Biochemical,biophysical and molecular dynamics studies on the proteoglycan-like domain of carbonic anhydrase IX

Human carbonic anhydrase IX (hCA IX) is a tumour-associated enzyme present in a limited number of normal tissues, but overexpressed in several malignant human tumours. It is a transmembrane protein, where the extracellular region consists of a greatly investigated catalytic CA domain and a much less investigated proteoglycan-like (PG) domain. Considering its important role in tumour biology, here, we report for the first time the full characterization of the PG domain, providing insights into its structural and functional features. In particular, this domain has been produced at high yields in bacterial cells and characterized by means of biochemical, biophysical and molecular dynamics studies. Results show that it belongs to the family of intrinsically disordered proteins, being globally unfolded with only some local residual polyproline II secondary structure. The observed conformational flexibility may have several important roles in tumour progression, facilitating interactions of hCA IX with partner proteins assisting tumour spreading and progression.     

Reproductive biology of two species of Hyalella Smith, 1874 (Crustacea: Amphipoda: Hyalellidae) from southern Brazil

Reproduction is a critical event in the life cycle of animals, and a high amount of energy is allocated to it. The aim of this study was to characterize the reproductive biology of two recently described species of Hyalella from the north-east region of state of Rio Grande do Sul in southern Brazil. The specimens were collected during 12 months from a stream source (Hyalella georginae) and from a water reservoir (H. gauchensis), in Palmeira das Missões municipality, state of Rio Grande do Sul. Upon sampling, ovigerous females and pairs in precopulatory behaviour were separated into microtubes. In the laboratory, the cephalothorax length (CL in mm) of all specimens was measured. Male size at onset of sexual maturity was larger than that of the females, in both species. There was a correlation between the size of paired males and females: larger males carried larger females, in both Hyalella species. Precopulatory pairs were found throughout the year; however, H. georginae had a higher frequency in spring, and H. gauchensis in autumn. There was a positive correlation between the CL of ovigerous females and the number of eggs/juveniles. A reduction in the number of eggs throughout the embryonic development was observed in H. georginae. The average fecundity of H. georginae was significantly higher than H. gauchensis. The differences between the reproductive biology of these species may be related to the physical and environmental conditions at which they are exposed in their habitats.     

GABA transporter lysine 448: a key residue for tricyclic antidepressants interaction

The effects of three tricyclic antidepressants (TCAs) and two serotonin selective reuptake inhibitors (SSRIs) have been studied with an electrophysiological approach on Xenopus laevis oocytes expressing the rat GABA (γ-Aminobutyric-acid) transporter rGAT1. All tested TCAs and SSRIs inhibit the GABA-associated current in a dose-dependent way with low but comparable efficacy. The pre-steady-state and uncoupled currents appear substantially unaffected. The efficacy of desipramine, but not of the other drugs, is strongly increased in the lysine-glutamate or -aspartate mutants K448E and K448D. Comparison of I _max and K _0.5GABA in the absence and presence of desipramine showed that both parameters are reduced by the drug in the wild-type and in the K448E mutant. This suggests an uncompetitive inhibition, in which the drug can bind only after the substrate, an explanation in agreement with the lack of effects on the pre-steady-state and leak currents, and with the known structural data.     

Anandamide extends platelets survival through CB1-dependent Akt signaling

Platelets are stored at 22°C, since incubation at 37°C results in loss of viability. Nonetheless, in our body (37°C), platelets survive for 8–10 days. This discrepancy has been explained in terms of deprivation of viability factors or accumulation of apoptotic factors during storage. We report that the endocannabinoid anandamide (AEA) may be one of the agents allowing platelet survival. In fact, at 37°C, human platelets enhance the expression of pro-apoptotic proteins (caspases, Bax, Bak) and decrease the expression of Bcl-xL, thus changing the Bcl-xL/Bak ratio, a key platelet biological clock. AEA or its non-hydrolyzable analogue, methanandamide, extend platelet life span, without reversing the changes in Bcl-xL/Bak ratio induced by heat stress. Instead, AEA binding to type-1 cannabinoid receptor activates Akt, which regulates, through phosphorylation of Bad, the interactions among different Bcl-2 family members. These findings could have implications for platelet collection and, potentially, for their clinical use.     

