Mesozoic retroposons reveal parrots as the closest living relatives of passerine birds

The relationships of passerines (such as the well-studied zebra finch) with non-passerine birds is one of the great enigmas of avian phylogenetic research, because decades of extensive morphological and molecular studies yielded highly inconsistent results between and within data sets. Here we show the first application of the virtually homoplasy-free retroposon insertions to this controversy. Our study examined ~200,000 retroposon-containing loci from various avian genomes and retrieved 51 markers resolving early bird phylogeny. Among these, we obtained statistically significant evidence that parrots are the closest and falcons the second-closest relatives of passerines, together constituting the Psittacopasserae and the Eufalconimorphae, respectively. Our new and robust phylogenetic framework has substantial implications for the interpretation of various conclusions drawn from passerines as model organisms. This includes insights of relevance to human neuroscience, as vocal learning (that is, birdsong) probably evolved in the psittacopasseran ancestor, >30 million years earlier than previously assumed

The genome of the green anole lizard and a comparative analysis with birds and mammals

The evolution of the amniotic egg was one of the great evolutionary innovations in the history of life, freeing vertebrates from an obligatory connection to water and thus permitting the conquest of terrestrial environments1. Among amniotes, genome sequences are available for mammals2 and birds3–5, but not for non-avian reptiles. Here we report the genome sequence of the North American green anole lizard, Anolis carolinensis. We find that A. carolinensis microchromosomes are highly syntenic with chicken microchromosomes, yet do not exhibit the high GC and low repeat content that are characteristic of avian microchromosomes3. Also, A. carolinensis mobile elements are very young and diverse – more so than in any other sequenced amniote genome. This lizard genome’s GC content is also unusual in its homogeneity, unlike the regionally variable GC content found in mammals and birds6. We describe and assign sequence to the previously unknown A. carolinensis X chromosome. Comparative gene analysis shows that amniote egg proteins have evolved significantly more rapidly than other proteins. An anole phylogeny resolves basal branches to illuminate the history of their repeated adaptive radiations

Blocking TLR7- and TLR9-mediated IFN-α Production by Plasmacytoid Dendritic Cells Does Not Diminish Immune Activation in Early SIV Infection

Persistent production of type I interferon (IFN) by activated plasmacytoid dendritic cells (pDC) is a leading model to explain chronic immune activation in human immunodeficiency virus (HIV) infection but direct evidence for this is lacking. We used a dual antagonist of Toll-like receptor (TLR) 7 and TLR9 to selectively inhibit responses of pDC but not other mononuclear phagocytes to viral RNA prior to and for 8 weeks following pathogenic simian immunodeficiency virus (SIV) infection of rhesus macaques. We show that pDC are major but not exclusive producers of IFN-α that rapidly become unresponsive to virus stimulation following SIV infection, whereas myeloid DC gain the capacity to produce IFN-α, albeit at low levels. pDC mediate a marked but transient IFN-α response in lymph nodes during the acute phase that is blocked by administration of TLR7 and TLR9 antagonist without impacting pDC recruitment. TLR7 and TLR9 blockade did not impact virus load or the acute IFN-α response in plasma and had minimal effect on expression of IFN-stimulated genes in both blood and lymph node. TLR7 and TLR9 blockade did not prevent activation of memory CD4+ and CD8+ T cells in blood or lymph node but led to significant increases in proliferation of both subsets in blood following SIV infection. Our findings reveal that virus-mediated activation of pDC through TLR7 and TLR9 contributes to substantial but transient IFN-α production following pathogenic SIV infection. However, the data indicate that pDC activation and IFN-α production are unlikely to be major factors in driving immune activation in early infection. Based on these findings therapeutic strategies aimed at blocking pDC function and IFN-α production may not reduce HIV-associated immunopathology. © 2013 Kader et al

Day-4 Myeloid Dendritic Cells Pulsed with Whole Tumor Lysate Are Highly Immunogenic and Elicit Potent Anti-Tumor Responses

