Differences in elemental composition of tailings, soils, and plant tissues following five decades of native plant colonization on a gold mine site in Northwestern Québec.

Mining activities have significant environmental impacts, such as the production of acid mine drainage and the typical absence of vegetation on mine tailings whose absence can facilitate the migration of metals to adjacent ecosystems. We investigated the metal and metalloid composition of plants and substrates on, and near a former gold mine site to understand elemental dynamics in such environments. A mine tailings deposit rich in Mo and As in Northwestern Québec was studied following the natural colonization of the deposit by boreal plant species. The site and surrounding forest were categorized into 6 vegetation density classes (VDC) to determine if and how vegetation density, and plant elemental composition, and soil properties were linked. Macroelemental composition of plant tissues (P, K and Ca) was relatively stable, despite differences in macroelemental levels of substrates between different VDC (with lower macronutrient levels associated with less dense areas), indicating the adaptability of the three species studied (Alnus incana spp. rugosa, Betula papyrifera and Picea spp.). Results showed that across a wide range of substrate properties, it was plant species and density that explained metal and metalloid composition in plant tissues (leaves, stems, and roots), while the main environmental determinants for this were VDC, pH, Ca and Cu. Increasing vegetation density was associated with decreasing As and Mo concentrations in substrates. This study sheds light on the plasticity of alder, spruce and birch growing on mine sites, allowing us to better understand elemental dynamics on such sites, and ultimately improve their management.

Uncovering the prevalence and diversity of integrating conjugative elements in actinobacteria.

Horizontal gene transfer greatly facilitates rapid genetic adaptation of bacteria to shifts in environmental conditions and colonization of new niches by allowing one-step acquisition of novel functions. Conjugation is a major mechanism of horizontal gene transfer mediated by conjugative plasmids and integrating conjugative elements (ICEs). While in most bacterial conjugative systems DNA translocation requires the assembly of a complex type IV secretion system (T4SS), in Actinobacteria a single DNA FtsK/SpoIIIE-like translocation protein is required. To date, the role and diversity of ICEs in Actinobacteria have received little attention. Putative ICEs were searched for in 275 genomes of Actinobacteria using HMM-profiles of proteins involved in ICE maintenance and transfer. These exhaustive analyses revealed 144 putative FtsK/SpoIIIE-type ICEs and 17 putative T4SS-type ICEs. Grouping of the ICEs based on the phylogenetic analyses of maintenance and transfer proteins revealed extensive exchanges between different sub-families of ICEs. 17 ICEs were found in Actinobacteria from the genus Frankia, globally important nitrogen-fixing microorganisms that establish root nodule symbioses with actinorhizal plants. Structural analysis of ICEs from Frankia revealed their unexpected diversity and a vast array of predicted adaptive functions. Frankia ICEs were found to excise by site-specific recombination from their host's chromosome in vitro and in planta suggesting that they are functional mobile elements whether Frankiae live as soil saprophytes or plant endosymbionts. Phylogenetic analyses of proteins involved in ICEs maintenance and transfer suggests that active exchange between ICEs cargo-borne and chromosomal genes took place within the Actinomycetales order. Functionality of Frankia ICEs in vitro as well as in planta lets us anticipate that conjugation and ICEs could allow the development of genetic manipulation tools for this challenging microorganism and for many other Actinobacteria.

Sequence analysis of rpoB and rpoD gene fragments reveals the phylogenetic diversity of actinobacteria of genus Frankia.

Partial rpoD, rpoB, and 16S rRNA gene sequences were obtained from databases and (or) amplified from 12 strains of Frankia. These strains belonged to either Cluster 1 (Alnus-, Myrica-, Comptonia-, and Casuarina-infective strains) or Cluster 3 (Elaeagnus-infective strain). An rpoD gene-based PCR approach was designed to allow the detection of frankiae in complex samples. Additionally, partial gene sequences obtained using 2 rpoB gene primer sets (named rpoB-1 and rpoB-2) were used to generate phylogenetic eurograms to find a molecular tool able to assess biodiversity among Frankia strains. The rpoB-2 primer set allowed separation of closely related strains and groupings representative of host plant compatibility groups. One exception to this was for strains ACN10a and ACN14a, isolated from the same geographical location. Results obtained showed that rpoB-2 is a tool of great interest to evaluate relatedness of Frankia strains, and assess biodiversity in this genus. Additionally, since rpoB-2 phylogenetic profiles of the Frankia strains studied reflected the species of host plants they were isolated from, the study of rpoB (a house-keeping gene) shows promise for future ecological studies on these symbioses.

MUC16 (CA125) regulates epithelial ovarian cancer cell growth, tumorigenesis and metastasis.

OBJECTIVES: MUC16 (CA125) protein is a high molecular weight mucin overexpressed in the majority of epithelial ovarian cancers (EOC) but not in the epithelium of normal ovaries suggesting that it might play a role in EOC pathogenesis. Here, we explored the phenotypic consequences of MUC16 knockdown and expression of its C-terminal domain with the aim of establishing a role for MUC16 in tumorigenesis. METHODS: MUC16 was down-regulated by stably expressing an anti-MUC16 endoplasmic reticulum-targeted single-chain antibody which prevented MUC16 cell surface localization in NIH:OVCAR3 cells. In addition, we generated epitope tagged, N-terminal region-deleted MUC16 constructs with (MUC16TMU) and without (MUC16CTD) cytoplasmic tail deletions and stably expressed them in SKOV3 cells. RESULTS: Although MUC16 knockdown did not affect the cell growth rate, knockdown cells reached a stationary growth phase after 4 days whereas control cells continued to grow for up to 7 days. Colony formation assays in soft agar demonstrated that MUC16 knockdown cells had >8-fold reduction in their ability to form colonies. Importantly, MUC16 knockdown completely prevents the formation of subcutaneous tumors in nude mice. Conversely, we show that ectopic expression of the MUC16CTD enhances SKOV3 tumor cell growth, colony formation in soft agar and enhances tumor growth and metastases in SCID mice. In addition, MUC16CTD expression increases cell motility, invasiveness, and metastatic property. Deletion of the cytoplasmic tail from the MUC16CTD completely abolished its ability to enhance tumor cell growth, cell motility and invasiveness. Furthermore, the increased invasive properties of MUC16CTD-expressing cells correlated with decreased expression of E-cadherin and increased expression of N-cadherin and vimentin. CONCLUSION: These findings provide the first evidence for a critical role of MUC16 in tumor cell growth, tumorigenesis and metastases.

High-throughput screening of microbial adaptation to environmental stress.

We developed a microwell plate, high-throughput, screening method aimed at quantitating the tolerance of a panel of Gram-positive and Gram-negative bacteria to metals (Frankia sp., Escherichia coli, Cupriavidus metallidurans, Rhizobium leguminosarum, and Streptomyces scabies). Microbial viability was quantified using MTS; a tetrazolium salt converted to a water-soluble formazan through microbial reduction. In this paper, we present the stepwise development of the method, highlighting the main elements underlying its reliability, and compare results obtained with literature. We conclude the method is well suited to efficiently screen bacteria, including those that are filamentous and slow-growing, without the need for large amounts of inoculum which may not always be available. The method allows testing of compound gradients with sufficient replicates to generate statistically robust results, and is transposable to other types of cell proliferation assays such as those for antimicrobial susceptibility, and chemoresistance.