A role for PKC-ɛ in FcγR-mediated phagocytosis by RAW 264.7 cells

Protein kinase C (PKC) plays a prominent role in immune signaling, and the paradigms for isoform selective signaling are beginning to be elucidated. Real-time microscopy was combined with molecular and biochemical approaches to demonstrate a role for PKC-ɛ in Fcγ receptor (FcγR)–dependent phagocytosis. RAW 264.7 macrophages were transfected with GFP-conjugated PKC isoforms, and GFP movement was followed during phagocytosis of fluorescent IgG–opsonized beads. PKC-ɛ, but not PKC-δ, concentrated around the beads. PKC-ɛ accumulation was transient; apparent as a “flash” on target ingestion. Similarly, endogenous PKC-ɛ was specifically recruited to the nascent phagosomes in a time-dependent manner. Overexpression of PKC-ɛ, but not PKC-α, PKC-δ, or PKC-γ enhanced bead uptake 1.8-fold. Additionally, the rate of phagocytosis in GFP PKC-ɛ expressors was twice that of cells expressing GFP PKC-δ. Expression of the regulatory domain (ɛRD) and the first variable region (ɛV1) of PKC-ɛ inhibited uptake, whereas the corresponding PKC-δ region had no effect. Actin polymerization was enhanced on expression of GFP PKC-ɛ and ɛRD, but decreased in cells expressing ɛV1, suggesting that the ɛRD and ɛV1 inhibition of phagocytosis is not due to effects on actin polymerization. These results demonstrate a role for PKC-ɛ in FcγR-mediated phagocytosis that is independent of its effects on actin assembly

Combined Operational Taxonomic Unit (OTU) Table

This operational taxonomic unit (OTU) table is the same as the final OTU (fOTU) table, EXCEPT the technical replicates have been combined into one sample. Therefore there are no A and B samples anymore and they are combined and treated as one sample

Data from: Metabarcoding reveals environmental factors influencing spatio-temporal variation in pelagic micro-eukaryotes

Marine environments harbour a vast diversity of micro-eukaryotic organisms (protists and other small eukaryotes) that play important roles in structuring marine ecosystems. However, micro-eukaryote diversity is not well understood. Likewise, knowledge is limited regarding micro-eukaryote spatial and seasonal distribution, especially over long temporal scales. Given the importance of this group for mobilizing energy from lower trophic levels near the base of the food chain to larger organisms, assessing community stability, diversity and resilience is important to understand ecosystem health. Herein, we use a metabarcoding approach to examine pelagic micro-eukaryote communities over a 2.5-year time series. Bimonthly surface sampling (July 2009 to December 2011) was conducted at four locations within Mobile Bay (Bay) and along the Alabama continental shelf (Shelf). Alpha-diversity only showed significant differences in Shelf sites, with the greatest differences observed between summer and winter. Beta-diversity showed significant differences in community composition in relation to season and the Bay was dominated by diatoms, while the Shelf was characterized by dinoflagellates and copepods. The northern Gulf of Mexico is heavily influenced by the Mobile River Basin, which brings low-salinity nutrient-rich water mostly during winter and spring. Community composition was correlated with salinity, temperature and dissolved silicate. However, species interactions (e.g. predation and parasitism) may also contribute to the observed variation, especially on the Shelf, which warrants further exploration. Metabarcoding revealed clear patterns in surface pelagic micro-eukaryote communities that were consistent over multiple years, demonstrating how these techniques could be greatly beneficial to ecological monitoring and management over temporal scales

The genus Grateloupia C. Agardh (Halymeniaceae, Rhodophyta) in the Thau Lagoon (France, Mediterranean): A case study of marine plurispecific introductions

Based on morphological data and molecular analyses [Nuclear ribosomal internal transcribed spacer (ITS), rbcL and mitochondrial cox2-cox3 spacer sequences] of Grateloupia spp. populations in the Thau Lagoon (France, Mediterranean) we demonstrated that at least five exotic species of Grateloupia were introduced. These include: (1) Grateloupia asiatica, a recently described species that was previously misidentified as G. filicina in Japan and Grateloupia sp. in the Thau Lagoon; (2) G. lanceolata from Japan; (3) G. luxurians, a Pacific species described as G. filicina var. luxurians; (4) G. patens from Japan; and (5) G. turuturu, a Japanese species previously misidentified as G. doryphora in the NE and NW Atlantic and Mediterranean Sea. These nonnative species probably were introduced in the Thau Lagoon in the 1970s along with the massive importations of Japanese oysters, Crassostrea gigas, into Europe for mariculture purposes. Since their introduction, they all have established large, reproductive populations with the exception of G. patens. The Mediterranean Grateloupia specimens are genetically and morphologically similar to Pacific specimens of the same species, although in the Thau Lagoon, G. asiatica specimens are morphologically more variable than those found in Japanese populations. This is the first report of G. asiatica in the Mediterranean Sea and Europe. Based on morphological data and molecular analyses (rbcL sequences) G. subpectinata is placed in synonymy with G. luxurians

