Permeabilization ofNeurospora crassa hyphae with toluene-ethanol and filipin

Young hyphae ofNeurospora crassa were made permeable to UDP-glucose and trypan blue by treatment with toluene-ethanol and filipin. Less than 2% of treated cells survived treatment with 8% and 16% toluene-ethanol, while 25% survived treatment with 4% toluene-ethanol. Similarly, 98% of treated cells were killed by treatment with 16 g/ml filipin. Electron microscopy revealed that toluene-ethanol-treated cells lost pieces of plasma membrane and contained a number of vacuole-like structures; filipin-treated cells were less affected. Both filipin- and toluene-ethanol-treated cells were able to incorporate UDP-glucose into insoluble material (likely glycogen and glucan).     

Reproduction and the central project of evolutionary theory

In much of the discourse of evolutionary theory, reproduction is treated as an autonomous function of the individual organism — even in discussions of sexually reproducing organisms. In this paper, I examine some of the functions and consequences of such manifestly peculiar language. In particular, I suggest that it provides crucial support for the central project of evolutionary theory — namely that of locating causal efficacy in intrinsic properties of the individual organism. Furthermore, I argue that the language of individual reproduction is maintained by certain methodological conventions that both obscure many of the problems it generates and serve to actively impede attempts to redress those difficulties that can be identified. Finally, I suggest that inclusion of the complexities introduced by sexual reproduction — in both language and methodology — may radically undermine the individualist focus of evolutionary theory.I am indepted to the Rockefeller Foundation for a Humanities Fellowship that supported this research during the spring of 1986. I am also grateful to Richard Lewontin, Diane Paul, and Lisa Lloyd for many extremely helpful conversations.     

Physicochemical Properties of Niigata Waxy Rices

The crude lipase powder has been purified 216-fold in specific activity by means of pH adjustment, DEAE-Cellu1ose, DEAE-Sephadex A-50, CM-Sephadex C-50 and Sephadex G-200 column chromatography and the recovery of the activity was 30%. The purified lipase was confirmed to be homogeneous with disc electrophoresis and ultracentrifugal analyses. The purified lipase was stable in the pH range from 7.0 to 10.0. Optimal pH for the lipolysis of polyvinyl alcohol-emulsified olive oil at 45°C was 8.0 and optimal temperature was 60°C. The purified lipase was stable up to 60°C and retained 55% of full activity after heating at 70°C for 20 min.     

Root apoplastic transport and water relations cannot account for differences in Cl− transport and Cl−/NO3 − interactions of two grapevine rootstocks differing in salt tolerance

Grapevine is moderately sensitive to salinity and accumulation of toxic levels of Cl^? in leaves is the major reason for salt-induced symptoms. In this study, apoplastic Cl^? uptake and transport mechanism(s) were investigated in two grapevine (Vitis sp.) rootstock hybrids differing in salt tolerance; 1103 Paulsen (salt tolerant) and K 51–40 (salt sensitive). Increased external salinity caused high Cl^? accumulation in shoots of the salt sensitive K 51–40 in comparison to Paulsen. Measurement of ¹⁵NO₃ ^? net fluxes under high salinity showed that by increasing external Cl^? concentrations K 51–40 roots showed reduced NO₃ ^? accumulation. This was associated with increased accumulation of Cl^?. In comparison to Paulsen, K 51–40 showed reduced NO₃ ^?/Cl^? root selectivity with increased salinity, but Paulsen had lower selectivity over the whole salinity range (0–45 mM). To examine if root hydraulic and permeability characterisations accounted for differences between varieties, the root pressure probe was used on excised roots. This showed that the osmotic Lp_r was significantly smaller than hydrostatic Lp_r, but no obvious difference was observed between the rootstocks. The reflection coefficient (σ) values (0.48–0.59) were the same for both rootstocks, and root anatomical studies showed no obvious difference in apoplastic barriers of the main and lateral roots. Comparing the uptake of Cl^? with an apoplastic tracer, PTS (3-hydroxy-5,8,10-pyrentrisulphonic acid), showed that there was no correlation between Cl^? and PTS transport. These results indicated that bypass flow of salts to the xylem is the same for both rootstocks (0.77 ± 0.2 and 1.05 ± 0.12 %) and hence pointed to differences in membrane transport to explain difference in Cl^? transport to the shoot.     

