Differential expression of a C-terminal splice variant of phosphatidylinositol transfer protein beta lacking the constitutive-phosphorylated Ser262 that localizes to the Golgi compartment

Mammalian PITPbeta (phosphatidylinositol transfer protein beta) is a 272-amino-acid polypeptide capable of transferring PtdIns, PtdCho and SM (sphingomyelin) between membrane bilayers. It has been reported that Ser262 present in the C-terminus of PITPbeta is constitutively phosphorylated and determines Golgi localization. We provide evidence for the expression of an sp (splice) variant of PITPbeta (PITPbeta-sp2) where the C-terminal 15 amino acids of PITPbeta-sp1 are replaced by an alternative C-terminus of 16 amino acids. PITPbeta-sp1 is the product of the first 11 exons, whereas PITPbeta-sp2 is a product of the first 10 exons followed by the twelfth exon--exon 11 being 'skipped'. Both splice variants are capable of PtdIns and PtdCho transfer, with PITPbeta-sp2 being unable to transport SM. PITPbeta is ubiquitously expressed, with the highest amounts of PITPbeta found in HL60 cells and in rat liver; HL60 cells express only PITPbeta-sp1, whereas rat liver expresses both sp variants in similar amounts. In both cell types, PITPbeta-sp1 is constitutively phosphorylated and both the PtdIns and PtdCho forms of PITPbeta-sp1 are present. In contrast, PITPbeta-sp2 lacks the constitutively phosphorylated Ser262 (replaced with glutamine). Nonetheless, both PITPbeta variants localize to the Golgi and, moreover, dephosphorylation of Ser262 of PITPbeta-sp1 does not affect its Golgi localization. The presence of PITPbeta sp variants adds an extra level of proteome complexity and, in rat liver, the single gene for PITPbeta gives rise to seven distinct protein species that can be resolved on the basis of their charge differences.     

Genetic Variation and the Role of Insect Life History Traits in the Ability of Drosophila Larvae to Develop in the Presence of a Competing Filamentous Fungus

Competitive interactions between organisms from distantly related phylogenetical branches have been suggested as being one of the most pervasive forms of interspecific competition. However, so-called inter-kingdom competition has rarely been the focus of ecological and evolutionary studies. Thus, a relatively novel hypothesis has been proposed on the basis that saprophagous insects might intensively compete with filamentous fungi for ephemeral resources (e.g. decaying plant tissue). Consideration that life history traits (e.g. developmental time) are adaptive in determining developmental success in the presence of con- or hetero-specifics competitors implies that these traits have been progressively established by natural selection. Because a similar scenario may apply to antagonistic interactions between saprophagous insects and filamentous fungi, one can expect the existence of heritable variation in developmental success when insect larvae are forced to grow in the presence of noxious mould. Therefore, this study aimed at discovering whether a local population of Drosophila melanogaster indeed harbours genetic variation in developmental success in the presence of the mould Aspergillus niger. By using the isofemale line technique, single larvae forced to feed on fungal infected or uninfected substrate were analysed for variation in survival probability to the adult stage, developmental time and body size of emerged adults. I found genetic variation in survival probability in fungal infected substrates but not in uninfected larval food sources. Mean developmental time and body size varied significantly among isofemale lines in both types of larval environment. Survival was negatively correlated with developmental time on fungal infected substrate, but variation in developmental time on fungal-free substrates was not correlated with survival on fungal infected food patches. Within-trait correlation between fungal infected and uninfected substrates was surprisingly weak, and developmental time was not correlated with body size. The results of this study demonstrate (a) the existence of genetic variation for larval developmental success in the presence of A. niger in a Drosophila population, and (b) heritability of important insect life history traits differed as a function of the larval environment (fungal infected or uninfected feeding substrate). I discuss models that might explain heritability differences and the evolutionary consequences of these results.     

Denture repairs in different regions of Croatia in relation to prosthodontic teams

The purpose of this paper is to evaluate the incidence of denture repairs in different districts of Croatia through the year of 2002. and to analyse the percentage of different repairs (relinings, simple repairs up to 2 elements and complicated repairs-more than 2 elements) in relation to prosthodontic teams. Data on the number of dentures, and the number and types of denture repairs delivered in the Croatian regions of Zagreb, Rijeka, Split and Karlovac were obtained from the Croatian Institute for Health Insurance for the whole of the year 2002. Information of the number of prosthodontic teams operating in those regions was also obtained. Proportionally more denture repairs were carried out in Karlovac (18%) than Split (5%). The smallest percantage of dentures that required relining was registered in Split and the highest in Rijeka (chi2 = 36.7, p < 0.01). The smallest percentage of simple repairs was registered in Rijeka and the highest in Split (chi2 = 24.3, p < 0.01). The smallest percentage of complicated repairs was registered in Split and the highest in Karlovac. In each region the proportion of denture repairs and types of repairs were correlated with a number of prosthodontic teams in that region. Karlovac had the smallest percentage of specialistic prosthodontic teams and the highest rate of denture repairs.     

Mouse stefins A1 and A2 (Stfa1 and Stfa2) differentiate between papain-like endo- and exopeptidases

Stefin A (Stfa) acts as a competitive inhibitor of intracellular papain-like cysteine proteases which play important roles in normal cellular functions such as general protein turnover, antigen processing and ovarian follicular growth and maturation. In the mouse there are at least three different variants of Stfa (Stfa1, Stfa2 and Stfa3). Recent genetic studies identified structural polymorphisms in Stfa1 and Stfa2 as candidates for Aod1b, a locus controlling susceptibility to day three thymectomy (D3Tx)-induced autoimmune ovarian disease (AOD). To evaluate the functional significance of these polymorphisms, recombinant allelic proteins were expressed in Escherichia coli, purified and characterized. The polymorphisms do not markedly alter the folding characteristics of the two proteins. Stfa1 and Stfa2 both act as fast and tight binding inhibitors of endopeptidases papain and cathepsins L and S, however their interaction with exopeptidases cathepsins B, C and H was several orders of magnitude weaker compared to human, porcine and bovine Stfa. Notwithstanding, the K(i) values for the interactions of Stfa1-b from AOD resistant C57BL/6J mice was 10-fold higher than that of the Stfa1-a allele from susceptible A/J mice for papain, cathepsins B, C and H but not L and S. In contrast, the inhibitory activities of Stfa2-a and Stfa2-b were found to be roughly equivalent for all targets peptidases.     

