MicroRNA 10a marks regulatory T cells

MicroRNAs (miRNAs) are crucial for regulatory T cell (Treg) stability and function. We report that microRNA-10a (miR-10a) is expressed in Tregs but not in other T cells including individual thymocyte subsets. Expression profiling in inbred mouse strains demonstrated that non-obese diabetic (NOD) mice with a genetic susceptibility for autoimmune diabetes have lower Treg-specific miR-10a expression than C57BL/6J autoimmune resistant mice. Inhibition of miR-10a expression in vitro leads to reduced FoxP3 expression levels and miR-10a expression is lower in unstable "exFoxP3" T cells. Unstable in vitro TGF-?-induced, iTregs do not express miR-10a unless cultured in the presence of retinoic acid (RA) which has been associated with increased stability of iTreg, suggesting that miR-10a might play a role in stabilizing Treg. However, genetic ablation of miR-10a neither affected the number and phenotype of natural Treg nor the capacity of conventional T cells to induce FoxP3 in response to TGFβ, RA, or a combination of the two. Thus, miR-10a is selectively expressed in Treg but inhibition by antagomiRs or genetic ablation resulted in discordant effects on FoxP3.     

Gamma Interferon (IFN-γ) Receptor Restricts Systemic Dengue Virus Replication and Prevents Paralysis in IFN-α/β Receptor-Deficient Mice

We previously reported that mice lacking alpha/beta and gamma interferon receptors (IFN-α/βR and -γR) uniformly exhibit paralysis following infection with the dengue virus (DENV) clinical isolate PL046, while only a subset of mice lacking the IFN-γR alone and virtually no mice lacking the IFN-α/βR alone develop paralysis. Here, using a mouse-passaged variant of PL046, strain S221, we show that in the absence of the IFN-α/βR, signaling through the IFN-γR confers approximately 140-fold greater resistance against systemic vascular leakage-associated dengue disease and virtually complete protection from dengue-induced paralysis. Viral replication in the spleen was assessed by immunohistochemistry and flow cytometry, which revealed a reduction in the number of infected cells due to IFN-γR signaling by 2 days after infection, coincident with elevated levels of IFN-γ in the spleen and serum. By 4 days after infection, IFN-γR signaling was found to restrict DENV replication systemically. Clearance of DENV, on the other hand, occurred in the absence of IFN-γR, except in the central nervous system (CNS) (brain and spinal cord), where clearance relied on IFN-γ from CD8⁺ T cells. These results demonstrate the roles of IFN-γR signaling in protection from initial systemic and subsequent CNS disease following DENV infection and demonstrate the importance of CD8⁺ T cells in preventing DENV-induced CNS disease.     

Caffeine relaxes smooth muscle through actin depolymerization

Caffeine is sometimes used in cell physiological studies to release internally stored Ca(2+). We obtained evidence that caffeine may also act through a different mechanism that has not been previously described and sought to examine this in greater detail. We ruled out a role for phosphodiesterase (PDE) inhibition, since the effect was 1) not reversed by inhibiting PKA or adenylate cyclase; 2) not exacerbated by inhibiting PDE4; and 3) not mimicked by submillimolar caffeine nor theophylline, both of which are sufficient to inhibit PDE. Although caffeine is an agonist of bitter taste receptors, which in turn mediate bronchodilation, its relaxant effect was not mimicked by quinine. After permeabilizing the membrane using β-escin and depleting the internal Ca(2+) store using A23187, we found that 10 mM caffeine reversed tone evoked by direct application of Ca(2+), suggesting it functionally antagonizes the contractile apparatus. Using a variety of molecular techniques, we found that caffeine did not affect phosphorylation of myosin light chain (MLC) by MLC kinase, actin-filament motility catalyzed by MLC kinase, phosphorylation of CPI-17 by either protein kinase C or RhoA kinase, nor the activity of MLC-phosphatase. However, we did obtain evidence that caffeine decreased actin filament binding to phosphorylated myosin heads and increased the ratio of globular to filamentous actin in precontracted tissues. We conclude that, in addition to its other non-RyR targets, caffeine also interferes with actin function (decreased binding by myosin, possibly with depolymerization), an effect that should be borne in mind in studies using caffeine to probe excitation-contraction coupling in smooth muscle.     

A novel and versatile computational tool to model translation

MOTIVATION: Much is now known about the mechanistic details of gene translation. There are also rapid advances in high-throughput technologies to determine quantitative aspects of the system. As a consequence-realistic and system-wide simulation models of translation are now feasible. Such models are also needed as devices to integrate a large volume of highly fragmented data known about translation. Software: In this application note, we present a novel, highly efficient software tool to model translation. The tool represents the main aspects of translation. Features include a representation of exhaustible tRNA pools, ribosome-ribosome interactions and differential initiation rates for different mRNA species. The tool is written in Java, and is hence portable and can be parameterized for any organism. AVAILABILITY: The model can be obtained from the authors or directly downloaded from the authors' home-page (http://goo.gl/JUWvI).     

