, 50 and 54), according to their TR (tandem repeats) lengths. Interestingly, short rare alleles were associated with gastric cancer (OR = 5.6, 95% CI: 1.93-16.42; p = 0.00036). Moreover, hypervariable MUC2 minisatellites were analyzed in matched blood and cancer tissue from 28 patients with gastric cancer and in 4 cases of MUC2-MS2, minisatellites were found to have undergone rearrangement. CONCLUSIONS/SIGNIFICANCE: Our observations suggest that the short rare MUC2-MS6 alleles could function as identifiers for risk of gastric cancer. Additionally, we suggest that minisatellite instability might be associated with MUC2 function in cancer cells

Control of Visceral Leishmaniasis in Latin America—A Systematic Review

Visceral leishmaniasis is a vector-borne disease characterized by fever, spleen and liver enlargement, and low blood cell counts. In the Americas VL is zoonotic, with domestic dogs as main animal reservoirs, and is caused by the intracellular parasite Leishmania infantum (syn. Leishmania chagasi). Humans acquire the infection through the bite of an infected sand fly. The disease is potentially lethal if untreated. VL is reported from Mexico to Argentina, with recent trends showing a rapid spread in Brazil. Control measures directed against the canine reservoir and insect vectors have been unsuccessful, and early detection and treatment of human cases remains as the most important strategy to reduce case fatality. Well-designed studies evaluating diagnosis, treatment, and prevention/control interventions are scarce. The available scientific evidence reasonably supports the use of rapid diagnostic tests for the diagnosis of human disease. Properly designed randomized controlled trials following good clinical practices are needed to inform drug policy. Routine control strategies against the canine reservoirs and insect vectors are based on weak and conflicting evidence, and vector control strategies and vaccine development should constitute research priorities

One Health: The global challenge of epidemic and endemic leishmaniasis

'One Health' proposes the unification of medical and veterinary sciences with the establishment of collaborative ventures in clinical care, surveillance and control of cross-species disease, education, and research into disease pathogenesis, diagnosis, therapy and vaccination. The concept encompasses the human population, domestic animals and wildlife, and the impact that environmental changes ('environmental health') such as global warming will have on these populations. Visceral leishmaniasis is a perfect example of a small companion animal disease for which prevention and control might abolish or decrease the suffering of canine and human patients, and which aligns well with the One Health approach. In this review we discuss how surveillance for leishmaniases is undertaken globally through the control of anthroponootic visceral leishmaniasis (AVL) and zoonotic visceral leishmaniasis (ZVL). The ZVL epidemic has been managed to date by the culling of infected dogs, treatment of human cases and control of the sandfly vector by insecticidal treatment of human homes and the canine reservoir. Recently, preventive vaccination of dogs in Brazil has led to reduction in the incidence of the canine and human disease. Vaccination permits greater dog owner compliance with control measures than a culling programme. Another advance in disease control in Africa is provided by a surveillance programme that combines remote satellite sensing, ecological modelling, vector surveillance and geo-spatial mapping of the distribution of vectors and of the animal-to-animal or animal-to-human pathogen transmission. This coordinated programme generates advisory notices and alerts on emerging infectious disease outbreaks that may impede or avoid the spreading of visceral leishmaniasis to new areas of the planet as a consequence of global warming

Cooperation of distinct Rac-dependent pathways to stabilize E-cadherin adhesion

AbstractThe precise mechanisms via which Rac1 is activated by cadherin junctions are not fully known. In keratinocytes Rac1 activation by cadherin junctions requires EGFR signalling, but how EGFR does so is unclear. To address which activator could mediate E-cadherin signalling to Rac1, we investigated EGFR and two Rac1 GEFs, SOS1 and DOCK180. EGFR RNAi prevented junction-induced Rac1 activation and led to fragmented localization of E-cadherin at cadherin contacts. In contrast, depletion of another EGFR family member, ErbB3, did not interfere with either process. DOCK180 RNAi, but not SOS1, prevented E-cadherin-induced Rac1 activation. However, in a strong divergence from EGFR RNAi phenotype, DOCK180 depletion did not perturb actin recruitment or cadherin localisation at junctions. Rather, reduced DOCK180 levels impaired the resistance to mechanical stress of pre-formed cell aggregates. Thus, within the same cell type, EGFR and DOCK180 regulate Rac1 activation by newly-formed contacts, but control separate cellular events that cooperate to stabilise junctions

Vascular permeability: flow-mediated, non-canonical notch signalling promotes barrier integrity

The vascular permeability barrier must be maintained in response to changes to vessel calibre, shear stress and blood pressure. A new study reveals a remarkable mechanism for flow-mediated regulation of permeability: Notch1 activation leads to the assembly of GTPase signalling complexes at VE-cadherin contacts and a strengthening of the endothelial barrier