Dynamic and Reversible Aggregation of the Human CAP Superfamily Member GAPR-1 in Protein Inclusions in Saccharomyces cerevisiae

Many proteins that can assemble into higher order structures termed amyloids can also concentrate into cytoplasmic inclusions via liquid–liquid phase separation. Here, we study the assembly of human Golgi-Associated plant Pathogenesis Related protein 1 (GAPR-1), an amyloidogenic protein of the Cysteine-rich secretory proteins, Antigen 5, and Pathogenesis-related 1 proteins (CAP) protein superfamily, into cytosolic inclusions in Saccharomyces cerevisiae. Overexpression of GAPR-1-GFP results in the formation GAPR-1 oligomers and fluorescent inclusions in yeast cytosol. These cytosolic inclusions are dynamic and reversible organelles that gradually increase during time of overexpression and decrease after promoter shut-off. Inclusion formation is, however, a regulated process that is influenced by factors other than protein expression levels. We identified N-myristoylation of GAPR-1 as an important determinant at early stages of inclusion formation. In addition, mutations in the conserved metal-binding site (His54 and His103) enhanced inclusion formation, suggesting that these residues prevent uncontrolled protein sequestration. In agreement with this, we find that addition of Zn²⁺ metal ions enhances inclusion formation. Furthermore, Zn²⁺ reduces GAPR-1 protein degradation, which indicates stabilization of GAPR-1 in inclusions. We propose that the properties underlying both the amyloidogenic properties and the reversible sequestration of GAPR-1 into inclusions play a role in the biological function of GAPR-1 and other CAP family members.     

Temporal and spatial trends in heavy metal concentrations in the marine mussel Mytilus edulis from the Western Scheldt estuary (The Netherlands)

Since the first North Sea Conference (1984, Bremen), all countries bordering the North Sea made commitments to reduce discharge of hazardous substances into the North Sea. From Belgium and The Netherlands, large reductions (upto 90) in heavy metal emissions from land-based sources have been reported between 1985 and 2000. Recently, some studies in the Western Scheldt estuary (WS) have shown that total metal concentration in the water, sediments and suspended particles have decreased compared to levels in the 70s. However, data on aquatic organisms is still very limited and it is therefore difficult to confirm whether the reductions in pollution input and generally improving water quality in the WS have a corresponding impact on the levels of heavy metals in aquatic organisms. The current study measured metal concentrations in the soft tissues of mussels, Mytilus edulis (known to be good indicators of environmental metal pollution) during the period 1996–2002. Spatial (salinity and pollution gradients), temporal and seasonal variations were also studied. Results showed a down-stream decreasing trend for the metals studied (Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb and Zn) during all sampling campaigns. There was also a significant seasonal effect on tissue metal concentrations, with a peak observed around spring in both WS and the nearby less polluted Eastern Scheldt (ES). On temporal trends, a clear drop of metals in mussels was observed in the early 80s coinciding with the start of the efforts to reduce chemical pollution input into the North Sea. Since those early reductions, metal concentrations in mussels generally remained unchanged upto mid 90s. However, in recent times metal concentration in mussels have increased significantly, for example Cd in 2002 was almost 10 times the values in 1983 and similar to levels seen during the peak in the 70s. Other metals also increased in the 90s also reaching levels seen in the 70s. As there is no indication of recent increase in metal input into the estuary, we suggest that increased metal concentrations observed in mussels in recent years especially in the upper estuary are most likely a result of changes in physical and chemical speciation and metal bioavailability. Such changes may be caused by changes in some water quality parameters in the estuary (i.e. increased dissolved oxygen, concentration of organic matter), resulting in conditions that favour releases of sediment-bound metals into the water column. The relationship between metal content and season showed very similar annual profiles in the polluted WS and less polluted ES. Thus, seasonal variations in metal concentrations appear to be largely controlled by biological processes, while total body burdens are dependent on environmental levels and bioavailability.     

