A flexible computational framework for detecting, characterizing, and interpreting statistical patterns of epistasis in genetic studies of human disease susceptibility

Detecting, characterizing, and interpreting gene-gene interactions or epistasis in studies of human disease susceptibility is both a mathematical and a computational challenge. To address this problem, we have previously developed a multifactor dimensionality reduction (MDR) method for collapsing high-dimensional genetic data into a single dimension (i.e. constructive induction) thus permitting interactions to be detected in relatively small sample sizes. In this paper, we describe a comprehensive and flexible framework for detecting and interpreting gene-gene interactions that utilizes advances in information theory for selecting interesting single-nucleotide polymorphisms (SNPs), MDR for constructive induction, machine learning methods for classification, and finally graphical models for interpretation. We illustrate the usefulness of this strategy using artificial datasets simulated from several different two-locus and three-locus epistasis models. We show that the accuracy, sensitivity, specificity, and precision of a na?ve Bayes classifier are significantly improved when SNPs are selected based on their information gain (i.e. class entropy removed) and reduced to a single attribute using MDR. We then apply this strategy to detecting, characterizing, and interpreting epistatic models in a genetic study (n = 500) of atrial fibrillation and show that both classification and model interpretation are significantly improved.     

Effects of 900 MHz GSM wireless communication signals on DMBA-induced mammary tumors in rats

The purpose of the study was to investigate whether exposure to 900 MHz GSM wireless communication signals enhances mammary tumor development and growth induced by low-dose DMBA. Five hundred female Sprague-Dawley rats were treated with a single dose of 35 mg/kg DMBA and then divided into five groups in a blinded fashion: one cage control group and four exposure groups, including three microwave exposure groups and one sham exposure with specific absorption rates (SARs) of 4.0, 1.33, 0.44 and 0 W/kg, respectively. Exposure started on the day after DMBA administration and lasted 4 h/day, 5 days/week for 26 weeks. Rats were weighed and palpated weekly for the presence of tumors and were killed humanely at the end of the 26-week exposure period. All mammary glands were examined histologically. There were no statistically significant differences in body weight between sham- and GSM microwave-exposed groups. No significant differences in overall mammary tumor incidence, latency to tumor onset, tumor multiplicity, or tumor size were observed between microwave- and sham-exposed groups. There was a tendency for reduction of mammary adenocarcinoma incidence in the lowest microwave exposure group (0.44 W/ kg) compared with the sham-exposed group (P = 0.058). Additionally, a higher incidence of adenocarcinoma was noticed in the 4.0 W/kg group from the 15th to 26th weeks, especially in the 19th week (P = 0.358 compared to sham). However, neither tendency was statistically significant; thus this study does not provide evidence that GSM microwave exposure promotes mammary tumor development in rats. In the present study there were significant differences between the cage controls and the experimental groups (sham and exposure). Body weight and mammary tumor (malignant plus benign) incidence in the cage control group were significantly higher than in the sham- and GSM microwave-exposed groups. The latency to the mammary tumor onset was significantly shorter in the cage control group than in the other groups.     

Taxonomic re-assessment of <Emphasis Type="Italic">Phaseolus dasycarpus</Emphasis> (Leguminosae): Systematic position,chromosome studies and re-description

Based on new information on floral structures, seedling, fruit, seed, root, and leaflet characters, Phaseolus dasycarpus is re-described and illustrated. Its chromosome number was determined as 2n  = 22, with metacentric and submetacentric chromosomes, and karyotype formula of 9m + 2sm. Previous phylogenetic analyses of ITS and trnK sequence data, and those obtained in the present study, show P. dasycarpus to be aligned within section Pedicellati.

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Resumen  Con nueva información de estructuras florales, plántulas, frutos, semillas, raíz y folíolos, se redescribe e ilustra a Phaseolus dasycarpus. Se determina el número cromosómico 2n  = 22, con cromosomas metacéntricos y submetacéntricos y fórmula cariotípica = 9m + 2sm. Análisis filogenéticos previos con secuencias de ITS y trnK y los resultados obtenidos en el presente estudio, establecen a P. dasycarpus en la sección Pedicellati.

     

Andrographolide could inhibit human colorectal carcinoma Lovo cells migration and invasion via down-regulation of MMP-7 expression

Andrographolide (Andro), a diterpenoid lactone isolated from a traditional herbal medicine Andrographis paniculata, is known to possess multiple pharmacological activities. In our previous study, Andro had been shown to have potent anti-cancer activity against human colorectal carcinoma Lovo cells by inhibiting cell-cycle progression. To further investigate the mechanism for the anti-cancer properties of Andro, it was used to examine the effect on migration and invasion of Lovo cells. The results of wound-healing assay and in vitro transwell assay revealed that Andro inhibited dose-dependently the migration and invasion of Lovo cells under non-cytotoxic concentrations. Using zymographic assay and RT-PCR, the results revealed that Andro diminished the activity and the mRNA and protein levels of MMP-7, but not MMP-2 or MMP-9. The down-regulation of MMP-7 appeared to be via the inactivation of activator protein-1 (AP-1) since the treatment with Andro suppressed the nuclear protein level of AP-1, which was accompanied by a decrease in DNA-binding level of the factor. Taken together, these results indicated that Andro reduces the MMP-7-mediated cellular events in Lovo cells, and provided a new mechanism for its anti-cancer activity.     

