Single-molecule transcript counting of stem-cell markers in the mouse intestine

available in PMC 2012 July 1.Determining the molecular identities of adult stem cells requires technologies for sensitive transcript detection in tissues. In mouse intestinal crypts, lineage-tracing studies indicated that different genes uniquely mark spatially distinct stem-cell populations, residing either at crypt bases or at position +4, but a detailed analysis of their spatial co-expression has not been feasible. Here we apply three-colour single-molecule fluorescent in situ hybridization to study a comprehensive panel of intestinal stem-cell markers during homeostasis, ageing and regeneration. We find that the expression of all markers overlaps at crypt-base cells. This co-expression includes Lgr5, Bmi1 and mTert, genes previously suggested to mark distinct stem cells. Strikingly, Dcamkl1 tuft cells, distributed throughout the crypt axis, co-express Lgr5 and other stem-cell markers that are otherwise confined to crypt bases. We also detect significant changes in the expression of some of the markers following irradiation, indicating their potential role in the regeneration process. Our approach can enable the sensitive detection of putative stem cells in other tissues and in tumours, guiding complementary functional studies to evaluate their stem-cell properties.National Institutes of Health (U.S.) (U54CA143874)National Cancer Institute (U.S.) (Physical Sciences Oncology Center at MIT (U54CA143874))National Institutes of Health (U.S.) (NIH Pioneer award (1DP1OD003936))National Cancer Institute (U.S.) (Cancer Center Support (core) grant P30-CA14051)European Molecular Biology Organization (postdoctoral fellowship)Human Frontier Science Program (Strasbourg, France)Machiah FoundationHoward Hughes Medical Institute (Gilliam fellowship

Family model of HIV care and treatment: a retrospective study in Kenya

<p>Abstract</p> <p>Background</p> <p>Nyanza Province, Kenya, had the highest HIV prevalence in the country at 14.9% in 2007, more than twice the national HIV prevalence of 7.1%. Only 16% of HIV-infected adults in the country accurately knew their HIV status. Targeted strategies to reach and test individuals are urgently needed to curb the HIV epidemic. The family unit is one important portal.</p> <p>Methods</p> <p>A family model of care was designed to build on the strengths of Kenyan families. Providers use a family information table (FIT) to guide index patients through the steps of identifying family members at HIV risk, address disclosure, facilitate family testing, and work to enrol HIV-positive members and to prevent new infections. Comprehensive family-centred clinical services are built around these steps. To assess the approach, a retrospective study of patients receiving HIV care between September 2007 and September 2009 at Lumumba Health Centre in Kisumu was conducted. A random sample of FITs was examined to assess family reach.</p> <p>Results</p> <p>Through the family model of care, for each index patient, approximately 2.5 family members at risk were identified and 1.6 family members were tested. The approach was instrumental in reaching children; 61% of family members identified and tested were children. The approach also led to identifying and enrolling a high proportion of HIV- positive partners among those tested: 71% and 89%, respectively.</p> <p>Conclusions</p> <p>The family model of care is a feasible approach to broaden HIV case detection and service reach. The approach can be adapted for the local context and should continue to utilize index patient linkages, FIT adaption, and innovative methods to package services for families in a manner that builds on family support and enhances patient care and prevention efforts. Further efforts are needed to increase family member engagement.</p

Differential Affinity and Catalytic Activity of CheZ in E. coli Chemotaxis

Push–pull networks, in which two antagonistic enzymes control the activity of a messenger protein, are ubiquitous in signal transduction pathways. A classical example is the chemotaxis system of the bacterium Escherichia coli, in which the kinase CheA and the phosphatase CheZ regulate the phosphorylation level of the messenger protein CheY. Recent experiments suggest that both the kinase and the phosphatase are localized at the receptor cluster, and Vaknin and Berg recently demonstrated that the spatial distribution of the phosphatase can markedly affect the dose–response curves. We argue, using mathematical modeling, that the canonical model of the chemotaxis network cannot explain the experimental observations of Vaknin and Berg. We present a new model, in which a small fraction of the phosphatase is localized at the receptor cluster, while the remainder freely diffuses in the cytoplasm; moreover, the phosphatase at the cluster has a higher binding affinity for the messenger protein and a higher catalytic activity than the phosphatase in the cytoplasm. This model is consistent with a large body of experimental data and can explain many of the experimental observations of Vaknin and Berg. More generally, the combination of differential affinity and catalytic activity provides a generic mechanism for amplifying signals that could be exploited in other two-component signaling systems. If this model is correct, then a number of recent modeling studies, which aim to explain the chemotactic gain in terms of the activity of the receptor cluster, should be reconsidered

