Improved Isolation of SlaA and SlaB S-layer proteins in Sulfolobus acidocaldarius

The Sulfolobus acidocaldarius S-layer is composed of two main proteins: SlaA, which forms the ordered structure of the S-layer matrix, and SlaB, which supports and anchors the S-layer into the tetraether lipid membrane. While SlaA has previously been purified by exploiting its thermotolerance and high resistance to detergents, SlaB has resisted isolation, particularly from the cell membrane. Removal of proteins other than those of the S-layer is especially difficult if large batch-scale culture volumes are unavailable. Here, we describe a benchtop-scale protocol for the purification of SlaA from S. acidocaldarius, enabling isolation of SlaB using size exclusion chromatography (gel filtration). Using this protocol, we were able to identify for the first time tetraether lipids strongly attached to SlaB via heat- and detergent-resistant interactions.

     

SUV39H2 epigenetic silencing controls fate conversion of epidermal stem and progenitor cells

Epigenetic histone trimethylation on lysine 9 (H3K9me3) represents a major molecular signal for genome stability and gene silencing conserved from worms to man. However, the functional role of the H3K9 trimethylases SUV39H1/2 in mammalian tissue homeostasis remains largely unknown. Here, we use a spontaneous dog model with monogenic inheritance of a recessive SUV39H2 loss-of-function variant and impaired differentiation in the epidermis, a self-renewing tissue fueled by stem and progenitor cell proliferation and differentiation. Our results demonstrate that SUV39H2 maintains the stem and progenitor cell pool by restricting fate conversion through H3K9me3 repressive marks on gene promoters encoding components of the Wnt/p63/adhesion axis. When SUV39H2 function is lost, repression is relieved, and enhanced Wnt activity causes progenitor cells to prematurely exit the cell cycle, a process mimicked by pharmacological Wnt activation in primary canine, human, and mouse keratinocytes. As a consequence, the stem cell growth potential of cultured SUV39H2-deficient canine keratinocytes is exhausted while epidermal differentiation and genome stability are compromised. Collectively, our data identify SUV39H2 and potentially also SUV39H1 as major gatekeepers in the delicate balance of progenitor fate conversion through H3K9me3 rate-limiting road blocks in basal layer keratinocytes.     

Multiple stages of plant root calcification deciphered by chemical and micromorphological analyses

Rhizoliths, that is, roots fossilized by secondary carbonates, have been known for ages and are increasingly used for paleoenvironmental reconstructions. However, knowledge about their formation mechanisms remains limited. This study reports the mineralogical and chemical characterization of rhizoliths at different stages of mineralization and fossilization in the Late Pleistocene loess–paleosol sequence of Nussloch (SW Germany). Scanning electron microscopy coupled with elemental mapping and ¹³C solid‐state nuclear magnetic resonance were used to concomitantly characterize the mineral and organic matter of the rhizoliths. These joint analyses showed for the first time that large rhizoliths are not necessarily remains of single large roots but consist of numerous microrhizoliths as remains of fine roots, formed mainly by calcium carbonates with only low amounts of Mg and Si. They further revealed that the precipitation of secondary carbonates occurs not only around, but also within the plant root and that fossilization leads to the selective preservation of recalcitrant root biopolymers—lignin and suberin. The precipitation of secondary carbonates was observed to occur first around fine roots, the epidermis acting as a first barrier, and then within the root, within the cortex cells, and even sometimes around the phloem and within the xylem. This study suggests that the calcification of plant roots starts during the lifetime of the plant and continues after its death. This has to be systematically investigated to understand the stratigraphic context before using (micro)rhizoliths for paleoenvironmental reconstructions in terrestrial sediments.     

Proteomic Analysis of the Reproductive Organs of the Hermaphroditic Gastropod Lymnaea stagnalis Exposed to Different Endocrine Disrupting Chemicals

Many studies have reported perturbations of mollusc reproduction following exposure to low concentrations (ng/L range) of endocrine disrupting chemicals (EDCs). However, the mechanisms of action of these molecules on molluscs are still poorly understood. Investigation of the modifications of protein expression in organisms exposed to chemicals using proteomic methods can provide a broader and more comprehensive understanding of adverse impacts of pollution on organisms than conventional biochemical biomarkers (e.g., heat-shock proteins, metallothioneins, GST, EROD). In this study we have investigated the impacts of four chemicals, which exhibit different endocrine disrupting properties in vertebrates, on the proteome of the hermaphroditic freshwater pulmonate gastropod Lymnaea stagnalis after 21 days of exposure. Testosterone, tributyltin, chlordecone and cyproterone acetate were chosen as tested compounds as they can induce adverse effects on the reproduction of this snail. The 2D-DIGE method was used to identify proteins whose expression was affected by these compounds. In addition to modifying the expression of proteins involved in the structure and function of the cytoskeleton, chemicals had impacts on the expression of proteins involved in the reproduction of L. stagnalis. Exposure to 19.2 µg/L of chlordecone increased the abundance of ovipostatin, a peptide transmitted during mating through seminal fluid, which reduces oviposition in this species. The expression of yolk ferritin, the vitellogenin equivalent in L. stagnalis, was reduced after exposure to 94.2 ng Sn/L of tributyltin. The identification of yolk ferritin and the modification of its expression in snails exposed to chemicals were refined using western blot analysis. Our results showed that the tested compounds influenced the abundance of yolk ferritin in the reproductive organs. Alteration in proteins involved in reproductive pathways (e.g., ovipostatin and yolk ferritin) could constitute relevant evidence of interaction of EDCs with reproductive pathways that are under the control of the endocrine system of L. stagnalis.     

