Cloning and characterization of the platypus mitochondrial genome

The vertebrate mitochondrial genome is highly conserved in size and gene content. Among the chordates there appears to be one basic gene arrangement, but rearrangements in the mitochondrial gene order of the avian lineages have indicated that the mitochondrial genome may be more variable than once thought. Different gene orders in marsupials and eutherian mammals leave the ancestral mammalian order in some doubt. We have investigated the mitochondrial gene order in the platypus (Ornithorhynchus anatinus), a representative of the third major group of mammals, to determine which mitochondrial gene arrangement is ancestral in mammals. We have found that the platypus mtDNA conforms to the basic chordate gene arrangement, common to fish, amphibians, and eutherian mammals, indicating that this arrangement was the original mammalian arrangement, and that the unusual rearrangements observed in the avians and marsupials are probably lineage-specific. Correspondence to: N.J. Gemmell     

LncRNA and mRNA integration network reconstruction reveals novel key regulators in esophageal squamous-cell carcinoma

Many experimental and computational studies have identified key protein coding genes in initiation and progression of esophageal squamous cell carcinoma (ESCC). However, the number of researches that tried to reveal the role of long non-coding RNAs (lncRNAs) in ESCC has been limited. LncRNAs are one of the important regulators of cancers which are transcribed dominantly in the genome and in various conditions. The main goal of this study was to use a systems biology approach to predict novel lncRNAs as well as protein coding genes associated with ESCC and assess their prognostic values. By using microarray expression data for mRNAs and lncRNAs from a large number of ESCC patients, we utilized “Weighted Gene Co-expression Network Analysis” (WGCNA) method to make a big coding-non-coding gene co-expression network, and discovered important functional modules. Gene set enrichment and pathway analysis revealed major biological processes and pathways involved in these modules. After selecting some protein coding genes involved in biological processes and pathways related to cancer, we used “LncTar”, a computational tool to predict potential interactions between these genes and lncRNAs. By combining interaction results with Pearson correlations, we introduced some novel lncRNAs with putative key regulatory roles in the network. Survival analysis with Kaplan-Meier estimator and Log-rank test statistic confirmed that most of the introduced genes are associated with poor prognosis in ESCC. Overall, our study reveals novel protein coding genes and lncRNAs associated with ESCC, along with their predicted interactions. Based on the promising results of survival analysis, these genes can be used as good estimators of patients' survival, or even can be analyzed further as new potential signatures or targets for the therapy of ESCC disease.     

Suppression of RNA interference increases alphavirus replication and virus-associated mortality in Aedes aegypti mosquitoes

Background  

Arthropod-borne viruses (arboviruses) can persistently infect and cause limited damage to mosquito vectors. RNA interference (RNAi) is a mosquito antiviral response important in restricting RNA virus replication and has been shown to be active against some arboviruses. The goal of this study was to use a recombinant Sindbis virus (SINV; family Togaviridae; genus Alphavirus) that expresses B2 protein of Flock House virus (FHV; family Nodaviridae; genus Alphanodavirus), a protein that inhibits RNAi, to determine the effects of linking arbovirus infection with RNAi inhibition.     

A place to land: spatiotemporal drivers of stopover habitat use by migrating birds

Migrating birds require en route habitats to rest and refuel. Yet, habitat use has never been integrated with passage to understand the factors that determine where and when birds stopover during spring and autumn migration. Here, we introduce the stopover‐to‐passage ratio (SPR), the percentage of passage migrants that stop in an area, and use 8 years of data from 12 weather surveillance radars to estimate over 50% SPR during spring and autumn through the Gulf of Mexico and Atlantic coasts of the south‐eastern US, the most prominent corridor for North America’s migratory birds. During stopovers, birds concentrated close to the coast during spring and inland in forested landscapes during autumn, suggesting seasonal differences in habitat function and highlighting the vital role of stopover habitats in sustaining migratory communities. Beyond advancing understanding of migration ecology, SPR will facilitate conservation through identification of sites that are disproportionally selected for stopover by migrating birds.     