Engineering evolution to study speciation in yeasts

The Saccharomyces 'sensu stricto' yeasts are a group of species that will mate with one another, but interspecific pairings produce sterile hybrids. A retrospective analysis of their genomes revealed that translocations between the chromosomes of these species do not correlate with the group's sequence-based phylogeny (that is, translocations do not drive the process of speciation). However, that analysis was unable to infer what contribution such rearrangements make to reproductive isolation between these organisms. Here, we report experiments that take an interventionist, rather than a retrospective approach to studying speciation, by reconfiguring the Saccharomyces cerevisiae genome so that it is collinear with that of Saccharomyces mikatae. We demonstrate that this imposed genomic collinearity allows the generation of interspecific hybrids that produce a large proportion of spores that are viable, but extensively aneuploid. We obtained similar results in crosses between wild-type S. cerevisiae and the naturally collinear species Saccharomyces paradoxus, but not with non-collinear crosses. This controlled comparison of the effect of chromosomal translocation on species barriers suggests a mechanism for the generation of redundancy in the S. cerevisiae genome.     

Essential role for the peroxiredoxin Prdx1 in erythrocyte antioxidant defence and tumour suppression

Reactive oxygen species are involved in many cellular metabolic and signalling processes and are thought to have a role in disease, particularly in carcinogenesis and ageing. We have generated mice with targeted inactivation of Prdx1, a member of the peroxiredoxin family of antioxidant enzymes. Here we show that mice lacking Prdx1 are viable and fertile but have a shortened lifespan owing to the development beginning at about 9 months of severe haemolytic anaemia and several malignant cancers, both of which are also observed at increased frequency in heterozygotes. The haemolytic anaemia is characterized by an increase in erythrocyte reactive oxygen species, leading to protein oxidation, haemoglobin instability, Heinz body formation and decreased erythrocyte lifespan. The malignancies include lymphomas, sarcomas and carcinomas, and are frequently associated with loss of Prdx1 expression in heterozygotes, which suggests that this protein functions as a tumour suppressor. Prdx1-deficient fibroblasts show decreased proliferation and increased sensitivity to oxidative DNA damage, whereas Prdx1-null mice have abnormalities in numbers, phenotype and function of natural killer cells. Our results implicate Prdx1 as an important defence against oxidants in ageing mice.     

Regulatory functions of γδ T cells

γδ T cells share characteristics of innate and adaptive immune cells and are involved in a broad spectrum of pro-inflammatory functions. Nonetheless, there is accumulating evidence that γδ T cells also exhibit regulatory functions. In this review, we describe the different phenotypes of regulatory γδ T cells in correlation with the identified mechanisms of suppression.     

On a Transvariation Based Measure of Group Separability

In this paper, the potentialities of transvariation (Gini, 1959) in measuring the separation between two groups of multivariate observations are explored. With this aim, a modified version of Gini’s notion of multidimensional transvariation is proposed. According to Gini (1959), two groups G₁ and G₂ are said to transvary on the k-dimensional variable X = (X₁,...,X_h,...,X_k) if there exists at least one pair of units, belonging to different groups, such that for h = 1,...,k the sign of the difference between their X_h values is opposite to that of m_1h −m_2h, where m_1h and m_2h are the corresponding group mean values of X_h. We introduce a modification that allows us to derive a measure of group separation, which can be profitably used in discriminating between two groups. The performance of the measure is tested through simulation experiments. The results show that the proposed measure is not sensitive to distributional assumptions and highlight its robustness against outliers.     

The metaphor of epigenesis: Kant,Blumenbach and Herder

Over the last few decades, the meaning of the scientific theory of epigenesis and its significance for Kant's critical philosophy have become increasingly central questions. Most recently, scholars have argued that epigenesis is a key factor in the development of Kant's understanding of reason as self-grounding and self-generating. Building on this work, our claim is that Kant appealed to not just any epigenetic theory, but specifically Johann Friedrich Blumenbach's account of generation, and that this appeal must be understood not only in terms of self-organization, but also in terms of the demarcation of a specific domain of inquiry: for Blumenbach, the study of life; for Kant, the study of reason. We argue that Kant adopted this specific epigenetic model as a result of his dispute with Herder regarding the independence of reason from nature. Blumenbach's conception of epigenesis and his separation of a domain of the living from the non-living lent Kant the tools to demarcate metaphysics, and to guard reason against Herder's attempts to naturalize it.