Original Operational Taxonomic Unit (OTU) Table

This is the original operational taxonomic unit (OTU) table resulting from the sequences clustering prior to any filtering of OTUs

Geographic variation in allele frequency of the gamete recognition protein M7 lysin throughout a mosaic blue mussel hybrid zone

Divergence at gamete recognition loci is hypothesized to result in speciation in broadcast spawning invertebrates. Many gamete recognition loci evolve quickly and show patterns of positive selection, yet the advantage of divergence is rarely known. M7 lysin is a sperm protein in the Mytilus edulis species complex that shows evidence of adaptive evolution. This locus is polymorphic with two distinct clades within Mytilus galloprovincialis, one of which, the D clade, shows the strongest signal of positive selection. We tested whether the geographic patterns in allele frequency were consistent with the hypothesis that positive selection on D clade alleles (GD) was due to reinforcement. Populations of M. edulis showed little evidence of introgression of GD alleles, but there was no consistent evidence of reproductive character displacement. The lack of consistent patterns expected of reinforcement suggests that another mechanism is likely responsible for the adaptive divergence of M7 lysin. © 2013 Springer-Verlag Berlin Heidelberg

Representative Operational Taxonomic Unit (OTU) Sequences

The sequences used as representatives for the different operational taxonomic unit (OTU) clusters. These sequences are the most abundant sequences found within the OTU cluster. These sequences were used for taxonomic assignment and sequence alignment as stated in the manuscript. Further detail can be found in the method section of the manuscript

Final Operational Taxonomic Unit (fOTU) Table

The filtered operational taxonomic unit (OTU) table that excludes OTUs that failed to align with PyNAST or were classified as Archaea and Bacteria. This OTU table corresponds to the final OTU (fOTU) table listed in the manuscript

Biogeochemical and microbial variation across 5500 km of Antarctic surface sediment implicates organic matter as a driver of benthic community structure

Western Antarctica, one of the fastest warming locations on Earth, is a unique environment that is underexplored with regards to biodiversity. Although pelagic microbial communities in the Southern Ocean and coastal Antarctic waters have been well studied, there are fewer investigations of benthic communities and most have a focused geographic range. We sampled surface sediment from 24 sites across a 5,500 km region of Western Antarctica (covering the Ross Sea to the Weddell Sea) to examine relationships between microbial communities and sediment geochemistry. Sequencing of the 16S and 18S rRNA genes showed microbial communities in sediments from the Antarctic Peninsula (AP) and Western Antarctica (WA), including the Ross, Amundsen, and Bellingshausen Seas, could be distinguished by correlations with organic matter concentrations and stable isotope fractionation (total organic carbon; TOC, nitrogen, and δ13C). Overall, samples from the AP were higher in nutrient content (TOC, nitrogen, and NH4+) and communities in these samples had higher relative abundances of operational taxonomic units (OTUs) classified as the diatom, Chaetoceros, a marine cercozoan and four OTUs classified as Cytophaga or Flavobacteria. As these OTUs were strongly correlated with TOC, the data suggests the diatoms could be a source of organic matter and the Bacteroidetes and cercozoan are grazers that consume the organic matter. Additionally, samples from WA have lower nutrients and were dominated by Thaumarchaeota, which could be related to their known ability to thrive as lithotrophs. This study documents the largest analysis of benthic microbial communities to date in the Southern Ocean, representing almost half the continental shoreline of Antarctica, and documents trophic interactions and coupling of pelagic and benthic communities. Our results indicate potential modifications in carbon sequestration processes related to change in community composition, identifying a prospective mechanism that links climate change to carbon availability

Reconstruction of Cyclooxygenase Evolution in Animals Suggests Variable, Lineage-Specific Duplications, and Homologs with Low Sequence Identity

Cyclooxygenase (COX) enzymatically converts arachidonic acid into prostaglandin G/H in animals and has importance during pregnancy, digestion, and other physiological functions in mammals. COX genes have mainly been described from vertebrates, where gene duplications are common, but few studies have examined COX in invertebrates. Given the increasing ease in generating genomic data, as well as recent, although incomplete descriptions of potential COX sequences in Mollusca, Crustacea, and Insecta, assessing COX evolution across Metazoa is now possible. Here, we recover 40 putative COX orthologs by searching publicly available genomic resources as well as ~250 novel invertebrate transcriptomic datasets. Results suggest the common ancestor of Cnidaria and Bilateria possessed a COX homolog similar to those of vertebrates, although such homologs were not found in poriferan and ctenophore genomes. COX was found in most crustaceans and the majority of molluscs examined, but only specific taxa/lineages within Cnidaria and Annelida. For example, all octocorallians appear to have COX, while no COX homologs were found in hexacorallian datasets. Most species examined had a single homolog, although species-specific COX duplications were found in members of Annelida, Mollusca, and Cnidaria. Additionally, COX genes were not found in Hemichordata, Echinodermata, or Platyhelminthes, and the few previously described COX genes in Insecta lacked appreciable sequence homology (although structural analyses suggest these may still be functional COX enzymes). This analysis provides a benchmark for identifying COX homologs in future genomic and transcriptomic datasets, and identifies lineages for future studies of COX