Alcoholic fermentation of carbon sources in biomass hydrolysates by Saccharomyces cerevisiae: current status

Fuel ethanol production from plant biomass hydrolysates by Saccharomyces cerevisiae is of great economic and environmental significance. This paper reviews the current status with respect to alcoholic fermentation of the main plant biomass-derived monosaccharides by this yeast. Wild-type S. cerevisiae strains readily ferment glucose, mannose and fructose via the Embden–Meyerhof pathway of glycolysis, while galactose is fermented via the Leloir pathway. Construction of yeast strains that efficiently convert other potentially fermentable substrates in plant biomass hydrolysates into ethanol is a major challenge in metabolic engineering. The most abundant of these compounds is xylose. Recent metabolic and evolutionary engineering studies on S. cerevisiae strains that express a fungal xylose isomerase have enabled the rapid and efficient␣anaerobic fermentation of this pentose. l-Arabinose fermentation, based on the expression of a prokaryotic pathway in S. cerevisiae, has also been established, but needs further optimization before it can be considered for industrial implementation. In addition to these already investigated strategies, possible approaches for metabolic engineering of galacturonic acid and rhamnose fermentation by S. cerevisiae are discussed. An emerging and major challenge is to achieve the rapid transition from proof-of-principle experiments under ‘academic’ conditions (synthetic media, single substrates or simple substrate mixtures, absence of toxic inhibitors) towards efficient conversion of complex industrial substrate mixtures that contain synergistically acting inhibitors.     

Parameters influencing the accuracy and practical applicability of UV indicator cards.

In order to evaluate the potential hazard for the skin and the eye presented by solar ultraviolet radiation, and to take appropriate precautions, it is necessary to characterise the biological effective irradiance or dose. Recently, inexpensive UV indicator cards became available that in principle would provide an attractive means to roughly indicate the local level of erythemal solar UV irradiance. We have characterised the properties of a number of different types of UV indicator cards. Several parameters which may influence the colour of the cards were examined with both outdoor trials under solar UV as well as indoor trials using a filtered xenon arc lamp. Our findings show that the tested cards do not give an appropriate estimation of the effective irradiance due to their spectral sensitivity and their temperature dependence. The application area of the tested UV indicator cards is therefore limited to certain temperature ranges and to seasons where a certain ratio between solar UV-A and solar UV-B occurs.     

Effects of bovine spermatozoa preparation on embryonic development in vitro

The aim of our research was to examine the ability of density gradient preparation BoviPure^? and swim up method on bull sperm separation and in vitro embryo production (IVP) systems. Frozen/thawed semen from six Simmental bulls was pooled and treated using both methods. The sperm motility, concentration, membrane activity, membrane integrity and acrosomal status were evaluated and compared before and after sperm processing using BoviPure^? and swim up methods. We also evaluated and compared cleavage rates, embryo yield and quality between the methods. There were significant differences (P < 0.05) between the sperm characteristics before and after BoviPure^?, but not after swim up method. However, there were significant differences for sperm results among those two mentioned methods. A total of 641 oocytes were matured and fertilized in vitro and cultured in SOFaaBSA. The percentage of cleavage (Day 2) and the percentage of hatched embryos (Day 9) were similar for both methods. However, embryo production rate (Day 7) was significantly higher using BoviPure^? method (P < 0.05). Also, total cell number and embryo differential staining (inner cell mass and trophectoderm cells) of Day 7 morulas and blastocysts showed that BoviPure^? treated sperm displayed higher quality embryos compared to swim up method (P < 0.05). Our results indicate that BoviPure^? method has an enhanced capacity in sperm selection for in vitro embryo production when compared with swim up method. So, we concluded that BoviPure^? could be considered as a better alternative to swim up method for separating bull spermatozoa from frozen/thawed semen for IVP of bovine embryos.     

Large-scale analysis of plasmid relationships through gene-sharing networks

Plasmids are vessels of genetic exchange in microbial communities. They are known to transfer between different host organisms and acquire diverse genetic elements from chromosomes and/or other plasmids. Therefore, they constitute an important element in microbial evolution by rapidly disseminating various genetic properties among different communities. A paradigmatic example of this is the dissemination of antibiotic resistance (AR) genes that has resulted in the emergence of multiresistant pathogenic bacterial strains. To globally analyze the evolutionary dynamics of plasmids, we built a large graph in which 2,343 plasmids (nodes) are connected according to the proteins shared by each other. The analysis of this gene-sharing network revealed an overall coherence between network clustering and the phylogenetic classes of the corresponding microorganisms, likely resulting from genetic barriers to horizontal gene transfer between distant phylogenetic groups. Habitat was not a crucial factor in clustering as plasmids from organisms inhabiting different environments were often found embedded in the same cluster. Analyses of network metrics revealed a statistically significant correlation between plasmid mobility and their centrality within the network, providing support to the observation that mobile plasmids are particularly important in spreading genes in microbial communities. Finally, our study reveals an extensive (and previously undescribed) sharing of AR genes between Actinobacteria and Gammaproteobacteria, suggesting that the former might represent an important reservoir of AR genes for the latter.