Atrial septal defect is one of the most common forms of congenital heart malformation. We identified a new locus linked with atrial septal defect on chromosome 14q12 in a large family with dominantly inherited atrial septal defect. The underlying mutation is a missense substitution, I820N, in alpha-myosin heavy chain (MYH6), a structural protein expressed at high levels in the developing atria, which affects the binding of the heavy chain to its regulatory light chain. The cardiac transcription factor TBX5 strongly regulates expression of MYH6, but mutant forms of TBX5, which cause Holt-Oram syndrome, do not. Morpholino knock-down of expression of the chick MYH6 homolog eliminates the formation of the atrial septum without overtly affecting atrial chamber formation. These data provide evidence for a link between a transcription factor, a structural protein and congenital heart disease.

A region-specific ENU mutagenesis screen was conducted to elucidate the functional content of proximal mouse Chr 5. We used the visibly marked, recessive, lethal inversion Rump White (Rw) as a balancer in a three-generation breeding scheme to identify recessive mutations within the ∼50 megabases spanned by Rw. A total of 1003 pedigrees were produced, representing the largest inversion screen performed in mice. Test-class animals, homozygous for the ENU-mutagenized proximal Chr 5 and visibly distinguishable from nonhomozygous littermates, were screened for fertility, hearing, vestibular function, DNA repair, behavior, and dysmorphology. Lethals were identifiable by failure to derive test-class animals within a pedigree. Embryonic lethal mutations (total of 34) were overwhelmingly the largest class of mutants recovered. We characterized them with respect to the time of embryonic death, revealing that most act at midgestation (8.5-10.5) or sooner. To position the mutations within the Rw region and to guide allelism tests, we performed complementation analyses with a set of new and existing chromosomal deletions, as well as standard recombinational mapping on a subset of the mutations. By pooling the data from this and other region-specific mutagenesis projects, we calculate that the mouse genome contains ∼3479-4825 embryonic lethal genes, or about 13.7%-19% of all genes.

Background and aimsBacteroides fragilis (BF) are Gram-negative anaerobe symbionts present in the colon. Recent studies have reported the beneficial role of BF in maintaining intestinal homeostasis, stimulating host immunologic development, and preventing infectious colitis caused by pathogenic bacteria. Our previous studies showed that monocolonization of germ-free mice with BF significantly reduced colon inflammations and damage.MethodsIn order to investigate the Toll-like receptor-2 (TLR2), TLR4, and interleukin 10 (IL-10) molecular signaling pathways involved in BF-mediated prevention of dextran sulfate sodium (DSS)-induced colitis. The wild-type (WT), TLR4, TLR2, and IL-10 knockout (-/-) germ-free mice grown were with or without BF colonization for 28 days, and then administered 1% DSS in drinking water for 7 day to induce acute ulcerative colitis.ResultsWe compared phenotypes such as weight loss, disease activity, intestinal histological scores, and immunohistochemistry for inflammatory cells. Unlike WT and TLR4-/- mice, the severity of DSS-colitis did not improve in TLR2-/- animals after BF colonization. The BF enhanced anti-inflammatory cytokines IL-10 expression and inhibited pro-inflammatory-related tumor necrosis factor (TNF-α) and IL-6 mRNA expression in both WT and TLR4-/- mice. In contrast, the failed to up-regulated IL-10 and down-regulated the TNF-α and IL-6 in BF colonization TLR2-/- mice. In addition, we further perform IL-10-/- mice to clarify whether the BF through TLR2 /IL-10 pathway to alleviate DSS-colitis. There were no significant differences in colitis severity and pro-inflammatory related genes expression in the IL-10-/- mice with or without BF colonization.ConclusionsThese results indicate the disease-preventing effects of BF in acute DSS-induced colitis may occur through the TLR2/IL-10 signal pathway.

