Habitat use of the ocelot (Leopardus pardalis) in Brazilian Amazon

Amazonia forest plays a major role in providing ecosystem services for human and sanctuaries for wildlife. However, ongoing deforestation and habitat fragmentation in the Brazilian Amazon has threatened both. The ocelot is an ecologically important mesopredator and a potential conservation ambassador species, yet there are no previous studies on its habitat preference and spatial patterns in this biome. From 2010 to 2017, twelve sites were surveyed, totaling 899 camera trap stations, the largest known dataset for this species. Using occupancy modeling incorporating spatial autocorrelation, we assessed habitat use for ocelot populations across the Brazilian Amazon. Our results revealed a positive sigmoidal correlation between remote‐sensing derived metrics of forest cover, disjunct core area density, elevation, distance to roads, distance to settlements and habitat use, and that habitat use by ocelots was negatively associated with slope and distance to river/lake. These findings shed light on the regional scale habitat use of ocelots and indicate important species–habitat relationships, thus providing valuable information for conservation management and land‐use planning.     

Escherichia coli “TatExpress” strains export several g/L human growth hormone to the periplasm by the Tat pathway

Escherichia coli is a heavily used platform for the production of biotherapeutic and other high-value proteins, and a favored strategy is to export the protein of interest to the periplasm to simplify downstream processing and facilitate disulfide bond formation. The Sec pathway is the standard means of transporting the target protein but it is unable to transport complex or rapidly folding proteins because the Sec system can only transport proteins in an unfolded state. The Tat system also operates to transport proteins to the periplasm, and it has significant potential as an alternative means of recombinant protein production because it transports fully folded proteins. Here, we have tested the Tat system's full potential for the production of biotherapeutics for the first time using fed-batch fermentation. We expressed human growth hormone (hGH) with a Tat signal peptide in E. coli W3110 “TatExpress” strains that contain elevated levels of the Tat apparatus. This construct contained four amino acids from TorA at the hGH N-terminus as well as the initiation methionine from hGH, which is removed in vivo. We show that the protein is efficiently exported to the periplasm during extended fed-batch fermentation, to the extent that it is by far the most abundant protein in the periplasm. The protein was shown to be homogeneous, disulfide bonded, and active. The bioassay showed that the yields of purified periplasmic hGH are 5.4 g/L culture whereas an enzyme-linked immunosorbent assay gave a figure of 2.39 g/L. Separate analysis of a TorA signal peptide linked to hGH construct lacking any additional amino acids likewise showed efficient export to the periplasm, although yields were approximately two-fold lower.     

CPEB3 deficiency in mice affect ovarian follicle development and causes premature ovarian insufficiency

Premature ovarian insufficiency (POI) is a heterogeneous and multifactorial disorder. In recent years, there has been an increasing interest in research on the pathogenesis and treatment of POI, owing to the implementation of the second-child policy in China. Cytoplasmic polyadenylation element-binding protein 3 (CPEB3) is an RNA-binding protein that can bind to specific RNA sequences. CPEB3 can bind to and affect the expression, cellular location, and stability of target RNAs. Cpeb3 is highly expressed in the ovary; however, its functions remain unknown. In this study, Cpeb3-mutant mice were used to characterize the physiological functions of CPEB3. Cpeb3-mutant female mice manifested signs of gradual loss of ovarian follicles, ovarian follicle development arrest, increased follicle atresia, and subfertility with a phenotype analogous to POI in women. Further analysis showed that granulosa cell proliferation was inhibited and apoptosis was markedly increased in Cpeb3-mutant ovaries. In addition, the expression of Gdf9, a potential target of CPEB3, was decreased in Cpeb3-mutant ovaries and oocytes. Altogether, these results reveal that CPEB3 is essential for ovarian follicle development and female fertility as it regulates the expression of Gdf9 in oocytes, disruption of which leads to impaired ovarian follicle development and POI.Subject terms: RNA-binding proteins, Infertility     

Anti-inflammatory effects of the products from Wilbrandia ebracteata on carrageenan-induced pleurisy in mice.

