Breast cancer resistance protein (BCRP/ABCG2) is expressed by progenitor cells/reactive ductules and hepatocytes and its expression pattern is influenced by disease etiology and species type: possible functional consequences.

Breast cancer resistance protein (BCRP/ABCG2) is an ATP-binding cassette transport protein that is expressed in several organs including the liver. Previous studies have shown that ABC transport proteins play an important pathophysiological role in several liver diseases. However, to date, expression pattern and possible role of BCRP in human liver diseases and animal models have not been studied in detail. Here we investigated the expression pattern of BCRP in normal liver, chronic parenchymal and biliary human liver diseases, and parallel in different rat models of liver diseases. Expression was studied by immunohistochemistry and additionally by RT-PCR analysis in Thy-1-positive rat oval cells. Bile ducts, hepatic progenitor cells, reactive bile ductules, and blood vessel endothelium were immunoreactive for BCRP in normal liver and all types of human liver diseases and in rat models. BCRP was expressed by the canalicular membrane of hepatocytes in normal and diseased human liver, but never in rat liver. Remarkably, there was also expression of BCRP at the basolateral pole of human hepatocytes, and this was most pronounced in chronic biliary diseases. In conclusion, BCRP positivity in the progenitor cells/reactive ductules could contribute to the resistance of these cells to cytotoxic agents and xenotoxins. Basolateral hepatocytic expression in chronic biliary diseases may be an adaptive mechanism to pump bile constituents back into the sinusoidal blood. Strong differences between human and rat liver must be taken into account in future studies with animal models.     

Stringent analysis of gene function and protein-protein interactions using fluorescently tagged genes

In Drosophila collections of green fluorescent protein (GFP) trap lines have been used to probe the endogenous expression patterns of trapped genes or the subcellular localization of their protein products. Here, we describe a method, based on nonoverlapping, highly specific, shRNA transgenes directed against GFP, that extends the utility of these collections to loss-of-function studies. Furthermore, we used a MiMIC transposon to generate GFP traps in Drosophila cell lines with distinct subcellular localization patterns, which will permit high-throughput screens using fluorescently tagged proteins. Finally, we show that fluorescent traps, paired with recombinant nanobodies and mass spectrometry, allow the study of endogenous protein complexes in Drosophila.     

Overexpression of NRPS4 leads to increased surface hydrophobicity in fusarium graminearum

The plant pathogen Fusarium graminearum is the infamous cause of Fusarium head blight worldwide resulting in significant losses of yield and reduced grain feed quality. It also has the potential to produce a range of small bioactive peptides produced by the non ribosomal peptide synthetases (NRPSs). Most of these are unknown as F. graminearum contains 19 NRPS encoding genes, but only three have been assigned products. For the first time, we use deletion and overexpression mutants to investigate the functions and product of NRPS4 in F. graminearum. Deletion of NRPS4 homologues in Alternaria brassicicola and Cochloibolus heterostrophus has been shown to result in mutants unable to repel water. In a time study of surface hydrophobicity we observed that water droplets could penetrate 7 d old colonies of the NRPS4 deletion mutants. Loss in ability to repel water was first observed on 13 d old cultures of the wild type strain, whereas the overexpression strain remained water repellant throughout the 38 d time study. The conidia of both mutants were examined and those of the overexpression mutant showed distinct morphological differences in form of collapsed cells. These observations might suggest that the peptide product of NRPS4 could be an architectural factor in the cell walls of Fusarium or an indirect regulator of hydrophobicity.     