To sense numerous odorants and chemicals, animals have evolved a large number of olfactory receptor genes (Olfrs) in their genome. In particular, the house mouse has ∼1,100 genes in the Olfr gene family. This makes the mouse a good model organism to study Olfr genes and olfaction-related genes. To date, whether male and female mice possess the same ability in detecting environmental odorants is still unknown. Using the next generation sequencing technology (paired-end mRNA-seq), we detected 1,088 expressed Olfr genes in both male and female olfactory epithelium. We found that not only Olfr genes but also odorant-binding protein (Obp) genes have evolved rapidly in the mouse lineage. Interestingly, Olfr genes tend to express at a higher level in males than in females, whereas the Obp genes clustered on the X chromosome show the opposite trend. These observations may imply a more efficient odorant-transporting system in females, whereas a more active Olfr gene expressing system in males. In addition, we detected the expression of two genes encoding major urinary proteins, which have been proposed to bind and transport pheromones or act as pheromones in mouse urine. This observation suggests a role of main olfactory system (MOS) in pheromone detection, contrary to the view that only accessory olfactory system (AOS) is involved in pheromone detection. This study suggests the sexual differences in detecting environmental odorants in MOS and demonstrates that mRNA-seq provides a powerful tool for detecting genes with low expression levels and with high sequence similarities.

Ulcerative colitis is inflammatory conditions of the colon caused by interplay of genetic and environmental factors. Previous studies indicated that the gut microflora may be involved in the colonic inflammation. Bacteroides fragilis (BF) is a Gram-negative anaerobe belonging to the colonic symbiotic. We aimed to investigate the protective role of BF in a colitis model induced in germ-free (GF) mice by dextran sulfate sodium (DSS). GF C57BL/6JNarl mice were colonized with BF for 28 days before acute colitis was induced by DSS. BF colonization significantly increased animal survival by 40%, with less reduction in colon length, and decreased infiltration of inflammatory cells (macrophages and neutrophils) in colon mucosa following challenge with DSS. In addition, BF could enhance the mRNA expression of anti-inflammatory-related cytokine such as interleukin 10 (IL-10) with polymorphism cytokine IL-17 and diminish that of proinflammatory-related tumor necrosis factor α with inducible nitric oxide synthase in the ulcerated colon. Myeloperoxidase activity was also decreased in BF-DSS mice. Taking these together, the BF colonization significantly ameliorated DSS-induced colitis by suppressing the activity of inflammatory-related molecules and inducing the production of anti-inflammatory cytokines. BF may play an important role in maintaining intestinal immune system homeostasis and regulate inflammatory responses.

A selective sweep is the result of strong positive selection driving newly occurring or standing genetic variants to fixation, and can dramatically alter the pattern and distribution of allelic diversity in a population. Population-level sequencing data have enabled discoveries of selective sweeps associated with genes involved in recent adaptations in many species. In contrast, much debate but little evidence addresses whether "selfish" genes are capable of fixation-thereby leaving signatures identical to classical selective sweeps-despite being neutral or deleterious to organismal fitness. We previously described R2d2, a large copy-number variant that causes nonrandom segregation of mouse Chromosome 2 in females due to meiotic drive. Here we show population-genetic data consistent with a selfish sweep driven by alleles of R2d2 with high copy number (R2d2 HC ) in natural populations. We replicate this finding in multiple closed breeding populations from six outbred backgrounds segregating for R2d2 alleles. We find that R2d2 HC rapidly increases in frequency, and in most cases becomes fixed in significantly fewer generations than can be explained by genetic drift. R2d2HC is also associated with significantly reduced litter sizes in heterozygous mothers, making it a true selfish allele. Our data provide direct evidence of populations actively undergoing selfish sweeps, and demonstrate that meiotic drive can rapidly alter the genomic landscape in favor of mutations with neutral or even negative effects on overall Darwinian fitness. Further study will reveal the incidence of selfish sweeps, and will elucidate the relative contributions of selfish genes, adaptation and genetic drift to evolution.