Conquered from the Deep Sea? A New Deep-Sea Isopod Species from the Antarctic Shelf Shows Pattern of Recent Colonization

The Amundsen Sea, Antarctica, is amongst the most rapidly changing environments of the world. Its benthic inhabitants are barely known and the BIOPEARL 2 project was one of the first to biologically explore this region. Collected during this expedition, Macrostylis roaldi sp. nov. is described as the first isopod discovered on the Amundsen-Sea shelf. Amongst many characteristic features, the most obvious characters unique for M. roaldi are the rather short pleotelson and short operculum as well as the trapezoid shape of the pleotelson in adult males. We used DNA barcodes (COI) and additional mitochondrial markers (12S, 16S) to reciprocally illuminate morphological results and nucleotide variability. In contrast to many other deep-sea isopods, this species is common and shows a wide distribution. Its range spreads from Pine Island Bay at inner shelf right to the shelf break and across 1,000 m bathymetrically. Its gene pool is homogenized across space and depth. This is indicative for a genetic bottleneck or a recent colonization history. Our results suggest further that migratory or dispersal capabilities of some species of brooding macrobenthos have been underestimated. This might be relevant for the species’ potential to cope with effects of climate change. To determine where this species could have survived the last glacial period, alternative refuge possibilities are discussed.     

Incorporation of beta-selenolo[3,2-b]pyrrolyl-alanine into proteins for phase determination in protein X-ray crystallography

beta-Selenolo[3,2-b]pyrrolyl-L-alanine that mimics tryptophan with the benzene ring of the indole moiety replaced by selenophene, was incorporated into human annexin V and barstar. This was achieved by fermentation and expression in a Trp-auxotrophic Escherichia coli host strain using the selective pressure incorporation method. The seleno- proteins were obtained in yields comparable to those of the wild-type proteins and exhibit full crystallographic isomorphism to the parent proteins, but expectedly show altered absorbance profiles and quenched tryptophan fluorescence. Since the occurrence of tryptophan residues in proteins is rare, incorporation of the electron-rich selenium-containing tryptophan surrogate into proteins represents a useful supplementation and even a promising novel alternative to selenomethionine for solving the phase problem in protein X-ray crystallography.     

Wnt8 is required for growth-zone establishment and development of opisthosomal segments in a spider

The Wnt genes encode secreted glycoprotein ligands that regulate many developmental processes from axis formation to tissue regeneration [1]. In bilaterians, there are at least 12 subfamilies of Wnt genes [2]. Wnt3 and Wnt8 are required for somitogenesis in vertebrates [3-7] and are thought to be involved in posterior specification in deuterostomes in general [8]. Although TCF and beta-catenin have been implicated in the posterior patterning of some short-germ insects [9, 10], the specific Wnt ligands required for posterior specification in insects and other protostomes remained unknown. Here we investigated the function of Wnt8 in a chelicerate, the common house spider Achaearanea tepidariorum[11]. Knockdown of Wnt8 in Achaearanea via parental RNAi caused misregulation of Delta, hairy, twist, and caudal and resulted in failure to properly establish a posterior growth zone and truncation of the opisthosoma (abdomen). In embryos with the most severe phenotypes, the entire opisthosoma was missing. Our results suggest that in the spider, Wnt8 is required for posterior development through the specification and maintenance of growth-zone cells. Furthermore, we propose that Wnt8, caudal, and Delta/Notch may be parts of an ancient genetic regulatory network that could have been required for posterior specification in the last common ancestor of protostomes and deuterostomes.     

Tissue microarray study for classification of breast tumors

Clinical and pathological heterogeneity of breast cancer hinders selection of appropriate treatment for individual cases. Molecular profiling at gene or protein levels may elucidate the biological variance of tumors and provide a new classification system that correlates better with biological, clinical and prognostic parameters. We studied the immunohistochemical profile of a panel of seven important biomarkers using tumor tissue arrays. The tumor samples were then classified with a monothetic (binary variables) clustering algorithm. Two distinct groups of tumors are characterized by the estrogen receptor (ER) status and tumor grade (p = 0.0026). Four biomarkers, c-erbB2, Cox-2, p53 and VEGF, were significantly overexpressed in tumors with the ER-negative (ER-) phenotype. Eight subsets of tumors were further identified according to the expression status of VEGF, c-erbB2 and p53. The malignant potential of the ER-/VEGF+ subgroup was associated with the strong correlations of Cox-2 and c-erbB2 with VEGF. Our results indicate that this molecular classification system, based on the statistical analysis of immunohistochemical profiling, is a useful approach for tumor grouping. Some of these subgroups have a relative genetic homogeneity that may allow further study of specific genetically-controlled metabolic pathways. This approach may hold great promise in rationalizing the application of different therapeutic strategies for different subgroups of breast tumors.     