Virulence of a Klebsiella pneumoniae strain carrying the New Delhi metallo-beta-lactamase-1 (NDM-1)

The aim of the study was to compare and evaluate virulence in five strains of Klebsiella pneumoniae, including an isolate carrying New Delhi metallo-beta-lactamase-1 (NDM-1). In vivo virulence was assessed using a murine sepsis model and using the nematode Caenorhabditis elegans killing model, and in vitro virulence by assessing various virulence factors. The NDM-1 carrying K. pneumoniae isolate was the most virulent in the murine sepsis model but there was no clear cut correlation to in vitro virulence factors or killing in C. elegans. It is concluded that K. pneumoniae carrying NDM-1 have an intrinsic virulence potential, which in coexistence with its multiresistance could promote and partly explain its epidemiological success.     

An N-Myristoylated Globin with a Redox-Sensing Function That Regulates the Defecation Cycle in Caenorhabditis elegans

Globins occur in all kingdoms of life where they fulfill a wide variety of functions. In the past they used to be primarily characterized as oxygen transport/storage proteins, but since the discovery of new members of the globin family like neuroglobin and cytoglobin, more diverse and complex functions have been assigned to this heterogeneous family. Here we propose a function for a membrane-bound globin of C. elegans, GLB-26. This globin was predicted to be myristoylated at its N-terminus, a post-translational modification only recently described in the globin family. In vivo, this globin is found in the membrane of the head mesodermal cell and in the tail stomato-intestinal and anal depressor muscle cells. Since GLB-26 is almost directly oxidized when exposed to oxygen, we postulate a possible function as electron transfer protein. Phenotypical studies show that GLB-26 takes part in regulating the length of the defecation cycle in C. elegans under oxidative stress conditions.     

Developmental causes of allometry: new models and implications for phenotypic plasticity and evolution

Shapes change during development because tissues, organs, and various anatomical features differ in onset, rate, and duration of growth. Allometry is the study of the consequences of differences in the growth of body parts on morphology, although the field of allometry has been surprisingly little concerned with understanding the causes of differential growth. The power-law equation y?=?ax(b), commonly used to describe allometries, is fundamentally an empirical equation whose biological foundation has been little studied. Huxley showed that the power-law equation can be derived if one assumes that body parts grow with exponential kinetics, for exactly the same amount of time. In life, however, the growth of body parts is almost always sigmoidal, and few, if any, grow for exactly the same amount of time during ontogeny. Here, we explore the shapes of allometries that result from real growth patterns and analyze them with new allometric equations derived from sigmoidal growth kinetics. We use an extensive ontogenetic dataset of the growth of internal organs in the rat from birth to adulthood, and show that they grow with Gompertz sigmoid kinetics. Gompertz growth parameters of body and internal organs accurately predict the shapes of their allometries, and that nonlinear regression on allometric data can accurately estimate the underlying kinetics of growth. We also use these data to discuss the developmental relationship between static and ontogenetic allometries. We show that small changes in growth kinetics can produce large and apparently qualitatively different allometries. Large evolutionary changes in allometry can be produced by small and simple changes in growth kinetics, and we show how understanding the development of traits can greatly simplify the interpretation of how they evolved.     

Nos2 inactivation promotes the development of medulloblastoma in Ptch1(+/-) mice by deregulation of Gap43-dependent granule cell precursor migration

