Comparative analysis of novel and common reference genes in adult tissues of the mussel Mytilus galloprovincialis

Background Real-time quantitative PCR is a widely used method for gene expression analyses in various organisms. Its accuracy mainly relies on the correct selection of reference genes. Any experimental plan involving real-time PCR needs to evaluate the characteristics of the samples to be examined and the relative stability of reference genes. Most studies in mollusks rely on reference genes commonly used in vertebrates. Results In this study, we focused on the transcriptome of the bivalve mollusk Mytilus galloprovincialis in physiological state to identify suitable reference genes in several adult tissues. Candidate genes with highly stable expression across 51 RNA-seq datasets from multiple tissues were selected through genome-wide bioinformatics analysis. This approach led to the identification of three genes (Rpl14, Rpl32 and Rpl34), whose suitability was evaluated together with 7 other reference genes commonly reported in literature (Act, Cyp-A, Ef1 alpha, Gapdh, 18S, 28S and Rps4). The stability analyses performed with geNorm, NormFinder and Bestkeeper identified specific either single or pairs of genes suitable as references for gene expression analyses in specific tissues and revealed the Act/Cyp-A pair as the most appropriate to analyze gene expression across different tissues. Conclusion Mytilus galloprovincialis is a model system increasingly used in ecotoxicology and molecular studies. Our transcriptome-wide approach represents the first comprehensive investigation aimed at the identification of suitable reference genes for expression studies in this species

Selection and validation of a set of reliable reference genes for quantitative RT-PCR studies in the brain of the Cephalopod Mollusc Octopus vulgaris

<p>Abstract</p> <p>Background</p> <p>Quantitative real-time polymerase chain reaction (RT-qPCR) is valuable for studying the molecular events underlying physiological and behavioral phenomena. Normalization of real-time PCR data is critical for a reliable mRNA quantification. Here we identify reference genes to be utilized in RT-qPCR experiments to normalize and monitor the expression of target genes in the brain of the cephalopod mollusc <it>Octopus vulgaris</it>, an invertebrate. Such an approach is novel for this taxon and of advantage in future experiments given the complexity of the behavioral repertoire of this species when compared with its relatively simple neural organization.</p> <p>Results</p> <p>We chose <it>16S</it>, and <it>18S </it>rRNA, <it>actB</it>, <it>EEF1A</it>, <it>tubA </it>and <it>ubi </it>as candidate reference genes (housekeeping genes, HKG). The expression of <it>16S </it>and <it>18S </it>was highly variable and did not meet the requirements of candidate HKG. The expression of the other genes was almost stable and uniform among samples. We analyzed the expression of HKG into two different set of animals using tissues taken from the central nervous system (brain parts) and mantle (here considered as control tissue) by BestKeeper, geNorm and NormFinder. We found that HKG expressions differed considerably with respect to brain area and octopus samples in an HKG-specific manner. However, when the mantle is treated as control tissue and the entire central nervous system is considered, NormFinder revealed <it>tubA </it>and <it>ubi </it>as the most suitable HKG pair. These two genes were utilized to evaluate the relative expression of the genes <it>FoxP</it>, <it>creb, dat </it>and <it>TH </it>in <it>O. vulgaris</it>.</p> <p>Conclusion</p> <p>We analyzed the expression profiles of some genes here identified for <it>O. vulgaris </it>by applying RT-qPCR analysis for the first time in cephalopods. We validated candidate reference genes and found the expression of <it>ubi </it>and <it>tubA </it>to be the most appropriate to evaluate the expression of target genes in the brain of different octopuses. Our results also underline the importance of choosing a proper normalization strategy when analyzing gene expression by qPCR taking into appropriate account the experimental setting and variability of the sample of animals (and tissues), thus providing a set of HGK which expression appears to be unaffected by the experimental factor(s).</p

Cyanidiophyceae (Rhodophyta) Tolerance to Precious Metals: Metabolic Response to Palladium and Gold