MLK3 Limits Activated Gαq Signaling to Rho by Binding to p63RhoGEF

Mixed lineage kinase 3 (MLK3) is a MAP3K that activates the JNK-dependent MAPK pathways. Here we show that MLK3 is required for cell migration in a manner independent of its role as a MAP3K or MLK3 kinase activity. Rather, MLK3 functions in a regulated way to limit levels of the activated GTPase, Rho, by binding to the Rho activator, p63RhoGEF/GEFT, which, in turn, prevents its activation by Gαq. These findings demonstrate a scaffolding role for MLK3 in controlling the extent of Rho activation that modulates cell migration. Moreover, they suggest that MLK3 functions as a network hub that links a number of signaling pathways

D4.2 Evaluation of cycle 1 pilots

This report describes the efforts in cycle 1 pilots to evaluate the concept of TENCompetence and its first-release system implementation. Cycle 1 pilots are framed in two different domains: Digital Cinema and ICT Teacher Training. Both pilots are preceded by two preliminary experiences, which are very useful for planning the actual cycle 1 implementation, deployment and evaluation. The first results from cycle 1 pilots are encouraging and indicate that learners using the first-release of the TENCompetence infrastructure feel more in control of their own learning.The work on this publication has been sponsored by the TENCompetence Integrated Project that is funded by the European Commission's 6th Framework Programme, priority IST/Technology Enhanced Learning. Contract 027087 [http://www.tencompetence.org

Lubricating Bacteria Model for Branching growth of Bacterial Colonies

Various bacterial strains (e.g. strains belonging to the genera Bacillus, Paenibacillus, Serratia and Salmonella) exhibit colonial branching patterns during growth on poor semi-solid substrates. These patterns reflect the bacterial cooperative self-organization. Central part of the cooperation is the collective formation of lubricant on top of the agar which enables the bacteria to swim. Hence it provides the colony means to advance towards the food. One method of modeling the colonial development is via coupled reaction-diffusion equations which describe the time evolution of the bacterial density and the concentrations of the relevant chemical fields. This idea has been pursued by a number of groups. Here we present an additional model which specifically includes an evolution equation for the lubricant excreted by the bacteria. We show that when the diffusion of the fluid is governed by nonlinear diffusion coefficient branching patterns evolves. We study the effect of the rates of emission and decomposition of the lubricant fluid on the observed patterns. The results are compared with experimental observations. We also include fields of chemotactic agents and food chemotaxis and conclude that these features are needed in order to explain the observations.Comment: 1 latex file, 16 jpeg files, submitted to Phys. Rev.

Phonons and related properties of extended systems from density-functional perturbation theory

This article reviews the current status of lattice-dynamical calculations in crystals, using density-functional perturbation theory, with emphasis on the plane-wave pseudo-potential method. Several specialized topics are treated, including the implementation for metals, the calculation of the response to macroscopic electric fields and their relevance to long wave-length vibrations in polar materials, the response to strain deformations, and higher-order responses. The success of this methodology is demonstrated with a number of applications existing in the literature.Comment: 52 pages, 14 figures, submitted to Review of Modern Physic

Smc5/6 coordinates formation and resolution of joint molecules with chromosome morphology to ensure meiotic divisions

During meiosis, Structural Maintenance of Chromosome (SMC) complexes underpin two fundamental features of meiosis: homologous recombination and chromosome segregation. While meiotic functions of the cohesin and condensin complexes have been delineated, the role of the third SMC complex, Smc5/6, remains enigmatic. Here we identify specific, essential meiotic functions for the Smc5/6 complex in homologous recombination and the regulation of cohesin. We show that Smc5/6 is enriched at centromeres and cohesin-association sites where it regulates sister-chromatid cohesion and the timely removal of cohesin from chromosomal arms, respectively. Smc5/6 also localizes to recombination hotspots, where it promotes normal formation and resolution of a subset of joint-molecule intermediates. In this regard, Smc5/6 functions independently of the major crossover pathway defined by the MutLγ complex. Furthermore, we show that Smc5/6 is required for stable chromosomal localization of the XPF-family endonuclease, Mus81-Mms4Eme1. Our data suggest that the Smc5/6 complex is required for specific recombination and chromosomal processes throughout meiosis and that in its absence, attempts at cell division with unresolved joint molecules and residual cohesin lead to severe recombination-induced meiotic catastroph