Mass production of Dictyostelium discoideum in homogeneous and heterogeneous cultivation systems

The social amoeba Dictyostelium discoideum is a promising host for the expression of recombinant proteins requiring post-translational modifications. Limited maximal cell densities and slow growth rates, however, disfavor its application. Little attention has been paid to improve its cultivation. Here, some strategies are described, which allow the attainment of higher cell densities. This can be achieved by cultivation on an improved synthetic medium as well as in immobilized form. Two promising inorganic porous supports are presented—broken pumice and a ceramic catalyst carrier. Cell densities up to 4.5×10⁷ ml⁻¹ are obtained during suspension cultivation on the improved synthetic medium. This is about three times as much as can be expected for cultivations on conventional complex media. Cells in the pores of broken pumice and a ceramic support reach up to 15–20 times higher local cell densities compared with cells growing on conventional axenic media in suspension. Thus, the cell density of 3.5×10⁸ ml⁻¹ in the ceramic carrier (CeramTec^®) is the highest cell density for D. discoideum observed so far in cultivation systems. The immobilized cell density could be maintained for a long period of time by either repeated medium replacement or continuous cultivation. To minimize the external volume of the medium a trickle bed reactor was operated continuously.     

Nitrogen Cycles: Past, Present, and Future

This paper contrasts the natural and anthropogenic controls on the conversion of unreactive N₂ to more reactive forms of nitrogen (Nr). A variety of data sets are used to construct global N budgets for 1860 and the early 1990s and to make projections for the global N budget in 2050. Regional N budgets for Asia, North America, and other major regions for the early 1990s, as well as the marine N budget, are presented to Highlight the dominant fluxes of nitrogen in each region. Important findings are that human activities increasingly dominate the N budget at the global and at most regional scales, the terrestrial and open ocean N budgets are essentially disconnected, and the fixed forms of N are accumulating in most environmental reservoirs. The largest uncertainties in our understanding of the N budget at most scales are the rates of natural biological nitrogen fixation, the amount of Nr storage in most environmental reservoirs, and the production rates of N₂ by denitrification.     

Phosphorus accumulation in the littoral zone of a subtropical lake

In situ mesocosm experiments were performed under summer (1997) and winter (1999) conditions in the littoral zone of a subtropical lake in Florida, USA. The objective was to quantify phosphorus (P) accumulation by various components of the community after adding pulsed doses of dissolved inorganic P. A short-term experiment also was done to quantify the rate of P loss from the water column, with simultaneous use of an inert tracer to confirm that P depletion was not due to leakage of the tanks. In the experiments, added P was rapidly removed from the water; samples collected 3–4 days after adding spikes of near 100 μg l^?1 P contained little or no soluble reactive P. In the short-term experiment, we documented that the half-life of added P was approximately 6–8 h in the water column, and that the tanks were not exchanging water with the surrounding lake. Little of the added P ended up in plankton, rooted vascular plants, or sediments. The main sink for P was periphyton, including surface algal mats, benthic algal mats and detritus, and epiphyton. In the summer 1997 experiment, the periphyton was intimately associated with a non-rooted plant (Utricularia), which also may have sequestered P from the water. Structure of the littoral community varied between summer and winter, and this influenced which periphyton component accounted for most of the P removal. In regard to P mass balances, we accounted for 54% of the added P in 1997, when coarse sampling was done. In 1999, when there was more detailed sampling of the community, 92% of the added P was located in various community components. Subtropical littoral periphyton can be a large sink for P, as long as depth and underwater irradiance conditions favor its growth.     

Universal false discovery rate estimation methodology for genome-wide association studies

Genome-wide case-control association studies aim at identifying significant differential markers between sick and healthy populations. With the development of large-scale technologies allowing the genotyping of thousands of single nucleotide polymorphisms (SNPs) comes the multiple testing problem and the practical issue of selecting the most probable set of associated markers. Several False Discovery Rate (FDR) estimation methods have been developed and tuned mainly for differential gene expression studies. However they are based on hypotheses and designs that are not necessarily relevant in genetic association studies. In this article we present a universal methodology to estimate the FDR of genome-wide association results. It uses a single global probability value per SNP and is applicable in practice for any study design, using any statistic. We have benchmarked this algorithm on simulated data and shown that it outperforms previous methods in cases requiring non-parametric estimation. We exemplified the usefulness of the method by applying it to the analysis of experimental genotyping data of three Multiple Sclerosis case-control association studies.     