Polyextremophilic red algae, which belong to the class Cyanidiophyceae, are adapted to live in geothermal and volcanic sites. These sites often have very high concentrations of heavy and precious metals. In this study, we assessed the capacity of three strains of Galdieria (G. maxima, G. sulphuraria, and G. phlegrea) and one strain of Cyanidium caldarium to tolerate different concentrations of precious metals, such as palladium (Cl4K2Pd) and gold (AuCl4K) by monitoring algal growths in cultures exposed to metals, and we investigated the algae potential oxidative stress induced by the metals. This work provides further understanding of metals responses in the Cyanidiophyceae, as this taxonomic class is developed as a biological refinement tool

Cyanidiophyceae (Rhodophyta) Tolerance to Precious Metals: Metabolic Response to Palladium and Gold

Polyextremophilic red algae, which belong to the class Cyanidiophyceae, are adapted to live in geothermal and volcanic sites. These sites often have very high concentrations of heavy and precious metals. In this study, we assessed the capacity of three strains of Galdieria (G. maxima, G. sulphuraria, and G. phlegrea) and one strain of Cyanidium caldarium to tolerate different concentrations of precious metals, such as palladium (Cl4K2Pd) and gold (AuCl4K) by monitoring algal growths in cultures exposed to metals, and we investigated the algae potential oxidative stress induced by the metals. This work provides further understanding of metals responses in the Cyanidiophyceae, as this taxonomic class is developed as a biological refinement tool

Cyanidium chilense (Cyanidiophyceae, Rhodophyta) from tuff rocks of the archeological site of Cuma, Italy

Phlegrean Fields is a large volcanic area situated southwest of Naples (Italy), including both cave and thermoacidic habitats. These extreme environments host the genus Cyanidium; the species C. chilense represents a common phototrophic micro- organism living in anthropogenic caves. With a view to provide a comprehensive characterization for a correct taxonomic classification, morpho-ultrastructural investigations ofC. chilense from Syb’s cave (Phlegren Fields) was herein car-ried out and compared with the thermoacidophilic C. caldarium. The biofilm was also analyzed to define the roleofC. chilensein the establishment of a bio film within caveenvironments. Despite the peculiar ecological and molecular divergences,C. chilenseandC. caldariumshared all the maindiacritic features, suggesting morphological convergence within the genus; cytological identity was found amongC. chilensestrains geographically distant and adapted to different substrates, such as the porous yellow tuff of Sybil cave and calcyte, magnesite and basaltic rocks from other caves. C. chilense is generally dominant in all biofilms, developing monospecific islets, developing both super ficially or betweenfungal hyphae and coccoid cyanobacteria. Extracellular polymeric substances (EPS) were recorded in C. chilense bio filmsfrom Sybil cave, confirming the role of EPS in facilitating cellsadhesion to the surface, creating a cohesive network of inter-connecting biofilm cell

Tolerance and metabolic responses of Cyanidiophytina (Rhodophyta) towards exposition to Cl4K2Pd and AuCl4K

Polyextremophilic algae, such as unicellular red algae known as Cyanidiophyceae, have the intrinsic capacity to selectively mobilize and adsorb metals, since they are adapted to live in geothermal and volcanic sites characterized by elevated concentration of heavy and rare metals. In this work we evaluated the ability of 3 strains of the genus Galdieria (G. maxima, G. sulphuraria, G. phlegrea) along with one strain of Cyanidium caldarium to tolerate different concentrations of rare metal as Cl4K2Pd and AuCl4K by monitoring changes in algal growth in culture exposed to different concentration of each metal and investigating algae metabolic response and possible oxidative stress induced by these metals

Cyanidiophyceae (Rhodophyta) tolerance to precious metals: metabolic response to Cl4K2Pd and AuCl4K

Polyextremophilic red algae, which belong to the class Cyanidiophyceae, are adapted to live in geothermal and volcanic sites. These sites often have very high concentrations of heavy and precious metals. In this study, we assessed the capacity of three strains of Galdieria (G. maxima, G. sulphuraria, and G. phlegrea) and one strain of Cyanidiumcaldarium to tolerate different concentrations of precious metals, such as palladium (Cl4K2Pd) and gold (AuCl4K) by monitoring algal growths in cultures exposed to metals, and we investigated the algae potential oxidative stress induced by the metals. This work provides further understanding of metals responses in the Cyanidiophyceae, as this taxonomic class is developed as a biological refinement tool