Cbl Enforces an SLP76-dependent Signaling Pathway for T Cell Differentiation

A signaling pathway involving ZAP-70, LAT, and SLP76 has been regarded as essential for receptor-driven T cell development and activation. Consistent with this model, mice deficient in SLP76 have a complete block at the double negative 3 stage of T cell development. Recently, however, it has been reported that inactivation of Cbl, a ubiquitin-protein isopeptide ligase, partially rescues T cell development in SLP76-deficient mice. To probe the influence of Cbl on domain-specific SLP76 functions, we reconstituted SLP76^-/- Cbl^-/- mice with Slp76 transgenes bearing mutations in each of three functional domains of SLP76 as follows: Y3F, in which the amino-terminal tyrosine residues of SLP76 were mutated, eliminating sites of SLP76 interaction with Vav, Nck, and Itk; Δ20, in which 20 amino acids in the proline-rich region of SLP76 were deleted, removing a binding site for Gads; and RK, in which arginine 448 of SLP76 was replaced by lysine, abolishing function of the Src homology 2 domain. Although each of these transgenes has been shown to partially rescue T cell development in SLP76^-/- mice, we report here that Cbl inactivation completely reverses the severe double negative 3 developmental block that occurs in SLP76-deficient mice expressing the Y3F transgene (Y3F mice) and partially rescues the defect in positive selection in T cell receptor transgenic Y3F mice, but in contrast fails to rescue thymic development of SLP76-deficient mice expressing the Δ20 or RK transgene. Rescue in SLP76^-/-Cbl^-/-Y3F double-positive thymocytes is associated with enhanced tyrosine phosphorylation of signaling molecules, including Lck, Vav, PLC-γ1, and ERKs, but not Itk, in response to T cell receptor stimulation. Thus, our data demonstrate that Cbl suppresses activation of a bypass signaling pathway and thereby enforces SLP76 dependence of early T cell development.T cell development proceeds through multiple stages that regulate the generation and selection of T cells whose T cell receptors (TCR)² have an appropriate range of affinity for peptides presented by major histocompatibility complex (MHC) molecules (1). Precursors give rise to immature CD4^-CD8^- double negative (DN) cells that can be further divided into DN1, DN2, DN3, and DN4 stages, distinguished by cell surface phenotype as well as by critical events, including expansion of DN3 cells that have successfully rearranged TCRβ and have expressed and signaled through the pre-TCR complex (2). DN3 cells differentiate to the DN4 and then CD4⁺CD8⁺ double-positive (DP) stage following pre-TCR signaling. DP thymocytes rearrange TCRα, express a mature TCRαβ receptor, and develop into mature CD4⁺CD8^- or CD4^-CD8⁺ single-positive (SP) cells through a process of positive and negative selection that is based on signaling through this mature TCR and selection of a T cell repertoire that is tolerant to self but capable of responding to foreign-peptide-MHC (pMHC) complexes (1, 3, 4). Finally, SP cells exit from the thymus as mature T cells capable of recognizing and responding to foreign antigens.The signals from pre-TCR and TCR, which determine the fate of developing thymocytes, have been intensely studied. Ligation of the TCR by pMHC complexes results in activation of a signaling cascade initiated by phosphorylation and activation of TCR-ζ, Lck, and ZAP-70, which in turn phosphorylate downstream targets, including LAT and SLP76. ZAP-70, LAT and SLP76 proteins (3) have been shown to be essential for thymocyte development by studies, including genetic manipulation in mice (5–8). There are essentially no detectable DP or SP thymocytes or peripheral T cells in LAT^-/- or SLP76^-/- mice, in which thymocyte development is blocked at the DN3 stage (5, 7). ZAP70^-/- thymocytes are blocked at the DP stage of T cell development, and ZAP70^-/- mice have very few SP thymocytes or peripheral T cells (6). These studies suggest that signal transduction required for early T cell development proceeds through a pathway that involves critical roles of multiple molecules, including ZAP-70, LAT, and SLP76.SLP76 consists of three functional domains as follows: an amino-terminal domain containing targets for tyrosine phosphorylation, a proline-rich region, and a carboxyl-terminal SH2 domain (9). The amino-terminal tyrosine residues (Tyr-112, Tyr-128, and Tyr-145) are phosphorylated by tyrosine kinases following TCR engagement, enabling SLP76 to interact with Vav, a Rho guanine nucleotide exchange factor, Nck, an adaptor protein, and Itk, a member of Tec family PTK. The proline-rich region of SLP76 has the capacity to bind Gads, a Grb2 homolog, which results in the recruitment of SLP76 to cell surface membrane lipid rafts through binding to LAT following TCR engagement. The carboxyl-terminal SH2 domain of SLP76 interacts with ADAP (adhesion and degranulation-promoting protein) (10) an adaptor protein, and HPK-1, a serine kinase (9). Reconstitution of SLP76-deficient mice with transgenes containing mutations in each of these domains has demonstrated that each region is required for normal thymocyte development (5, 8). Two groups have reconstituted SLP76-deficient mice with T cell-specific expression of wild-type and mutant SLP76 transgenes, including a mutant in which three tyrosine residues (Tyr-112, Tyr-128, and Tyr-145) in the amino-terminal domain of SLP76 were substituted by phenylalanines (Y3F); a mutant in which 20 amino acids (amino acids 224–244) in the proline-rich region of SLP76 were deleted (Δ20); and a mutant in which arginine 448 of SLP76 was replaced by lysine (RK) (11, 12). The profound defects in T cell development and activation that are observed in SLP76 knock-out mice are completely reversed by reconstitution with a wild-type SLP76 transgene. In contrast, however, reconstitution with SLP76 that has been mutated in any of its three functional domains only partially rescues T cell development in SLP76 knock-out mice.c-Cbl (Cbl) is a ubiquitin ligase and adaptor protein (regulator) with multiple domains that associate with multiple molecules involved in signal transduction (13). Thymocytes from Cbl knock-out mice have enhanced cell surface expression of TCR and CD3 in comparison with control mice (14, 15). In addition, it has been observed that phosphorylation of ZAP-70, LAT, and SLP76 is increased in Cbl^-/- mouse thymocytes (14, 15). Recently, we reported that inactivation of Cbl partially rescues T cell development in LAT and SLP76-deficient mice (16), and Myers et al. (17) reported that inactivation of Cbl partially rescues T cell development in ZAP-70-deficient mice. These observations indicate that Cbl mediates requirements for LAT, SLP76, and ZAP-70 by preventing signaling that is capable of supporting T cell differentiation independent of LAT, SLP76, or ZAP-70. However, the rescue of T cell development in these model systems is strikingly incomplete, failing to substantially reconstitute development through the pre-TCR-dependent DN3-DN4 transition and thus failing to generate normal numbers of DP or functionally mature SP thymocytes. These findings suggest that Cbl inactivation functions to enable pathways that are capable of bypassing some but not all of the requirements for ZAP-70, LAT, and SLP76 during T cell development. To define these signaling pathways, normally suppressed by Cbl, that can support T cell development, we assessed the ability of Cbl inactivation to rescue T cell development in the presence of Y3F, Δ20, or RK SLP76 mutant transgenes. In this study, we report that Cbl inactivation completely reverses the DN3-DN4 developmental defect and partially reverses alterations in positive selection in thymocytes of SLP76 knock-out mice reconstituted with the SLP76 mutant Y3F, which lacks amino-terminal phosphotyrosine residues. In contrast, Cbl inactivation has no effect on the thymic developmental defects observed in SLP76 knock-out mice reconstituted with Slp76 transgenes mutated in the proline-rich Gads-binding region (Δ20) or the carboxyl-terminal SH2 domain (RK). Biochemical studies revealed that rescue of development in SLP76^-/-Y3F thymocytes by inactivation of Cbl was marked by reversal of defects in tyrosine phosphorylation of multiple molecules, including Lck, Vav, PLC-γ1, and ERKs in response to TCR stimulation of DP thymocytes. Thus, Cbl normally enforces SLP76 dependence of T cell development by inhibiting an alternative pathway that may be independent of SLP76 association with Vav, Nck, and Itk (18).     