Roles of mouse sperm‐associated alpha‐L‐fucosidases in fertilization

Sperm‐associated α‐L ‐fucosidases have been implicated in fertilization in many species. Previously, we documented the existence of α‐L ‐fucosidase in mouse cauda epididymal contents, and showed that sperm‐associated α‐L ‐fucosidase is cryptically stored within the acrosome and reappears within the sperm equatorial segment after the acrosome reaction. The enrichment of sperm membrane‐associated α‐L ‐fucosidase within the equatorial segment of acrosome‐reacted cells implicates its roles during fertilization. Here, we document the absence of α‐L ‐fucosidase in mouse oocytes and early embryos, and define roles of sperm associated α‐L ‐fucosidase in fertilization using specific inhibitors and competitors. Mouse sperm were pretreated with deoxyfuconojirimycin (DFJ, an inhibitor of α‐L ‐fucosidase) or with anti‐fucosidase antibody; alternatively, mouse oocytes were pretreated with purified human liver α‐L ‐fucosidase. Five‐millimolar DFJ did not inhibit sperm–zona pellucida (ZP) binding, membrane binding, or fusion and penetration, but anti‐fucosidase antibody and purified human liver α‐L ‐fucosidase significantly decreased the frequency of these events. To evaluate sperm‐associated α‐L ‐fucosidase enzyme activity in post‐fusion events, DFJ‐pretreated sperm were microinjected into oocytes, and 2‐pronuclear (2‐PN) embryos were treated with 5 mM DFJ with no significant effects, suggesting that α‐L ‐fucosidase enzyme activity does not play a role in post‐fusion events and/or early embryo development in mice. The recognition and binding of mouse sperm to the ZP and oolemma involves the glycoprotein structure of α‐L ‐fucosidase, but not its catalytic action. These observations suggest that deficits in fucosidase protein and/or the presence of anti‐fucosidase antibody may be responsible for some types of infertility. Mol. Reprod. Dev. 80: 273–285, 2013. © 2013 Wiley Periodicals, Inc.     

Right Bundle Branch Block: A Predictor of Mortality in Early Systemic Sclerosis

Objective

To evaluate the prognostic significance of baseline electrocardiogram (ECG) abnormalities in a multiethnic cohort of patients with early systemic sclerosis (SSc) and to determine the serological, clinical, and echocardiogram correlates of ECG findings.

Methods

SSc patients with disease duration of≤5 years were enrolled in the GENISOS (Genetics versus Environment in Scleroderma Outcome Study) cohort. At the first visit, a standard 12 lead ECG was obtained along with demographic information, clinical data, and autoantibodies. The results of echocardiograms were also recorded. All ECGs were interpreted by a cardiologist unaware of the patients'' clinical data.

Results

Of 265 SSc patients with average disease duration at enrollment of 2.5 years, 140 (52.8%) had abnormal ECG findings. These findings were not associated with SSc disease type or autoantibody profile but were associated with more severe heart and lung involvement. A total of 75 patients (28.3%) died over a follow up time of 9.9 years. Complete right bundle branch block (± left anterior hemiblock) on ECG, present in 7 (2.6%) patients, predicted a higher risk of mortality (HR: 5.3; 95% CI: 2.1 to 13.4; p<0.001). The predictive significance of right bundle branch block was independent of age at enrollment, gender, ethnicity and risk factors for coronary artery disease.

Conclusion

ECG abnormalities are common in patients with early SSc and are associated with the severity of lung and heart involvement. Right bundle branch block is an independent predictor of mortality, and should be considered a marker of disease severity in SSc.     

Compensatory Hemagglutinin Mutations Alter Antigenic Properties of Influenza Viruses

Influenza viruses routinely acquire mutations in antigenic sites on the globular head of the hemagglutinin (HA) protein. Since these antigenic sites are near the receptor binding pocket of HA, many antigenic mutations simultaneously alter the receptor binding properties of HA. We previously reported that a K165E mutation in the Sa antigenic site of A/Puerto Rico/8/34 (PR8) HA is associated with secondary neuraminidase (NA) mutations that decrease NA activity. Here, using reverse genetics, we show that the K165E HA mutation dramatically decreases HA binding to sialic acid receptors on cell surfaces. We sequentially passaged reverse-genetics-derived PR8 viruses with the K165E antigenic HA mutation in fertilized chicken eggs, and to our surprise, viruses with secondary NA mutations did not emerge. Instead, viruses with secondary HA mutations emerged in 3 independent passaging experiments, and each of these mutations increased HA binding to sialic acid receptors. Importantly, these compensatory HA mutations were located in the Ca antigenic site and prevented binding of Ca-specific monoclonal antibodies. Taken together, these data indicate that HA antigenic mutations that alter receptor binding avidity can be compensated for by secondary HA or NA mutations. Antigenic diversification of influenza viruses can therefore occur irrespective of direct antibody pressure, since compensatory HA mutations can be located in distinct antibody binding sites.     

Reprogramming EF-hands for design of catalytically amplified lanthanide sensors

We recently reported that a computationally designed catalyst nicknamed AlleyCat facilitates C–H proton abstraction in Kemp elimination at neutral pH in a selective and calcium-dependent fashion by a factor of approximately 100,000 (Korendovych et al. in Proc. Natl. Acad. Sci. USA 108:6823, 2011). Kemp elimination produced a colored product that can be easily read out, thus making AlleyCat a catalytically amplified metal sensor for calcium. Here we report that metal-binding EF-hand motifs in AlleyCat could be redesigned to incorporate trivalent metal ions without significant loss of catalytic activity. Mutation of a single neutral residue at position 9 of each of the EF-hands to glutamate results in almost a two orders of magnitude improvement of selectivity for trivalent metal ions over calcium. Development of this new lanthanide-dependent switchable Kemp eliminase, named CuSeCat EE, provides the foundation for further selectivity improvement and broadening the scope of the repertoire of metals for sensing. A concerted effort in the design of switchable enzymes has many environmental, sensing, and metal ion tracking applications.