Multi-Level Interactions between the Nuclear Receptor TRα1 and the WNT Effectors β-Catenin/Tcf4 in the Intestinal Epithelium

Intestinal homeostasis results from complex cross-regulation of signaling pathways; their alteration induces intestinal tumorigenesis. Previously, we found that the thyroid hormone nuclear receptor TRα1 activates and synergizes with the WNT pathway, inducing crypt cell proliferation and promoting tumorigenesis. Here, we investigated the mechanisms and implications of the cross-regulation between these two pathways in gut tumorigenesis in vivo and in vitro. We analyzed TRα1 and WNT target gene expression in healthy mucosae and tumors from mice overexpressing TRα1 in the intestinal epithelium in a WNT-activated genetic background (vil-TRα1/Apc mice). Interestingly, increased levels of β-catenin/Tcf4 complex in tumors from vil-TRα1/Apc mice blocked TRα1 transcriptional activity. This observation was confirmed in Caco2 cells, in which TRα1 functionality on a luciferase reporter-assay was reduced by the overexpression of β-catenin/Tcf4. Moreover, TRα1 physically interacted with β-catenin/Tcf4 in the nuclei of these cells. Using molecular approaches, we demonstrated that the binding of TRα1 to its DNA target sequences within the tumors was impaired, while it was newly recruited to WNT target genes. In conclusion, our observations strongly suggest that increased β-catenin/Tcf4 levels i) correlated with reduced TRα1 transcriptional activity on its target genes and, ii) were likely responsible for the shift of TRα1 binding on WNT targets. Together, these data suggest a novel mechanism for the tumor-promoting activity of the TRα1 nuclear receptor

Age and date for early arrival of the Acheulian in Europe (Barranc de la Boella, la Canonja, Spain)

The first arrivals of hominin populations into Eurasia during the Early Pleistocene are currently considered to have occurred as short and poorly dated biological dispersions. Questions as to the tempo and mode of these early prehistoric settlements have given rise to debates concerning the taxonomic significance of the lithic assemblages, as trace fossils, and the geographical distribution of the technological traditions found in the Lower Palaeolithic record. Here, we report on the Barranc de la Boella site which has yielded a lithic assemblage dating to ,1 million years ago that includes large cutting tools (LCT). We argue that distinct technological traditions coexisted in the Iberian archaeological repertoires of the late Early Pleistocene age in a similar way to the earliest sub-Saharan African artefact assemblages. These differences between stone tool assemblages may be attributed to the different chronologies of hominin dispersal events. The archaeological record of Barranc de la Boella completes the geographical distribution of LCT assemblages across southern Eurasia during the EMPT (Early-Middle Pleistocene Transition, circa 942 to 641 kyr). Up to now, chronology of the earliest European LCT assemblages is based on the abundant Palaeolithic record found in terrace river sequences which have been dated to the end of the EMPT and later. However, the findings at Barranc de la Boella suggest that early LCT lithic assemblages appeared in the SW of Europe during earlier hominin dispersal episodes before the definitive colonization of temperate Eurasia took place.The research at Barranc de la Boella has been carried out with the financial support of the Spanish Ministerio de Economı´a y Competitividad (CGL2012- 36682; CGL2012-38358, CGL2012-38434-C03-03 and CGL2010-15326; MICINN project HAR2009-7223/HIST), Generalitat de Catalunya, AGAUR agence (projects 2014SGR-901; 2014SGR-899; 2009SGR-324, 2009PBR-0033 and 2009SGR-188) and Junta de Castilla y Leo´n BU1004A09. Financial support for Barranc de la Boella field work and archaeological excavations is provided by the Ajuntament de la Canonja and Departament de Cultura (Servei d’Arqueologia i Paleontologia) de la Generalitat de Catalunya. A. Carrancho’s research was funded by the International Excellence Programme, Reinforcement subprogramme of the Spanish Ministry of Education. I. Lozano-Ferna´ndez acknowledges the pre-doctoral grant from the Fundacio´n Atapuerca. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Thyroid Hormone Signaling and Function: News from Classical and Emerging Models

According to Brown and Cai, Thyroid hormones (THs) have been considered &ldquo;the first developmental morphogen ever discovered&rdquo; [...