The N-terminus of TDP-43 promotes its oligomerization and enhances DNA binding affinity

TDP-43 is a DNA/RNA-binding protein associated with different neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD-U). Here, the structural and physical properties of the N-terminus on TDP-43 have been carefully characterized through a combination of nuclear magnetic resonance (NMR), circular dichroism (CD) and fluorescence anisotropy studies. We demonstrate for the first time the importance of the N-terminus in promoting TDP-43 oligomerization and enhancing its DNA-binding affinity. An unidentified structural domain in the N-terminus is also disclosed. Our findings provide insights into the N-terminal domain function of TDP-43.     

Regulation of cyclin A localization downstream of Par-1 function is critical for the centrosome orientation checkpoint in Drosophila male germline stem cells

Male germline stem cells (GSCs) in Drosophila melanogaster divide asymmetrically by orienting the mitotic spindle with respect to the niche, a microenvironment that specifies stem cell identity. The spindle orientation is prepared during interphase through stereotypical positioning of the centrosomes. We recently demonstrated that GSCs possess a checkpoint ("the centrosome orientation checkpoint") that monitors correct centrosome orientation prior to mitosis to ensure an oriented spindle and thus asymmetric outcome of the division. Here, we show that Par-1, a serine/threonine kinase that regulates polarity in many systems, is involved in this checkpoint. Par-1 shows a cell cycle-dependent localization to the spectrosome, a germline-specific, endoplasmic reticulum-like organelle. Furthermore, the localization of cyclin A, which is normally localized to the spectrosome, is perturbed in par-1 mutant GSCs. Interestingly, overexpression of mutant cyclin A that does not localize to the spectrosome and mutation in hts, a core component of the spectrosome, both lead to defects in the centrosome orientation checkpoint. We propose that the regulation of cyclin A localization via Par-1 function plays a critical role in the centrosome orientation checkpoint.