Evidence for the presence of lipopolysaccharide in Treponema phagedenis (biotype Reiterii) but not in Treponema pallidum (Nichols)

Abstract Immunoblotting profiles of whole or protease-K-digested organisms with homologous antisera demonstrated the presence of a characteristic ladder pattern of smooth LPS in Treponema phagedenis . Periodic acid silver staining of SDS-PAGE gels confirmed these findings. However, when heterologous or homologous serum was reacted with Treponema pallidum , no such pattern or cross-reactions were observed. The significance of apparent absence of LPS in T. pallidum is discussed.     

The rise of coastal Middle Bronze Age Levant – A multidisciplinary approach for investigating in Sidon,Lebanon

Objectives

The Levantine Middle Bronze Age (MBA, circa 2000–1500 BCE) marks a period of increased trade and regional interaction, spurred on by technological developments. In light of previous research exhibiting limited mobility in Sidon, further investigation was conducted using biodistance analysis to understand local population history and site development.

Materials and Methods

Dental nonmetric traits, a proxy for genetic information, were explored using ASUDAS on a sub-sample of primary inhumations (n = 35). The biodistance matrix was generated using Gower distance measures, and further tested using PERMDISP, PERMANOVA, Mantel test and hierarchical cluster analysis. The data was also contrasted to ⁸⁷Sr/⁸⁶Sr and δ¹⁸O as well as δ¹³C and δ¹⁵N values.

Results

There were no significant diachronic differences in isotopes values, and there was biological continuity (n = 35, Mantel test r = 0.11, p = 0.02, comparing local phases and biodistance). The analysis also suggested of a sub-group of individuals with biological proximity shared a more limited range of mobility and dietary habits.

Conclusions

The isotopes (⁸⁷Sr/⁸⁶Sr, δ¹⁸O, δ¹³C, δ¹⁵N) and biodistance analysis conducted on the Sidon College site skeletal assemblage exhibits stability and continuity of the people, despite the site's increasing role in the maritime network. This continuity may have been a key factor in Sidon's success, allowing it to accumulate wealth and resources for centuries to come.

     

Body size increase in insular rodent populations: a role for predators?

Insular mammalian populations living in areas of small size are often characterized by a drastic change in body mass compared to related continental populations or species. Generally, small mammals (less than 100 g) evolve into giant forms while large mammals (up to 100 g) evolve into dwarf forms. These changes, coupled with changes in other life, behavioural, physiological or demographic traits are referred to generally as the insular syndrome. We tested in this study the relative contribution of three factors — area of island, numbers of competitor species and number of predator species — to changes in body size of the woodmouse (Apodemus sylvaticus) in the Western Mediterranean Sea. Our results, based on a comparative analysis using the phylogenetic independent contrasts method, indicate that the increase in body size is related both to the decrease of island size and to the lower number of predator species. A decrease of competitor species does not seem to have an important effect.     

Presence and development of populations of the introduced brown alga Sargassum muticum in the southwest Netherlands

Of the three Brachionus species used in aquaculture, Brachionus rubens, B. calycilorus and B. plicatilis, the latter is most widely used in raising marine fish and shrimp larvae due to its tolerance to the marine environment. In freshwater aquaculture the use of B. rubens and B. calycilorus is limited, probably because inert food products are readily available as feed for freshwater larvae.The rotifer Brachionus plicatilis is used in large numbers as the first food organism in intensive cultures of marine fish and shrimp larvae. An adequate supply of these rotifers relies on mass cultures. The reproductive rate of rotifers in these cultures depends on food quality and quantity, salinity, temperature and pH of the medium. Removal of waste products from culture tanks leads to higher and more efficient production of rotifers over extended periods of time. Rotifers have to be enriched with polyunsaturated fatty acids, which are essential for proper development and survival of marine fish and shrimp larvae.The future use of preserved rotifers and their resting eggs may help to overcome unforeseen failures of live cultures and may lead to more efficient use of these organisms in raising freshwater and marine fish and shrimp larvae.     

