Virmid: accurate detection of somatic mutations with sample impurity inference

Detection of somatic variation using sequence from disease-control matched data sets is a critical first step. In many cases including cancer, however, it is hard to isolate pure disease tissue, and the impurity hinders accurate mutation analysis by disrupting overall allele frequencies. Here, we propose a new method, Virmid, that explicitly determines the level of impurity in the sample, and uses it for improved detection of somatic variation. Extensive tests on simulated and real sequencing data from breast cancer and hemimegalencephaly demonstrate the power of our model. A software implementation of our method is available at http://sourceforge.net/projects/virmid/.     

Whole-Exome Sequencing Identifies Mutated C12orf57 in Recessive Corpus Callosum Hypoplasia

The corpus callosum is the principal cerebral commissure connecting the right and left hemispheres. The development of the corpus callosum is under tight genetic control, as demonstrated by abnormalities in its development in more than 1,000 genetic syndromes. We recruited more than 25 families in which members affected with corpus callosum hypoplasia (CCH) lacked syndromic features and had consanguineous parents, suggesting recessive causes. Exome sequence analysis identified C12orf57 mutations at the initiator methionine codon in four different families. C12orf57 is ubiquitously expressed and encodes a poorly annotated 126 amino acid protein of unknown function. This protein is without significant paralogs but has been tightly conserved across evolution. Our data suggest that this conserved gene is required for development of the human corpus callosum.     

A framework for estimating the sensitivity of eDNA surveys

Imperfect sensitivity, or imperfect detection, is a feature of all survey methods that needs to be accounted for when interpreting survey results. Detection of environmental DNA (eDNA) is increasingly being used to infer species distributions, yet the sensitivity of the technique has not been fully evaluated. Sensitivity, or the probability of detecting target DNA given it is present at a site, will depend on both the survey method and the concentration and dispersion of target DNA molecules at a site. We present a model to estimate target DNA concentration and dispersion at survey sites and to estimate the sensitivity of an eDNA survey method. We fitted this model to data from a species‐specific eDNA survey for Oriental weatherloach, Misgurnus anguillicaudatus, at three sites sampled in both autumn and spring. The concentration of target DNA molecules was similar at all three sites in autumn but much higher at two sites in spring. Our analysis showed the survey method had ≥95% sensitivity at sites where target DNA concentrations were ≥11 molecules per litre. We show how these data can be used to compare sampling schemes that differ in the number of field samples collected per site and number of PCR replicates per sample to achieve ≥95% sensitivity at a given target DNA concentration. These models allow researchers to quantify the sensitivity of eDNA survey methods to optimize the probability of detecting target species, and to compare DNA concentrations spatially and temporarily.     

In silico identification and in vitro validation of nogalamycin N-oxide (NSC116555) as a potent anticancer compound against non–small-cell lung cancer cells

The epidermal growth factor receptor (EGFR) was found to be overexpressed in several cancers, especially in lung cancers. Finding new effective drug against EGFR is the key to cancer treatment. In this study, the GOLD docking algorithm was used to virtually screen for novel human EGFR inhibitors from the NCI database. Thirty-four hit compounds were tested for EGFR-tyrosine kinase (TK) inhibition. Two potent compounds, 1-amino-4-(4-[4-amino-2-sulfophenyl]anilino)-9,10-dioxoanthracene-2-sulfonic acid (NSC125910), and nogalamycin N-oxide (NSC116555) were identified with IC₅₀ values against EGFR-TK comparable to gefitinib; 16.14 and 37.71 nM, respectively. However, only NSC116555 demonstrated cytotoxic effects against non–small-cell lung cancer, A549, shown in the cell cytotoxicity assay with an IC₅₀ of 0.19 + 0.01 µM, which was more potent than gefitinib. Furthermore, NSC116555 showed cytotoxicity against A549 via apoptosis in a dose-dependent manner.     

A novel Golgi-localisation domain shared by a class of coiled-coil peripheral membrane proteins

The mechanism by which peripheral membrane proteins are targeted to the cytoplasmic face of the Golgi apparatus is poorly understood. Previously, we have identified a carboxy-terminal domain of the trans-Golgi-network (TGN) protein p230 that is responsible for Golgi localisation [1]. Here, we report the identification of a similar Golgi-localisation domain (GLD, also termed the 'GRIP' domain - see the paper by Munro and Nichols elsewhere in this issue) in a family of putative peripheral membrane proteins from lower and higher eucaryotes. The majority of family members have a domain structure similar to that of p230, with extensive coiled-coil regions (>80%) and the potential GLD located in a non-coiled-coil domain at the carboxyl terminus. Previously reported proteins in this family include human golgin-97 and Saccharomyces cerevisiae Imh1p. By constructing chimeric cDNAs encoding carboxy-terminal regions of these family members fused to green fluorescent protein (GFP), we have directly demonstrated that the GLD of p230, golgin-97, the newly identified human protein GCC1p and yeast Imh1p functions as a Golgi-targeting domain in transfected mammalian cells. Site-directed mutagenesis of the GLDs identified two conserved aromatic residues that are critical for the function of this targeting domain. Endogenous p230 was displaced from the Golgi membranes in transfected cells expressing high levels of GFP fused to the GLD of either p230 or golgin-97, indicating that different GLDs interact with similar membrane determinants. Thus, we have identified a family of coiled-coil proteins that share a domain shown to be sufficient for the localisation of peripheral membrane proteins to the Golgi apparatus.     

The insulin-like growth factor I generation test in adults

The insulin-like growth factor I (IGF-I) generation test has the potential to assess the ability of an individual to respond to an acute bolus of growth hormone (GH), in terms of IGF-I, IGF-binding protein 3 and acid-labile subunit responses. This article will discuss something of the history of the IGF-I generation test, and review some of the major studies to date. The IGF-I generation test was first used in adults by Lieberman et al., who studied the effects of ageing and oestrogen administration, and suggested that decreased responsiveness to GH occurs with increasing age and oral oestrogen administration. Our results, however, show that, while activity of the GH/IGF-I axis declines with age, peripheral responsiveness to GH is not affected. As in the Lieberman study, we found that oral oestrogen replacement reduces responses of GH-dependent peptides to GH stimulation in healthy post-menopausal women. Transdermal oestrogen administration also reduced responsiveness to GH, although to a lesser degree than orally administered oestrogen. In addition, utilizing a non-weight-based dose of GH we have demonstrated that obese individuals produce greater increases in IGF-I following an acute bolus of GH. In GH deficiency (GHD), data suggesting enhanced peripheral responsiveness should be interpreted with caution, and with awareness of differences between these groups in terms of age and obesity. The IGF-I generation test may allow a fresh approach to unanswered questions in the field of GHD, but as the IGF-I response to GH is not strictly associated with protein anabolism or clinical benefit, the question remains whether this test will predict the effect of longer-term GH administration.