Medulloblastoma is the most common malignant brain tumor in children. A subset of medulloblastoma originates from granule cell precursors (GCPs) of the developing cerebellum and demonstrates aberrant hedgehog signaling, typically due to inactivating mutations in the receptor PTCH1, a pathomechanism recapitulated in Ptch1(+/-) mice. As nitric oxide may regulate GCP proliferation and differentiation, we crossed Ptch1(+/-) mice with mice lacking inducible nitric oxide synthase (Nos2) to investigate a possible influence on tumorigenesis. We observed a two-fold higher medulloblastoma rate in Ptch1(+/-) Nos2(-/-) mice compared to Ptch1(+/-) Nos2(+/+) mice. To identify the molecular mechanisms underlying this finding, we performed gene expression profiling of medulloblastomas from both genotypes, as well as normal cerebellar tissue samples of different developmental stages and genotypes. Downregulation of hedgehog target genes was observed in postnatal cerebellum from Ptch1(+/+) Nos2(-/-) mice but not from Ptch1(+/-) Nos2(-/-) mice. The most consistent effect of Nos2 deficiency was downregulation of growth-associated protein 43 (Gap43). Functional studies in neuronal progenitor cells demonstrated nitric oxide dependence of Gap43 expression and impaired migration upon Gap43 knock-down. Both effects were confirmed in situ by immunofluorescence analyses on tissue sections of the developing cerebellum. Finally, the number of proliferating GCPs at the cerebellar periphery was decreased in Ptch1(+/+) Nos2(-/-) mice but increased in Ptch1(+/-) Nos2(-/) (-) mice relative to Ptch1(+/-) Nos2(+/+) mice. Taken together, these results indicate that Nos2 deficiency promotes medulloblastoma development in Ptch1(+/-) mice through retention of proliferating GCPs in the external granular layer due to reduced Gap43 expression. This study illustrates a new role of nitric oxide signaling in cerebellar development and demonstrates that the localization of pre-neoplastic cells during morphogenesis is crucial for their malignant progression.     

RNA interference-mediated repression of sucrose-phosphatase in transgenic potato tubers (Solanum tuberosum) strongly affects the hexose-to-sucrose ratio upon cold storage with only minor effects on total soluble carbohydrate accumulation

Storage of potato tubers at low temperatures leads to the accumulation of glucose and fructose in a process called 'cold sweetening'. The aim of this work was to investigate the role of sucrose-phosphatase (SPP) in potato tuber carbohydrate metabolism at low temperature (4 degrees C). To this end, RNA interference (RNAi) was used to reduce SPP expression in transgenic potato tubers. Analysis of SPP specific small interfering RNAs (siRNAs), SPP protein accumulation and enzyme activity indicated that SPP silencing in transgenic tubers was stable during the cold treatment. Analysis of soluble carbohydrates showed that in transgenic tubers, cold-induced hexogenesis was inhibited while, despite strongly reduced SPP activity, sucrose levels exceeded wild-type (WT) values four- to fivefold after 34 d of cold treatment. This led to a drastic change in the hexose-to-sucrose ratio from 1.9 in WT tubers to 0.15 to 0.11 in transgenic tubers, while the total amount of soluble sugars was largely unchanged in both genotypes. Sucrose-6(F)-phosphate (Suc6P), the substrate of SPP, accumulated in transgenic tubers in the cold which most likely enables the residual enzyme to operate with maximal catalytic activity in vivo and thus, in the long term, counterbalances reduced SPP activity in the transformants. Northern analysis revealed that cold-induced expression of vacuolar invertase (VI) was blocked in SPP-silenced tubers explaining a reduced sucrose-to-hexose conversion. Suc6P levels were found to negatively correlate with VI expression. A possible role of Suc6P in regulating VI expression is discussed.     

Assemblage structure and conservation value of spiders and carabid beetles from restored lowland river banks

We assessed the composition of spider and carabid beetle assemblages along river banks from the Common Meuse (Belgium) to determine their relationships with local topographical and landscape-related characteristics. Data were gathered with pitfall traps in 1998 and explored by ordination and grouping methods. Our analysis revealed that the presence of xerothermic spider species was limited to scarcely covered, less dynamic gravel banks. Riparian spider species were found on frequently flooded as well as on rather elevated and high gravel banks, while riparian carabid beetles were dominant on all sampled banks. The level of flooding disturbance, the vegetation density and the presence of silt appeared to be the most important environmental determinants of spider and carabid beetle assemblage structure. Consequently, local environmental factors influence species occurrence patterns in a similar way for the two investigated arthropod groups. Nevertheless, distinct ecological groups are differently ordered along the prime environmental gradients. Nature management should therefore consider the conservation, restoration and connectivity of both dynamic and more elevated banks in order to obtain a high degree of local and regional heterogeneity throughout the river system.