BackgroundForward genetic screens in mice provide an unbiased means to identify genes and other functional genetic elements in the genome. Previously, a large scale ENU mutagenesis screen was conducted to query the functional content of a ~50 Mb region of the mouse genome on proximal Chr 5. The majority of phenotypic mutants recovered were embryonic lethals.ResultsWe report the high resolution genetic mapping, complementation analyses, and positional cloning of mutations in the target region. The collection of identified alleles include several with known or presumed functions for which no mutant models have been reported (Tbc1d14, Nol14, Tyms, Cad, Fbxl5, Haus3), and mutations in genes we or others previously reported (Tapt1, Rest, Ugdh, Paxip1, Hmx1, Otoe, Nsun7). We also confirmed the causative nature of a homeotic mutation with a targeted allele, mapped a lethal mutation to a large gene desert, and localized a spermiogenesis mutation to a region in which no annotated genes have coding mutations. The mutation in Tbc1d14 provides the first implication of a critical developmental role for RAB-GAP-mediated protein transport in early embryogenesis.ConclusionThis collection of alleles contributes to the goal of assigning biological functions to all known genes, as well as identifying novel functional elements that would be missed by reverse genetic approaches.

Epstein-Barr Virus (EBV) Nuclear Antigen 1 (EBNA1)-mediated origin of plasmid replication (oriP) DNA episome maintenance is essential for EBV-mediated tumorigenesis. We have now found that EBNA1 binds to Ribosome Protein L4 (RPL4). RPL4 shRNA knockdown decreased EBNA1 activation of an oriP luciferase reporter, EBNA1 DNA binding in lymphoblastoid cell lines, and EBV genome number per lymphoblastoid cell line. EBV infection increased RPL4 expression and redistributed RPL4 to cell nuclei. RPL4 and Nucleolin (NCL) were a scaffold for an EBNA1-induced oriP complex. The RPL4 N terminus cooperated with NCL-K429 to support EBNA1 and oriPmediated episome binding and maintenance, whereas the NCL C-terminal K380 and K393 induced oriP DNA H3K4me2 modification and promoted EBNA1 activation of oriP-dependent transcription. These observations provide new insights into the mechanisms by which EBV uses NCL and RPL4 to establish persistent B-lymphoblastoid cell infection.pstein-Barr virus (EBV) was recognized as an oncogenic human pathogen after the discovery of its causal associations with B-cell lymphomas (BLs), nasopharyngeal carcinomas, and gastric carcinomas. EBV infects B-lymphocytes and epithelial cells and converts resting B cells into lymphoblastoid cell lines (LCLs). LCL maintenance requires expression of EBV nuclear antigens (EBNAs), latency-associated membrane proteins (LMPs), and noncoding RNAs (1). EBNA1 is the only EBV gene expressed in all types of EBV-infected cells and has a key role in EBV genome maintenance, replication, postmitotic EBV genome segregation, and LCL growth (1, 2). EBNA1-mediated episome maintenance depends on EBNA1 binding to the EBV origin of genome replication (oriP), which has two essential components, a dyad symmetry (DS) element and a family of repeats (FR) (3). Despite a 2.4-Å resolution crystal structure of the EBNA1 DNA binding domain bound to its cognate DNA element (4), mechanistic insights into EBNA1 and oriP-mediated episome maintenance mainly come from genetic studies using EBV recombinants and biochemical studies of EBNA1's association with cell proteins, including CTCF, Bromodomain Protein 4 (BRD4), Nucleosome Assembly Protein 1 (NAP1), the cell Origin Recognition Complex, and the Mini Chromosome Maintenance complex (5-8). Recent studies indicate that EBNA1 may use complex strategies for episome maintenance (9-16).EBNA1-associated ribosome biogenesis factors Nucleophosmin (NPM1) and Nucleolin (NCL) have been implicated in EBNA1 and oriP-dependent functions (17,18). Other viruses also use ribosomal proteins (RPs), such as RPL4, RPS19, and RPL40, to enhance virus protein translation (19-21). Indeed "extraribosomal functions" of RPs were discovered through RPS1 involvement in bacteriophage Qβ-mediated genome replication (22). Moreover, the noncoding EBV RNA, EBER1, causes RPL22 redistribution from the nucleolus to the nucleoplasm and stimulates cell proliferation (23, 24). We have now found complex protein interactions among EBNA1, RPL4, and NCL and have examined the role of these in...