Genetic structure of peripheral, island-like populations: a case study of Saponaria bellidifolia Sm. (Caryophyllaceae) from the Southeastern Carpathians

Geographically peripheral populations often experience a reduction of genetic diversity and divergence from the core populations. Habitat geometry and quality can induce a local genetic diversity pattern, which overlies the regional variability issued from the range-wide phylogeography. We evaluated the genetic variation and genetic divergence of Saponaria bellidifolia Sm. on limestone outcrops within peripheral island-like populations from the Southeastern Carpathians, using RAPD markers. We also determined the degree of isolation related to other European populations, using AFLP. The Romanian populations had a decreased overall genetic diversity shared among populations, with lower level in small populations. Potential habitat size had a positive effect on genetic diversity estimates. Fisher’s exact tests of genetic differentiation revealed significant divergences only between the geographically most distant populations. Romanian populations were genetically pauperised as compared to Bulgarian and Italian populations and our results suggest that they might have originated from a recent range expansion from southern glacial refugia.     

LINC01101 and LINC00277 expression levels as novel factors in HPV‐induced cervical neoplasia

Recently long non‐coding RNAs were identified as new factors involved in gene expression regulation. To gain insight into expression pattern of these factors related to E7 HPV18 oncogene, this study uses HeLa cell culture transfected with E7‐siRNA. Gene expression profile was investigated using microarray analysis. After analysing the microarray results, we identified 15,387 RNA species differentially expressed in E7‐siRNA‐transfected cells compared with controls (fold change >2). The expression profiles of lncRNA species highlighted 731 lncRNAs and 203 lincRNAs. We selected two lincRNAs (LINC01101 and LINC00277) and we evaluated the expression profile in HPV‐induced neoplasia. Both lincRNAs investigated display a significantly reduced pattern of expression in cervical lesions and cancer, associated with clinical parameters. A connection between HPV presence and lincRNAs was noted. hrHPV‐positive samples exhibit significantly reduced LINC01101 and LINC00277 expression level (P < 0.05). These results provide new insights into involvement of lncRNA in HPV‐induced cervical cancer, enriching our understanding of their potential role in this pathology.     

Association studies for untyped markers with TUNA

The software package TUNA (Testing UNtyped Alleles) implements a fast and efficient algorithm for testing association of genotyped and ungenotyped variants in genome-wide case-control studies. TUNA uses Linkage Disequilibrium (LD) information from existing comprehensive variation datasets such as HapMap to construct databases of frequency predictors using linear combination of haplotype frequencies of genotyped SNPs. The predictors are used to estimate untyped allele frequencies, and to perform association tests. The methods incorporated in TUNA achieve great accuracy in estimation, and the software is computationally efficient and does not demand a lot of system memory and CPU resources. AVAILABILITY: The software package is available for download from the website: http://www.stat.uchicago.edu/~wen/tuna/.     

MDR1 Ala893 polymorphism is associated with inflammatory bowel disease

Crohn disease (CD) and ulcerative colitis (UC) are overlapping chronic inflammatory bowel diseases (IBDs). Suggestive evidence for linkage at chromosome 7q has been reported for both CD and UC. Contained within this region is the gene for MDR1 (multidrug resistance), a membrane transport protein for which human polymorphisms have been reported in Ala893Ser/Thr and C3435T that alter pharmacokinetic profiles for a variety of drugs. Because mdr1 knockout mice spontaneously develop colitis, exonic regions were resequenced and tested for IBD association in a large, multicenter North American cohort. Two missense mutations, Asn21Asp and Ala893Ser/Thr, as well as the expression-associated polymorphism C3435T, described elsewhere, were genotyped in the entire cohort. Significant association of Ala893 with IBD was observed by both case-control analysis (P=.002) and the pedigree disequilibrium test (PDT [P=.00020-.00030]) but not for the Asn21Asp or C3435T polymorphisms. Significant association by PDT was observed within the subset with CD (P=.0014-.00090), with similar, nonsignificant trends in a smaller subset with UC. The Ala893Ser/Thr variant is triallelic, and the associated, common allele is Ala893, with undertransmission of the 893Ser (common) and the 893Thr (rare) variants. The Ala893 variant has decreased activity compared with the 893Ser variant; therefore, the association with human IBD is consistent with the murine model of mdr1 deficiency. Taken together, these data support the association of the common Ala893 polymorphism with IBD specifically and, more broadly, provides additional support for its contribution to interindividual pharmacogenetic variation.