“Day-7” myeloid DCs are commonly used in the clinic. However, there is a strong need to develop DCs faster that have the same potent immunostimulatory capacity as “Day-7” myeloid DCs and at the same time minimizing time, labor and cost of DC preparations. Although “2 days” DCs can elicit peptide-specific responses, they have not been demonstrated to engulf, process and present complex whole tumor lysates, which could be more convenient and personalized source of tumor antigens than defined peptides. In this preclinical study, we evaluated the T-cell stimulatory capacity of Day-2, Day-4, and Day-7 cultured monocyte-derived DCs loaded with SKOV3 cell whole lysate prepared by freeze-thaw or by UVB-irradiation followed by freeze-thaw, and matured with lipopolysaccharide (LPS) and interferon (IFN)-gamma. DCs were evaluated for antigen uptake, and following maturation with LPS and IFN-gamma, DCs were assessed for expression of CD80, CD40, CD86, ICAM-1 and CCR7, production of IL-12p70 and IP-10, and induction of tumor-specific T-cell responses. Day-4 and Day-7 DCs exhibited similar phagocytic abilities, which were superior to Day-2 DCs. Mature Day-7 DCs expressed the highest CD40 and ICAM-1, but mature Day-4 DCs produced the most IL-12p70 and IP-10. Importantly, Day-4 and Day-7 DCs derived from ovarian cancer patients stimulated equally strongly tumor-specific T-cell responses. This is the first study demonstrating the highly immunogenic and strong T-cell stimulatory properties of Day-4 myeloid DCs, and provided important preclinical data for rapid development of potent whole tumor lysate-loaded DC vaccines that are applicable to many tumor types

A DNA Vaccine against Chikungunya Virus Is Protective in Mice and Induces Neutralizing Antibodies in Mice and Nonhuman Primates

Chikungunya virus (CHIKV) is an emerging mosquito-borne alphavirus indigenous to tropical Africa and Asia. Acute illness is characterized by fever, arthralgias, conjunctivitis, rash, and sometimes arthritis. Relatively little is known about the antigenic targets for immunity, and no licensed vaccines or therapeutics are currently available for the pathogen. While the Aedes aegypti mosquito is its primary vector, recent evidence suggests that other carriers can transmit CHIKV thus raising concerns about its spread outside of natural endemic areas to new countries including the U.S. and Europe. Considering the potential for pandemic spread, understanding the development of immunity is paramount to the development of effective counter measures against CHIKV. In this study, we isolated a new CHIKV virus from an acutely infected human patient and developed a defined viral challenge stock in mice that allowed us to study viral pathogenesis and develop a viral neutralization assay. We then constructed a synthetic DNA vaccine delivered by in vivo electroporation (EP) that expresses a component of the CHIKV envelope glycoprotein and used this model to evaluate its efficacy. Vaccination induced robust antigen-specific cellular and humoral immune responses, which individually were capable of providing protection against CHIKV challenge in mice. Furthermore, vaccine studies in rhesus macaques demonstrated induction of nAb responses, which mimicked those induced in convalescent human patient sera. These data suggest a protective role for nAb against CHIKV disease and support further study of envelope-based CHIKV DNA vaccines

A sequence-based genetic linkage map as a reference for Brassica rapa pseudochromosome assembly

