Can ID Repetitive Elements Serve as Cis-acting Dendritic Targeting Elements? An In Vivo Study

Dendritic localization of mRNA/RNA involves interaction of cis-elements and trans-factors. Small, non-protein coding dendritic BC1 RNA is thought to regulate translation in dendritic microdomains. Following microinjections into cultured cells, BC1 RNA fused to larger mRNAs appeared to impart transport competence to these chimeras, and its 5′ ID region was proposed as the cis-acting dendritic targeting element. As these ID elements move around rodent genomes and, if transcribed, form a long RNA stem-loop, they might, thereby, lead to new localizations for targeted gene products. To test their targeting ability in vivo we created transgenic mice expressing various ID elements fused to the 3′ UTR of reporter mRNA for Enhanced Green Fluorescent Protein. In vivo, neither ID elements nor the BC1 RNA coding region were capable of transporting EGFP RNA to dendrites, although the 3′ UTR of α-CaMKII mRNA, an established cis-acting element did produce positive results. Other mRNAs containing naturally inserted ID elements are also not found in neuronal dendrites. We conclude that the 5′ ID domain from BC1 RNA is not a sufficient dendritic targeting element for mRNAs in vivo

Preclinical Evaluation of a Replication-Deficient Intranasal ΔNS1 H5N1 Influenza Vaccine

We developed a novel intranasal influenza vaccine approach that is based on the construction of replication-deficient vaccine viruses that lack the entire NS1 gene (ΔNS1 virus). We previously showed that these viruses undergo abortive replication in the respiratory tract of animals. The local release of type I interferons and other cytokines and chemokines in the upper respiratory tract may have a “self-adjuvant effect”, in turn increasing vaccine immunogenicity. As a result, ΔNS1 viruses elicit strong B- and T- cell mediated immune responses.), one dose of vaccine delivered intranasally was sufficient for the induction of antibodies against homologous A/Vietnam/1203/04 and heterologous A/Indonesia/5/05 H5N1 strains.Our findings show that intranasal immunization with the replication deficient H5N1 ΔNS1 vaccine candidate is sufficient to induce a protective immune response against H5N1 viruses. This approach might be attractive as an alternative to conventional influenza vaccines. Clinical evaluation of ΔNS1 pandemic and seasonal influenza vaccine candidates are currently in progress

Single HA2 Mutation Increases the Infectivity and Immunogenicity of a Live Attenuated H5N1 Intranasal Influenza Vaccine Candidate Lacking NS1

Our finding suggests that an efficient intranasal vaccination with a live attenuated H5N1 virus may require a certain level of pH and temperature stability of HA in order to achieve an optimal virus uptake by the nasal epithelial cells and induce a sufficient immune response. The pH of the activation of the H5 HA protein may play a substantial role in the infectivity of HPAIVs for mammals

Retroposed Elements as Archives for the Evolutionary History of Placental Mammals

Reconstruction of the placental mammalian (eutherian) evolutionary tree has undergone diverse revisions, and numerous aspects remain hotly debated. Initial hierarchical divisions based on morphology contained many misgroupings due to features that evolved independently by similar selection processes. Molecular analyses corrected many of these misgroupings and the superordinal hierarchy of placental mammals was recently assembled into four clades. However, long or rapid evolutionary periods, as well as directional mutation pressure, can produce molecular homoplasies, similar characteristics lacking common ancestors. Retroposed elements, by contrast, integrate randomly into genomes with negligible probabilities of the same element integrating independently into orthologous positions in different species. Thus, presence/absence analyses of these elements are a superior strategy for molecular systematics. By computationally scanning more than 160,000 chromosomal loci and judiciously selecting from only phylogenetically informative retroposons for experimental high-throughput PCR applications, we recovered 28 clear, independent monophyly markers that conclusively verify the earliest divergences in placental mammalian evolution. Using tests that take into account ancestral polymorphisms, multiple long interspersed elements and long terminal repeat element insertions provide highly significant evidence for the monophyletic clades Boreotheria (synonymous with Boreoeutheria), Supraprimates (synonymous with Euarchontoglires), and Laurasiatheria. More importantly, two retropositions provide new support for a prior scenario of early mammalian evolution that places the basal placental divergence between Xenarthra and Epitheria, the latter comprising all remaining placentals. Due to its virtually homoplasy-free nature, the analysis of retroposon presence/absence patterns avoids the pitfalls of other molecular methodologies and provides a rapid, unequivocal means for revealing the evolutionary history of organisms

Positions of Retroposed Elements as Landmarks of Evolution on the Bayesian-Based Placental Evolutionary Tree from Murphy et al. [ 2]

<p>The resultant tree is consistent with previous studies [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b001" target="_blank">1</a>, <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b002" target="_blank">2</a>, <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b004" target="_blank">4</a>, <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b005" target="_blank">5</a>, <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b007" target="_blank">7</a>, <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b008" target="_blank">8</a>, <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b010" target="_blank">10</a>, <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b038" target="_blank">38</a>, <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b039" target="_blank">39</a>] in most aspects. Note that the positions of afrotherians and xenarthras have been reversed, based on the presence of two retroposon insertions at node 2. Gray balls represent single insertion events. Supported splitting points are labeled with Arabic numerals. Superordinal clades, in the order shown, were established by Waddell et al. [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b006" target="_blank">6</a>] and supported by several major studies [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b001" target="_blank">1</a>, <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b002" target="_blank">2</a>, <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b007" target="_blank">7</a>, <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b008" target="_blank">8</a>], and are labeled with Roman numerals. The taxa shown represent only those from which we sampled LINEs and LTRs. Dotted lines indicate nodes in need of further confirmation. Asterisks represent retroposon evidence from the literature for monophyly of Afrotheria [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b027" target="_blank">27</a>], Primates [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b018" target="_blank">18</a>], Rodentia [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b045" target="_blank">45</a>], and Cetartiodactyla [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-b026" target="_blank">26</a>]. </p

Representative Alignments of the Presence/Absence Regions Indicating Support for the Five Investigated Evolutionary Divergences

<p>Potential direct repeats are boxed. The 5′ and 3′ ends of the retroposon insertions are partially shown in lower case letters on a gray background. Node designations corresponding to <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0040091#pbio-0040091-g002" target="_blank">Figure 2</a> and the names of the supported monophyletic groups are given above the inserted elements. </p