Functional expression of oxidative phosphorylation proteins in the rod outer segment disc

The rod Outer Segment (OS) disc, an organelle devoid of mitochondria, is specialized in phototransduction, a process requiring a continual chemical energy supply. We have shown that OS discs express functional mitochondrial electron transport chains, F_oF₁‐ATP synthase and the tricarboxylic acid cycle enzymes, all mitochondrial features. Here, we focus on oxygen consumption and adenosine triphosphate (ATP) synthesis by OS discs analysing electron transport chain I‐III‐IV and II‐II‐IV pathways, supported by reduced nicotinamide adenine dinucleotide and succinate, respectively. Interestingly, respiratory capacity of discs was measurable also in the presence of 3‐hydroxy‐butyrrate, a typical metabolic substrate for the brain. Data were supported by a two‐dimensional electrophoresis analyses conducted as our previous one, but focused to those mitochondrial proteins that are involved in oxidative phosphorylation. Carbonic anhydrase was also found active in OS discs. Moreover, colocalization of Rhodopsin with respiratory complex I and ATP synthase seems a further step in the characterization of some proteins typical of the mitochondrial inner membranes that are expressed in the rod discs. The existence of oxygen utilization in the outer retina, likely supplying ATP for phototransduction, may shed light on some retinal pathologies related to oxidative stress of the outer retina. Copyright © 2013 John Wiley & Sons, Ltd.     

Response of planktonic communities to calcium hydroxide addition in a hardwater eutrophic lake: results from a mesocosm experiment

The main focus of this study was to investigate the effects of lime treatment with Ca(OH)₂ on a hypereutrophic hardwater lake (Lake Alserio, Northern Italy), focusing on its impact on plankton communities. We performed a mesocosm experiment using two large enclosures, one for treatment and one control, in the lake. The addition of Ca(OH)₂ proved to be effective in reducing phosphorus concentration, but the effect was limited in time. Redissolution of the precipitating calcium carbonate as it came into contact with the deep water determined a phosphorus increase about 2 weeks after the liming. The liming had a strong impact on the plankton assemblage, as was demonstrated by the sharp decrease in phytoplankton and zooplankton biomass and density, and 1 month after the treatment we observed a different taxonomic composition inside the enclosure from that of the lake. The result of the experiment showed that liming with Ca(OH)₂ can be regarded as a suitable treatment for accelerating recovery of the lake. Multiple treatments will be necessary to improve the trophic level over a longer period.     

PTK 7 Is a Transforming Gene and Prognostic Marker for Breast Cancer and Nodal Metastasis Involvement

Protein Tyrosin Kinase 7 (PTK7) is upregulated in several human cancers; however, its clinical implication in breast cancer (BC) and lymph node (LN) is still unclear. In order to investigate the function of PTK7 in mediating BC cell motility and invasivity, PTK7 expression in BC cell lines was determined. PTK7 signaling in highly invasive breast cancer cells was inhibited by a dominant-negative PTK7 mutant, an antibody against the extracellular domain of PTK7, and siRNA knockdown of PTK7. This resulted in decreased motility and invasivity of BC cells. We further examined PTK7 expression in BC and LN tissue of 128 BC patients by RT-PCR and its correlation with BC related genes like HER2, HER3, PAI1, MMP1, K19, and CD44. Expression profiling in BC cell lines and primary tumors showed association of PTK7 with ER/PR/HER2-negative (TNBC-triple negative BC) cancer. Oncomine data analysis confirmed this observation and classified PTK7 in a cluster with genes associated with agressive behavior of primary BC. Furthermore PTK7 expression was significantly different with respect to tumor size (ANOVA, p = 0.033) in BC and nodal involvement (ANOVA, p = 0.007) in LN. PTK7 expression in metastatic LN was related to shorter DFS (Cox Regression, p = 0.041). Our observations confirmed the transforming potential of PTK7, as well as its involvement in motility and invasivity of BC cells. PTK7 is highly expressed in TNBC cell lines. It represents a novel prognostic marker for BC patients and has potential therapeutic significance.     

Saposin B mobilizes lipids from cholesterol-poor and bis(monoacylglycero)phosphate-rich membranes at acidic pH. Unglycosylated patient variant saposin B lacks lipid-extraction capacity

Sphingolipid activator proteins (SAPs), GM2 activator protein (GM2AP) and saposins (Saps) A-D are small, enzymatically inactive glycoproteins of the lysosome. Despite of their sequence homology, these lipid-binding and -transfer proteins show different specificities and varying modes of action. Water-soluble SAPs facilitate the degradation of membrane-bound glycosphingolipids with short oligosaccharide chains by exohydrolases at the membrane-water interface. There is strong evidence that degradation of endocytosed components of the cell membrane takes place at intraendosomal and intralysosomal membranes. The inner membranes of the lysosome differ from the limiting membrane of the organelle in some typical ways: the inner vesicular membranes lack a protecting glycocalix, and they are almost free of cholesterol, but rich in bis(monoacylglycero)phosphate (BMP), the anionic marker lipid of lysosomes. In this study, we prepared glycosylated Sap-B free of other Saps by taking advantage of the Pichia pastoris expression system. We used immobilized liposomes as a model for intralysosomal vesicular membranes to probe their interaction with recombinantly expressed Sap-B. We monitored this interaction using SPR spectroscopy and an independent method based on the release of radioactively labelled lipids from liposomal membranes. We show that, after initial binding, Sap-B disturbs the membrane structure and mobilizes the lipids from it. Lipid mobilization is dependent on an acidic pH and the presence of anionic lipids, whereas cholesterol is able to stabilize the liposomes. We also show for the first time that glycosylation of Sap-B is essential to achieve its full lipid-extraction activity. Removal of the carbohydrate moiety of Sap-B reduces its membrane-destabilizing quality. An unglycosylated Sap-B variant, Asn215His, which causes a fatal sphingolipid storage disease, lost the ability to extract membrane lipids at acidic pH in the presence of BMP.     