Activin promotes oocyte development in ovine preantral follicles <Emphasis Type="Italic">in vitro</Emphasis>

Activins have been implicated as important regulating factors for many reproductive processes. The aim of this study was to determine the effect of activin A on the development of ovine preantral follicles in vitro. Mechanically isolated preantral follicles (161 ± 2 microm) were cultured for 6 days in the presence of human recombinant activin A (0, 10 and 100 ng/ml). Half of the medium was replaced every second day and follicle diameters were measured. Conditioned medium was subsequently analysed for oestradiol content using a delayed enhancement lanthanide fluorometric immunoassay (DELFIA). At the end of the culture period, follicles were fixed and processed for histology, after which oocyte diameter and granulosa cell death were measured. There was significant follicle growth over 6 days in all groups (p < 0.001). Activin, at both concentrations, increased follicle growth over control levels by Day 6 (p < 0.05). Oocyte diameters were also significantly increased by Day 6 of culture in all groups (p < 0.05), with 100 ng/ml activin increasing oocyte diameter over control levels (p < 0.05). Activin, at both concentrations, increased oestradiol production on Day 2 of culture, but this increase was not sustained during the culture period. Moreover, activin did not have any effect on antrum formation or follicle survival. In conclusion, activin promoted ovine preantral follicle and oocyte growth in vitro, but did not accelerate follicle differentiation over a six-day culture period. These results support a paracrine role for activin A during early oocyte and follicular development.     

Organic phosphorus in soil water under a European beech (Fagus sylvatica L.) stand in northeastern Bavaria, Germany: seasonal variability and changes with soil depth

Organically bound phosphorus (P) is a mobile form of phosphorus in many soils and thus its dynamics relevant for the leaching and cycling of this element. Despite its importance, little is known about the chemical composition of dissolved organic P. We studied the concentrations, fluxes, and chemical composition of organic P in forest floor leachates and soil solutions in a Rendzic Leptosol under a 90-year-old European beech (Fagus sylvatica L.) forest over a 27-month period (1997–1999). The chemical composition of organic P was analysed using XAD-8 fractionation and ³¹P-nuclear magnetic resonance (NMR) spectroscopy. Organic P was the dominant P form in forest floor leachates as well as in porewaters of the mineral soil. The largest concentrations of organic P were observed during summer and peaked (330–400 g dissolved organic P l^–1) after rain storms following short dry periods, concurrently with the concentrations of organic carbon (OC). Because of high rainfall, fluxes of organic P (and C) were greatest in autumn although concentrations of organic C and P were lower than in summer. In forest floor leachates, the hydrophilic fraction of dissolved organic matter contained 83 ± 13% of the bulk organic P. In soil solutions from 90 cm depth, organic P was almost exclusively in the hydrophilic fraction. Because of the low retention of the hydrophilic fraction of dissolved organic matter in the mineral soils, concentrations of organic P in soil water remained almost constant with depth. Consequently, organic P contributed > 95% of the total P leached into deeper subsoils. The overall retention of organic P in the weakly developed mineral soils was little and so the average annual fluxes of organic P in subsoils at 90 cm depth (38 mg m^–2) comprised 67% of those from the forest floors (57 mg m^–2) during the study period. Hence, organic P proved to be mobile in the studied soil. ³¹P-NMR spectroscopy confirmed the dominance of organic P species in soil water. Signals due to inorganic P occurred only in spectra of samples collected in winter and spring months. Spectra of samples from summer and autumn revealed traces of condensed phosphates. Due to low P contents, identification of organic P species in samples from winter and spring was not always possible. In summer and autumn, monoester and diester phosphates were the dominant organic species and varied little in their relative distributions. The distribution of organic species changed little from forest floor leachates to the subsoil solutions indicating that the composition of P-containing compounds was not influenced by sorptive interactions or biological transformation.