Wilbrandia ebracteata Cogn. (Cucurbitaceae) is commonly known in Brazil as "Taiuia". The roots are employed in folk medicine for the treatment of several diseases, such as rheumatic disease. This study has evaluated the anti-inflammatory action of dicloromethane fraction (F-DCM), purified fraction (PFIII) and Cucurbitacin B extracted from crude extract of W. ebracteata in experimental models in vivo. The F-DCM (0.3 to 10 mg.kg(-1), i.p. or 3 to 30 mg.kg(-1) p.o.) produced significant but not dose-dependent inhibition of the carrageenan-induced cell influx and exsudate leakage in the pleural cavity of mice. The F-DCM 0.01 to 10 mg.kg(-1), i.p. or 0.1 to 10 mg.kg(-1) p.o.) decreased the levels of PGE2 in the exsudate leakage induced by carrageenan in the pleural cavity after 4 h with a calculated ID50 of 0.01 (0.002-0.09, i.p.) and 0.29 (0.05-1.45, p.o.) mg.kg(-1). The PFIII (3 mg.kg(-1), i.p.) inhibited 80% of cell migration (1.50 +/- 0.09 x 10(6) cells/cavity) and exsudate leakage by about 50% (3.09 +/- 0.71 microg/ml) in relation to the control group. Cucurbitacin B (0.1 mg.kg(-1), i.p.), the main compound of PFIII, reduced significantly the levels of PGE2 in the exsudate leakage by 40.7% (10.41 +/- 2.67 ng.ml(-1)). These data show that the active principle(s) present in the F-DCM of W. ebracteata elicited pronounced anti-inflammatory effects when assessed by i.p. or p.o. routes, as well as PFIII. The F-DCM was also able to prevent PGE2 formation in exsudate leakage induced by carrageenan, as well as Cucurbitacin B, its active principle. These results indicate that the anti-inflammatory activity of Wilbrandia ebracteata can be related with the inhibition of the production of PGE2.     

Docking study of ligands into the colchicine binding site of tubulin

Cancer is a major cause of mortality in developed countries, following only cardiovascular diseases. Death of cancerous cells can be achieved by stopping mitosis and the antimitotic class of drugs formed by the spindle poisons can be used for this purpose. Their role is to disorganize the mitotic spindle by targeting its main constituent, the microtubules, themselves made of heterodimers of alpha and beta-tubulin. They disrupt the dynamics of the microtubules either by stabilizing them, as do paclitaxel or epothilones, or destabilizing them, as do colchicine. The binding site of colchicine seems to lie between the two units of the tubulin dimer. Here, we report on the characterization of this site by the docking of a series of reference compounds, and the subsequent docking of ligands prepared in our laboratory.     

Expression,purification and characterisation of full-length histidine protein kinase RegB from Rhodobacter sphaeroides

The global redox switch between aerobic and anaerobic growth in Rhodobacter sphaeroides is controlled by the RegA/RegB two-component system, in which RegB is the integral membrane histidine protein kinase, and RegA is the cytosolic response regulator. Despite the global regulatory importance of this system and its many homologues, there have been no reported examples to date of heterologous expression of full-length RegB or any histidine protein kinases. Here, we report the amplified expression of full-length functional His-tagged RegB in Escherichia coli, its purification, and characterisation of its properties. Both the membrane-bound and purified solubilised RegB protein demonstrate autophosphorylation activity, and the purified protein autophosphorylates at the same rate under both aerobic and anaerobic conditions confirming that an additional regulator is required to control/inhibit autophosphorylation. The intact protein has similar activity to previously characterised soluble forms, but is dephosphorylated more rapidly than the soluble form (half-life ca 30 minutes) demonstrating that the transmembrane segment present in the full-length RegB may be an important regulator of RegB activity. Phosphotransfer from RegB to RegA (overexpressed and purified from E. coli) by RegB is very rapid, as has been reported for the soluble domain. Dephosphorylation of active RegA by full-length RegB has a rate similar to that observed previously for soluble RegB.     

Multiple sclerosis: re-expression of a developmental pathway that restricts oligodendrocyte maturation

During mammalian central nervous system (CNS) development, contact-mediated activation of Notch1 receptors on oligodendrocyte precursors by the ligand Jagged1 induces Hes5, which inhibits maturation of these cells. Here we tested whether the Notch pathway is re-expressed in the adult CNS in multiple sclerosis (MS), an inflammatory demyelinating disease in which remyelination is typically limited. We found that transforming growth factor-beta 1 (TGF-beta 1), a cytokine upregulated in MS, specifically re-induced Jagged1 in primary cultures of human astrocytes. Within and around active MS plaques lacking remyelination, Jagged1 was expressed at high levels by hypertrophic astrocytes, whereas Notch1 and Hes5 localized to cells with an immature oligodendrocyte phenotype, and TGF-beta 1 was associated with perivascular extracellular matrix in the same areas. In contrast, there was negligible Jagged1 expression in remyelinated lesions. Experiments in vitro showed that Jagged1 signaling inhibited process outgrowth from primary human oligodendrocytes. These data are the first to implicate the Notch pathway in the limited remyelination in MS. Thus, Notch may represent a potential target for therapeutic intervention in this disease.