Can population genetic structure be predicted from life-history traits?

Population genetic structure is a key parameter in evolutionary biology. Earlier comparative studies have shown that genetic structure depends on species ecological attributes and life-history traits, but species phylogenetic relatedness had not been accounted for. Here we reevaluate the relationships between genetic structure and species traits in seed plants. Each species is characterized by a set of life-history and ecological features as well as by its geographic range size, its heterozygote deficit, and its genetic structure at nuclear and organelle markers to distinguish between pollen- and seed-mediated gene flow. We use both a conventional regression approach and a method that controls for phylogenetic relationships. Once phylogenetic conservatism and covariation among traits are taken into account, genetic structure is shown to be related with only a few synthetic traits, such as mating system for nuclear markers and seed dispersal mode or geographic range size for organelle markers. Along with other studies on invasiveness or rarity, our work illustrates the fact that predicting the fate of species across a broad taxonomic assemblage on the basis of simple traits is rarely possible, a testimony of the highly contingent nature of evolution.     

Exploring the association between Trypanosoma cruzi infection in rural communities and environmental changes in the southern Gran Chaco

The association between land use and land cover changes between 1979-2004 in a 2.26-million-hectare area south of the Gran Chaco region and Trypanosoma cruzi infection in rural communities was analysed. The extent of cultural land, open and closed forests and shrubland up to 3,000 m around rural communities in the north, northwest and west of the province of Córdoba was estimated using Landsat satellite imagery. The T. cruzi prevalence was estimated with a cross-sectional serological survey conducted in the rural communities. The land cover showed the same patterns in the 1979, 1999 and 2004 satellite imagery in both the northwest and west regions, with shrinking regions of cultured land and expanding closed forests away from the community. The closed forests and agricultural land coverage in the north region showed the same trend as in the northwest and west regions in 1979 but not in 1999 or 2004. In the latter two years, the coverage remote from the communities was either constant or changed in opposite ways from that of the northwest and west regions. The changes in closed forests and cultured vegetation alone did not have a significant, direct relationship with the occurrence of rural communities with at least one person infected by T. cruzi. This study suggests that the overall decrease in the prevalence of T. cruzi is a consequence of a combined effect of vector control activities and changes in land use and land cover.     

Effects of contemporary agricultural land cover on Colorado potato beetle genetic differentiation in the Columbia Basin and Central Sands

Landscape structure, which can be manipulated in agricultural landscapes through crop rotation and modification of field edge habitats, can have important effects on connectivity among local populations of insects. Though crop rotation is known to influence the abundance of Colorado potato beetle (CPB; Leptinotarsa decemlineata Say) in potato (Solanum tuberosum L.) fields each year, whether crop rotation and intervening edge habitat also affect genetic variation among populations is unknown. We investigated the role of landscape configuration and composition in shaping patterns of genetic variation in CPB populations in the Columbia Basin of Oregon and Washington, and the Central Sands of Wisconsin, USA. We compared landscape structure and its potential suitability for dispersal, tested for effects of specific land cover types on genetic differentiation among CPB populations, and examined the relationship between crop rotation distances and genetic diversity. We found higher genetic differentiation between populations separated by low potato land cover, and lower genetic diversity in populations occupying areas with greater crop rotation distances. Importantly, these relationships were only observed in the Columbia Basin, and no other land cover types influenced CPB genetic variation. The lack of signal in Wisconsin may arise as a consequence of greater effective population size and less pronounced genetic drift. Our results suggest that the degree to which host plant land cover connectivity affects CPB genetic variation depends on population size and that power to detect landscape effects on genetic differentiation might be reduced in agricultural insect pest systems.     

Purification and characterization of phycocyanin from the blue-green alga Aphanizomenon flos-aquae

Aphanizomenon flos-aquae (AFA) is a blue-green alga and represents a nutrient-dense food source. In this study the presence of phycocyanin (PC), a blue protein belonging to the photosynthetic apparatus, has been demonstrated in AFA. An efficient method for its separation has been set up: PC can be purified by a simple single step chromatographic run using a hydroxyapatite column (ratio A620/A280 of 4.78), allowing its usage for health-enhancing properties while eliminating other aspecific algal components. Proteomic investigation and HPLC analysis of purified AFA phycobilisomes revealed that, contrary to the well-characterized Synechocystis and Spirulina spp., only one type of biliprotein is present in phycobilisomes: phycocyanins with no allo-phycocyanins. Two subunit polypeptides of PC were also separated: the beta subunit containing two bilins as chromophore and the alpha subunit containing only one.