Primary cilia are specialized calcium signaling organelles

Summary Primary cilia are solitary nonmotile extensions of the centriole found on nearly all nucleated eukaryotic cells between cell divisions. Only ∼200-300 nm in diameter and a few microns long, they are separated from the cytoplasm by the ciliary neck and basal body. Often called sensory cilia, they are hypothesized to receive chemical and mechanical stimuli and initiate specific cellular signal transduction pathways. When activated by a ligand, Hedgehog (Hh) pathway proteins, such as Gli2 and Smoothened (Smo), translocate from the cell into the cilium1,2. Mutations in primary ciliary proteins are associated with severe developmental defects3. The ionic conditions, permeability of the primary cilia membrane, and effectiveness of the diffusion barriers between the cilia and cell body are unknown. Here we show that cilia are a unique calcium compartment regulated by a heteromeric TRP channel, PKD1-L1/PKD2-L1. In contrast to the hypothesis that polycystin (PKD) channels initiate changes in ciliary calcium that are conducted into the cytoplasm4, we show that changes in ciliary calcium concentration ([Ca2+]cilia) occur without substantially altering global cytoplasmic calcium ([Ca2+]cyto). PKD1-L1/PKD2-L1 acts as a ciliary calcium channel controlling [Ca2+]cilia and thereby modifying Smo-activated Gli2 translocation and Gli1 expression

Microorganisms from aphid honeydew attract and enhance the efficacy of natural enemies

Aphids are one of the most serious pests of crops worldwide, causing major yield and economic losses. To control aphids, natural enemies could be an option but their efficacy is sometimes limited by their dispersal in natural environment. Here we report the first isolation of a bacterium from the pea aphid Acyrthosiphon pisum honeydew, Staphylococcus sciuri, which acts as a kairomone enhancing the efficiency of aphid natural enemies. Our findings represent the first case of a host-associated bacterium driving prey location and ovipositional preference for the natural enemy. We show that this bacterium has a key role in tritrophic interactions because it is the direct source of volatiles used to locate prey. Some specific semiochemicals produced by S. sciuri were also identified as significant attractants and ovipositional stimulants. The use of this host-associated bacterium could certainly provide a novel approach to control aphids in field and greenhouse systems

Aphids acquired symbiotic genes via lateral gene transfer

<p>Abstract</p> <p>Background</p> <p>Aphids possess bacteriocytes, which are cells specifically differentiated to harbour the obligate mutualist <it>Buchnera aphidicola </it>(γ-Proteobacteria). <it>Buchnera </it>has lost many of the genes that appear to be essential for bacterial life. From the bacteriocyte of the pea aphid <it>Acyrthosiphon pisum</it>, we previously identified two clusters of expressed sequence tags that display similarity only to bacterial genes. Southern blot analysis demonstrated that they are encoded in the aphid genome. In this study, in order to assess the possibility of lateral gene transfer, we determined the full-length sequences of these transcripts, and performed detailed structural and phylogenetic analyses. We further examined their expression levels in the bacteriocyte using real-time quantitative RT-PCR.</p> <p>Results</p> <p>Sequence similarity searches demonstrated that these fully sequenced transcripts are significantly similar to the bacterial genes <it>ldcA </it>(product, LD-carboxypeptidase) and <it>rlpA </it>(product, rare lipoprotein A), respectively. <it>Buchnera </it>lacks these genes, whereas many other bacteria, including <it>Escherichia coli</it>, a close relative of <it>Buchnera</it>, possess both <it>ldcA </it>and <it>rlpA</it>. Molecular phylogenetic analysis clearly demonstrated that the aphid <it>ldcA </it>was derived from a rickettsial bacterium closely related to the extant <it>Wolbachia </it>spp. (α-Proteobacteria, Rickettsiales), which are intracellular symbionts of various lineages of arthropods. The evolutionary origin of <it>rlpA </it>was not fully resolved, but it was clearly demonstrated that its double-ψ β-barrel domain is of bacterial origin. Real-time quantitative RT-PCR demonstrated that <it>ldcA </it>and <it>rlpA </it>are expressed 11.6 and 154-fold higher in the bacteriocyte than in the whole body, respectively. LdcA is an enzyme required for recycling murein (peptidoglycan), which is a component of the bacterial cell wall. As <it>Buchnera </it>possesses a cell wall composed of murein but lacks <it>ldcA</it>, a high level of expression of the aphid <it>ldcA </it>in the bacteriocyte may be essential to maintain <it>Buchnera</it>. Although the function of RlpA is not well known, conspicuous up-regulation of the aphid <it>rlpA </it>in the bacteriocyte implies that this gene is also essential for <it>Buchnera</it>.</p> <p>Conclusion</p> <p>In this study, we obtained several lines of evidence indicating that aphids acquired genes from bacteria via lateral gene transfer and that these genes are used to maintain the obligately mutualistic bacterium, <it>Buchnera</it>.</p