Immunochemically detectable phytochrome is present at normal levels but is photochemically nonfunctional in the hy 1 and hy 2 long hypocotyl mutants of Arabidopsis

The hy 1 and hy 2 long hypocotyl mutants of Arabidopsis thaliana contain less than 20% (the detection limit) of the phytochrome in wild-type tissue as measured by in vivo difference spectroscopy. In contrast, spectral measurements for the hy 3, hy 4, and hy 5 long hypocotyl mutants indicate that they each contain levels of phytochrome equivalent to the wild-type parent. Immunoblot analysis using a monoclonal antibody directed against the chromophore-bearing region of etiolated-oat phytochrome demonstrates that extracts of all mutant and wild-type Arabidopsis tissues, prepared by extraction of proteins into hot SDS-containing buffer, have identical levels of one major immunodetectable protein (116 kDa). An assay involving controlled in vitro proteolysis, known to produce distinctive fragmentation patterns for Pr and Pfr (Vierstra RD, Quail PH, Planta 156: 158–165, 1982), indicates that the 116 kDa polypeptide from the wild-type parent represents Arabidopsis phytochrome. The 116 kDa protein from either hy 3, hy 4, or hy 5 displays the same fragmentation pattern found for the wild type. Together with the spectral data, these results indicate that the mutant phenotype of these variants does not involve lesions in the polypeptide sequence that lead to gross conformational aberrations, and suggest that the genetic lesions may affect steps in the transduction chain downstream of the photoreceptor. In contrast, this same analysis for hy 1 and hy 2 has revealed that the 116 kDa protein from either of these mutants is not degraded differently in response to the different wavelengths of irradiation given in vitro. Moreover, whereas immunoblot analysis of tissue extracts from light-grown wild-type seedlings show that the 116 kDa phytochrome protein level is greatly reduced relative to dark-grown tissue as expected, similar extracts of light-grown hy 1 and hy 2 seedlings contain the 116 kDa polypeptide in amounts equivalent to those of dark-grown tissue. Combined, these data indicate that the hy 1 and hy 2 mutants both produce normal levels of immunochemically detectable phytochrome that is photochemically nonfunctional.     

Unusually strong immunological cross-reaction between elongation factor Tu of Escherichia coli and Bacillus subtilis

Polyclonal antibodies were prepared against the purified elongation factor Tu (EF-Tu) of Escherichia coli and Bacillus subtilis. Using the methods of Western blotting and microcomplement fixation the cross-reactivities of EF-Tu of 19 different prokaryotes were determined. The immunological distance were compared with the results of 16S rRNA oligonucleotide analysis. An unexpectedly high cross-reactivity was revealed between the EF-Tu of B. subtilis and the antiserum against the EF-Tu of E. coli. A comparison of the predicted amino acid sequences from the tuf-genes of E. coli and B. subtilis yielded two identical peptide fragments that are likely candidates for antibody binding sites.Abbreviations EF-Tu elongation factor Tu - GDP guanosine 5-diphosphate - GTP guanosine 5-triphosphate - MCF microcomplement fixation - T type strain     

Uplift of the roof of africa and its bearing on the evolution of mankind

Evidence concerning the geomorphological evolution of the Western Rift Valley, sedimentation within the valley and comparison of the fossil mammalian faunas of Western Uganda and East Africa indicate that the mountain ranges which now flank the Western Rift were uplifted in three or more stages beginning during the upper Miocene and that they reached climatically important altitudes during the upper Pliocene, at which time they began to modify regional climatic patterns in East Africa. Their main effect was the xerification of conditions over much of the region east of the mountains. The regional climatic effects due to the mountain ranges were themselves modified by global climatic changes related to the onset of the Glacial Period, the two phenomena combining to yield the Present day climatic regime of East Africa. As the climate changed, so did the flora and fauna. Faunal response was of three main kinds: a) dispersal into East Africa of pre-existing forms already adapted to more xeric conditions (many bovids, some cercopithecids), b) autochthonous evolution of forms adapted to mesic environments into forms adapted to more xeric conditions (suids, elephantids, some bovids, hominids), c) displacement of species ranges of those lineages unable to adapt to changing conditions (i.e. local extinctions) (Anancus, Brachypotherium). Autochthonous evolvers, including hominids, adopted two main strategies reflected in their hard anatomy: a) dietary shift (suids, proboscideans, bovids and later Pliocene hominids) and b) locomotor changes (early Pliocene hominids).