The Genetic Legacy of Religious Diversity and Intolerance: Paternal Lineages of Christians, Jews, and Muslims in the Iberian Peninsula

Most studies of European genetic diversity have focused on large-scale variation and interpretations based on events in prehistory, but migrations and invasions in historical times could also have had profound effects on the genetic landscape. The Iberian Peninsula provides a suitable region for examination of the demographic impact of such recent events, because its complex recent history has involved the long-term residence of two very different populations with distinct geographical origins and their own particular cultural and religious characteristics—North African Muslims and Sephardic Jews. To address this issue, we analyzed Y chromosome haplotypes, which provide the necessary phylogeographic resolution, in 1140 males from the Iberian Peninsula and Balearic Islands. Admixture analysis based on binary and Y-STR haplotypes indicates a high mean proportion of ancestry from North African (10.6%) and Sephardic Jewish (19.8%) sources. Despite alternative possible sources for lineages ascribed a Sephardic Jewish origin, these proportions attest to a high level of religious conversion (whether voluntary or enforced), driven by historical episodes of social and religious intolerance, that ultimately led to the integration of descendants. In agreement with the historical record, analysis of haplotype sharing and diversity within specific haplogroups suggests that the Sephardic Jewish component is the more ancient. The geographical distribution of North African ancestry in the peninsula does not reflect the initial colonization and subsequent withdrawal and is likely to result from later enforced population movement—more marked in some regions than in others—plus the effects of genetic drift.     

Steemann Nielsen and the zooplankton

E. Steemann Nielsen is remembered by most biological oceanographers and limnologists as having introduced the ¹⁴C method for measuring photosynthesis in 1952. The present paper is to recall that he was interested in the phytoplankton as part of the plankton community and was much aware of the role of grazing in affecting, if not determining, the concentrations of phytoplankton and, thus, also its rate of production. His principal statements to this effect were made with the open, oligotrophic subtropical and tropical oceans in mind where phytoplankton concentrations exhibit little seasonal change. This paper shows that Steemann Nielsen's sentiment also applies to non-static situations, especially phytoplankton blooms. Of the blooms in Cushing's North Sea Calanus patches of 1949 and 1954 and the two low-latitude, open-sea iron fertilization experiments (IronEx I, II) of the 1990s, more than half or even most of the newly formed cells were lost daily. In these examples, the same water was revisited, mixing was considered, and sinking was an unimportant loss term, so that grazing was the principal cause of mortality. Because of the grazing losses and the subsequent regeneration the CO₂ draw down in the fertilized water was much lower than the ¹⁴C uptake. Moreover the examples show that over the course of the blooms, the rate and even the sign of temporal change of phytoplankton abundance had little relation to the rate of cell division, as already postulated by Riley's 1946 model of the seasonal cycle of phytoplankton on Georges Bank. Thus, in most situations in the open sea and, presumably, large lakes, the rates of cell division (instead of photosynthesis by itself) and of mortality (most often from grazing) are needed for understanding and predicting the temporal change of phytoplankton abundance, a principal goal of biological oceanography. The mechanism maintaining the actual abundance of phytoplankton in the quasi-steady state prevailing over most of the ocean much of the time is still unclear.     

Zooplankton on the move: first results on the quantification of dispersal of zooplankton in a set of interconnected ponds

In systems of interconnected ponds or lakes, the dispersal of zooplankton may be mediated by the active population component, with rivulets and overflows functioning as dispersal pathways and the dispersal being unidirectional. Such systems offer the possibility to study the impact of dispersal rates on local population dynamics and community structure, and provide opportunities to quantify dispersal in the field in a straightforward manner. In this study, dispersal of active zooplankton populations among interconnected ponds was quantified directly in the field by sampling the small waterways connecting the ponds. The number of dispersing zooplankton sampled in connecting elements was on average high (almost 7000 ind h^–1). However, the contribution of dispersing individuals to total population size in the target ponds was very limited (< 1% 24 h^–1.). Only a weak diel pattern in dispersal rates was observed.     

Degradation of the Retinoblastoma Tumor Suppressor by the Human Papillomavirus Type 16 E7 Oncoprotein Is Important for Functional Inactivation and Is Separable from Proteasomal Degradation of E7

The steady-state level and metabolic half-life of retinoblastoma tumor suppressor protein pRB are decreased in cells that express high-risk human papillomavirus (HPV) E7 proteins. Here we show that pRB degradation is a direct activity of E7 and does not reflect a property of cell lines acquired during the selection process for E7 expression. An amino-terminal domain of E7 that does not directly contribute to pRB binding but is required for transformation is also necessary for E7-mediated pRB degradation. Treatment with inhibitors of the 26S proteasome not only blocks E7-mediated pRB degradation but also causes the stabilization of E7. Mutagenic analyses, however, reveal that the processes of proteasomal degradation of E7 and pRB are not linked processes. HPV type 16 E7 also targets the pRB-related proteins p107 and p130 for destabilization by a proteasome-dependent mechanism. Using the SAOS2 flat-cell assay as a biological indicator for pRB function, we demonstrate that pRB degradation, not solely binding, is important for the E7-induced inactivation of pRB.