An Alteration in ELMOD3, an Arl2 GTPase-Activating Protein,Is Associated with Hearing Impairment in Humans

Exome sequencing coupled with homozygosity mapping was used to identify a transition mutation (c.794T>C; p.Leu265Ser) in ELMOD3 at the DFNB88 locus that is associated with nonsyndromic deafness in a large Pakistani family, PKDF468. The affected individuals of this family exhibited pre-lingual, severe-to-profound degrees of mixed hearing loss. ELMOD3 belongs to the engulfment and cell motility (ELMO) family, which consists of six paralogs in mammals. Several members of the ELMO family have been shown to regulate a subset of GTPases within the Ras superfamily. However, ELMOD3 is a largely uncharacterized protein that has no previously known biochemical activities. We found that in rodents, within the sensory epithelia of the inner ear, ELMOD3 appears most pronounced in the stereocilia of cochlear hair cells. Fluorescently tagged ELMOD3 co-localized with the actin cytoskeleton in MDCK cells and actin-based microvilli of LLC-PK1-CL4 epithelial cells. The p.Leu265Ser mutation in the ELMO domain impaired each of these activities. Super-resolution imaging revealed instances of close association of ELMOD3 with actin at the plasma membrane of MDCK cells. Furthermore, recombinant human GST-ELMOD3 exhibited GTPase activating protein (GAP) activity against the Arl2 GTPase, which was completely abolished by the p.Leu265Ser mutation. Collectively, our data provide the first insights into the expression and biochemical properties of ELMOD3 and highlight its functional links to sound perception and actin cytoskeleton.     

Side Effects of Being Blue: Influence of Sad Mood on Visual Statistical Learning

It is well established that mood influences many cognitive processes, such as learning and executive functions. Although statistical learning is assumed to be part of our daily life, as mood does, the influence of mood on statistical learning has never been investigated before. In the present study, a sad vs. neutral mood was induced to the participants through the listening of stories while they were exposed to a stream of visual shapes made up of the repeated presentation of four triplets, namely sequences of three shapes presented in a fixed order. Given that the inter-stimulus interval was held constant within and between triplets, the only cues available for triplet segmentation were the transitional probabilities between shapes. Direct and indirect measures of learning taken either immediately or 20 minutes after the exposure/mood induction phase revealed that participants learned the statistical regularities between shapes. Interestingly, although participants from the sad and neutral groups performed similarly in these tasks, subjective measures (confidence judgments taken after each trial) revealed that participants who experienced the sad mood induction showed increased conscious access to their statistical knowledge. These effects were not modulated by the time delay between the exposure/mood induction and the test phases. These results are discussed within the scope of the robustness principle and the influence of negative affects on processing style.     

Integrative Biology of Idas iwaotakii (Habe, 1958), a ‘Model Species’ Associated with Sunken Organic Substrates

The giant bathymodioline mussels from vents have been studied as models to understand the adaptation of organisms to deep-sea chemosynthetic environments. These mussels are closely related to minute mussels associated to organic remains decaying on the deep-sea floor. Whereas biological data accumulate for the giant mussels, the small mussels remain poorly studied. Despite this lack of data for species living on organic remains it has been hypothesized that during evolution, contrary to their relatives from vents or seeps, they did not acquire highly specialized biological features. We aim at testing this hypothesis by providing new biological data for species associated with organic falls. Within Bathymodiolinae a close phylogenetic relationship was revealed between the Bathymodiolus sensu stricto lineage (i.e. “thermophilus” lineage) which includes exclusively vent and seep species, and a diversified lineage of small mussels, attributed to the genus Idas, that includes mostly species from organic falls. We selected Idas iwaotakii (Habe, 1958) from this latter lineage to analyse population structure and to document biological features. Mitochondrial and nuclear markers reveal a north-south genetic structure at an oceanic scale in the Western Pacific but no structure was revealed at a regional scale or as correlated with the kind of substrate or depth. The morphology of larval shells suggests substantial dispersal abilities. Nutritional features were assessed by examining bacterial diversity coupled by a microscopic analysis of the digestive tract. Molecular data demonstrated the presence of sulphur-oxidizing bacteria resembling those identified in other Bathymodiolinae. In contrast with most Bathymodiolus s.s. species the digestive tract of I. iwaotakii is not reduced. Combining data from literature with the present data shows that most of the important biological features are shared between Bathymodiolus s.s. species and its sister-lineage. However Bathymodiolus s.s. species are ecologically more restricted and also display a lower species richness than Idas species.