In vitro and in vivo mRNA delivery using lipid-enveloped pHresponsive polymer nanoparticles

Biodegradable core−shell structured nanoparticles with a poly(β-amino ester) (PBAE) core enveloped by a phospholipid bilayer shell were developed for in vivo mRNA delivery with a view toward delivery of mRNA-based vaccines. The pH-responsive PBAE component was chosen to promote endosome disruption, while the lipid surface layer was selected to minimize toxicity of the polycation core. Messenger RNA was efficiently adsorbed via electrostatic interactions onto the surface of these net positively charged nanoparticles. In vitro, mRNA-loaded particle uptake by dendritic cells led to mRNA delivery into the cytosol with low cytotoxicity, followed by translation of the encoded protein in these difficult-to-transfect cells at a frequency of 30%. Particles loaded with mRNA administered intranasally (i.n.) in mice led to the expression of the reporter protein luciferase in vivo as soon as 6 h after administration, a time point when naked mRNA given i.n. showed no expression. At later time points, luciferase expression was detected in naked mRNA-treated mice, but this group showed a wide variation in levels of transfection, compared to particle-treated mice. This system may thus be promising for noninvasive delivery of mRNA-based vaccines.United States. Dept. of Defense (Institute for Soldier Nanotechnology, contract W911NF-07-D-0004)Ragon Institute of MGH, MIT and HarvardSingapore. Agency for Science, Technology and ResearchHoward Hughes Medical Institute (Investigator

Genome Sequence of the Pea Aphid Acyrthosiphon pisum

Aphids are important agricultural pests and also biological models for studies of insect-plant interactions, symbiosis, virus vectoring, and the developmental causes of extreme phenotypic plasticity. Here we present the 464 Mb draft genome assembly of the pea aphid Acyrthosiphon pisum. This first published whole genome sequence of a basal hemimetabolous insect provides an outgroup to the multiple published genomes of holometabolous insects. Pea aphids are host-plant specialists, they can reproduce both sexually and asexually, and they have coevolved with an obligate bacterial symbiont. Here we highlight findings from whole genome analysis that may be related to these unusual biological features. These findings include discovery of extensive gene duplication in more than 2000 gene families as well as loss of evolutionarily conserved genes. Gene family expansions relative to other published genomes include genes involved in chromatin modification, miRNA synthesis, and sugar transport. Gene losses include genes central to the IMD immune pathway, selenoprotein utilization, purine salvage, and the entire urea cycle. The pea aphid genome reveals that only a limited number of genes have been acquired from bacteria; thus the reduced gene count of Buchnera does not reflect gene transfer to the host genome. The inventory of metabolic genes in the pea aphid genome suggests that there is extensive metabolite exchange between the aphid and Buchnera, including sharing of amino acid biosynthesis between the aphid and Buchnera. The pea aphid genome provides a foundation for post-genomic studies of fundamental biological questions and applied agricultural problems

Molecular evolutionary trends and feeding ecology diversification in the Hemiptera, anchored by the milkweed bug genome.

BACKGROUND: The Hemiptera (aphids, cicadas, and true bugs) are a key insect order, with high diversity for feeding ecology and excellent experimental tractability for molecular genetics. Building upon recent sequencing of hemipteran pests such as phloem-feeding aphids and blood-feeding bed bugs, we present the genome sequence and comparative analyses centered on the milkweed bug Oncopeltus fasciatus, a seed feeder of the family Lygaeidae. RESULTS: The 926-Mb Oncopeltus genome is well represented by the current assembly and official gene set. We use our genomic and RNA-seq data not only to characterize the protein-coding gene repertoire and perform isoform-specific RNAi, but also to elucidate patterns of molecular evolution and physiology. We find ongoing, lineage-specific expansion and diversification of repressive C2H2 zinc finger proteins. The discovery of intron gain and turnover specific to the Hemiptera also prompted the evaluation of lineage and genome size as predictors of gene structure evolution. Furthermore, we identify enzymatic gains and losses that correlate with feeding biology, particularly for reductions associated with derived, fluid nutrition feeding. CONCLUSIONS: With the milkweed bug, we now have a critical mass of sequenced species for a hemimetabolous insect order and close outgroup to the Holometabola, substantially improving the diversity of insect genomics. We thereby define commonalities among the Hemiptera and delve into how hemipteran genomes reflect distinct feeding ecologies. Given Oncopeltus's strength as an experimental model, these new sequence resources bolster the foundation for molecular research and highlight technical considerations for the analysis of medium-sized invertebrate genomes