<p>Abstract</p> <p>Background</p> <p><it>Brassica rapa </it>is an economically important crop and a model plant for studies concerning polyploidization and the evolution of extreme morphology. The multinational <it>B. rapa </it>Genome Sequencing Project (BrGSP) was launched in 2003. In 2008, next generation sequencing technology was used to sequence the <it>B. rapa </it>genome. Several maps concerning <it>B. rapa </it>pseudochromosome assembly have been published but their coverage of the genome is incomplete, anchoring approximately 73.6% of the scaffolds on to chromosomes. Therefore, a new genetic map to aid pseudochromosome assembly is required.</p> <p>Results</p> <p>This study concerns the construction of a reference genetic linkage map for <it>Brassica rapa</it>, forming the backbone for anchoring sequence scaffolds of the <it>B. rapa </it>genome resulting from recent sequencing efforts. One hundred and nineteen doubled haploid (DH) lines derived from microspore cultures of an F1 cross between a Chinese cabbage (<it>B. rapa </it>ssp. <it>pekinensis</it>) DH line (Z16) and a rapid cycling inbred line (L144) were used to construct the linkage map. PCR-based insertion/deletion (InDel) markers were developed by re-sequencing the two parental lines. The map comprises a total of 507 markers including 415 InDels and 92 SSRs. Alignment and orientation using SSR markers in common with existing <it>B. rapa </it>linkage maps allowed ten linkage groups to be identified, designated A01-A10. The total length of the linkage map was 1234.2 cM, with an average distance of 2.43 cM between adjacent marker loci. The lengths of linkage groups ranged from 71.5 cM to 188.5 cM for A08 and A09, respectively. Using the developed linkage map, 152 scaffolds were anchored on to the chromosomes, encompassing more than 82.9% of the <it>B. rapa </it>genome. Taken together with the previously available linkage maps, 183 scaffolds were anchored on to the chromosomes and the total coverage of the genome was 88.9%.</p> <p>Conclusions</p> <p>The development of this linkage map is vital for the integration of genome sequences and genetic information, and provides a useful resource for the international <it>Brassica </it>research community.</p

Analyses of carnivore microsatellites and their intimate association with tRNA-derived SINEs

BACKGROUND: The popularity of microsatellites has greatly increased in the last decade on account of their many applications. However, little is currently understood about the factors that influence their genesis and distribution among and within species genomes. In this work, we analyzed carnivore microsatellite clones from GenBank to study their association with interspersed repeats and elucidate the role of the latter in microsatellite genesis and distribution. RESULTS: We constructed a comprehensive carnivore microsatellite database comprising 1236 clones from GenBank. Thirty-three species of 11 out of 12 carnivore families were represented, although two distantly related species, the domestic dog and cat, were clearly overrepresented. Of these clones, 330 contained tRNA(Lys)-derived SINEs and 357 contained other interspersed repeats. Our rough estimates of tRNA SINE copies per haploid genome were much higher than published ones. Our results also revealed a distinct juxtaposition of AG and A-rich repeats and tRNA(Lys)-derived SINEs suggesting their coevolution. Both microsatellites arose repeatedly in two regions of the insterspersed repeat. Moreover, microsatellites associated with tRNA(Lys)-derived SINEs showed the highest complexity and less potential instability. CONCLUSION: Our results suggest that tRNA(Lys)-derived SINEs are a significant source for microsatellite generation in carnivores, especially for AG and A-rich repeat motifs. These observations indicate two modes of microsatellite generation: the expansion and variation of pre-existing tandem repeats and the conversion of sequences with high cryptic simplicity into a repeat array; mechanisms which are not specific to tRNA(Lys)-derived SINEs. Microsatellite and interspersed repeat coevolution could also explain different distribution of repeat types among and within species genomes. Finally, due to their higher complexity and lower potential informative content of microsatellites associated with tRNA(Lys)-derived SINEs, we recommend avoiding their use as genetic markers

CD4+ T Cell Effects on CD8+ T Cell Location Defined Using Bioluminescence

T lymphocytes of the CD8+ class are critical in delivering cytotoxic function and in controlling viral and intracellular infections. These cells are “helped” by T lymphocytes of the CD4+ class, which facilitate their activation, clonal expansion, full differentiation and the persistence of memory. In this study we investigated the impact of CD4+ T cells on the location of CD8+ T cells, using antibody-mediated CD4+ T cell depletion and imaging the antigen-driven redistribution of bioluminescent CD8+ T cells in living mice. We documented that CD4+ T cells influence the biodistribution of CD8+ T cells, favoring their localization to abdominal lymph nodes. Flow cytometric analysis revealed that this was associated with an increase in the expression of specific integrins. The presence of CD4+ T cells at the time of initial CD8+ T cell activation also influences their biodistribution in the memory phase. Based on these results, we propose the model that one of the functions of CD4+ T cell “help” is to program the homing potential of CD8+ T cells