Rat genetics: the next episode

More than a thousand quantitative trait loci (QTLs) relevant to many aspects of complex human disease have been identified in a wide range of rat inbred lines over the past few decades. With the complete rat genome available, it is now time for the next stage in rat genetic research: the identification and causal confirmation of underlying polymorphisms and genes. Recent developments in single nucleotide polymorphism-based genetic markers and technologies to manipulate the rat genome will undoubtedly be important tools in this next episode.     

Dissimilation of C3-sulfonates

Cysteate and sulfolactate are widespread natural products in the environment, while propanesulfonate, 3-aminopropanesulfonate and propane-1,3-disulfonate are xenobiotics. While some understanding of the bacterial assimilation of cysteate sulfur has been achieved, details of the dissimilation of cysteate and sulfolactate by microbes together with information on the degradation of the xenobiotics have only recently become available. This minireview centres on bacterial catabolism of the carbon moiety in these C₃-sulfonates and on the fate of the sulfonate group. Three mechanisms of desulfonation have been established. Firstly, cysteate is converted via sulfopyruvate to sulfolactate, which is desulfonated to pyruvate and sulfite; the latter is oxidized to sulfate by a sulfite dehydrogenase and excreted as sulfate in Paracoccus pantotrophus NKNCYSA. Secondly, sulfolactate can be converted to cysteate, which is cleaved in a pyridoxal 5′-phosphate-coupled reaction to pyruvate, sulfite and ammonium ions; in Silicibacter pomeroyi DSS-3, the sulfite is excreted largely as sulfite. Both desulfonation reactions seem to be widespread. The third desulfonation mechanism is oxygenolysis of, e.g. propanesulfonate(s), about which less is known.     

Changing Estuaries, Changing Views

During the design and the execution of the Delta project, carried out after the storm flood of 1953 in the SW Netherlands, the importance of the long-term effects of morphological and ecological developments driven by tides and currents, have been underestimated. Due to these processes the height of the dams will have to be increased for centuries to come, because the land behind the levees cannot grow in elevation anymore with the rising of the sea. Maintenance of the civil-engineering structures, and mitigating their unpredictable impacts on ecosystems, involve very high recurrent costs. The chance of flooding is reduced, but the potential damage after a storm flood is enlarged: seawalls and dykes provide a false sense of safety against flooding. Changes in the role of agricultural use in the European context, offer an opportunity to abandon arable fields and to retrocede them to the sea in order to absorb tidal energy and to allow the land to rise concomitant with the sea. A cost-benefit analysis of this approach should assess the direct and indirect economic values, as well as the non-use (intrinsic) values, whereby public engagement in management questions, facilitates decision-making processes. Reversible and resilient economic measures within the limits of the natural processes are preferable. A future, speculative perspective is an urbanised landscape, where people and investments are located in safe places, e.g. on floating, or sea-encircled artificial dwelling-mounds, surrounded by a landscape that is ruled by the forces of nature. New approaches such as developed in the Westerschelde offer flexible solutions to flooding problems, and are worth a broader evaluation. A worldwide platform of experts should be organised to study the future management of estuaries and deltas, and to develop and exchange new ideas and techniques.     

Sensitive fluorescence in situ hybridization signal detection in maize using directly labeled probes produced by high concentration DNA polymerase nick translation

The signal produced by fluorescence in situ hybridization (FISH) often is inconsistent among cells and sensitivity is low. Small DNA targets on the chromatin are difficult to detect. We report here an improved nick translation procedure for Texas red and Alexa Fluor 488 direct labeling of FISH probes. Brighter probes can be obtained by adding excess DNA polymerase I. Using such probes, a 30 kb yeast transgene, and the rp1, rp3 and zein multigene clusters were clearly detected.     

The GTPase Activity of Murine Guanylate-binding Protein 2 (mGBP2) Controls the Intracellular Localization and Recruitment to the Parasitophorous Vacuole of Toxoplasma gondii

One of the most abundantly IFN-γ-induced protein families in different cell types is the 65-kDa guanylate-binding protein family that is recruited to the parasitophorous vacuole of the intracellular parasite Toxoplasma gondii. Here, we elucidate the relationship between biochemistry and cellular host defense functions of mGBP2 in response to Toxoplasma gondii. The wild type protein exhibits low affinities to guanine nucleotides, self-assembles upon GTP binding, forming tetramers in the activated state, and stimulates the GTPase activity in a cooperative manner. The products of the two consecutive hydrolysis reactions are both GDP and GMP. The biochemical characterization of point mutants in the GTP-binding motifs of mGBP2 revealed amino acid residues that decrease the GTPase activity by orders of magnitude and strongly impair nucleotide binding and multimerization ability. Live cell imaging employing multiparameter fluorescence image spectroscopy (MFIS) using a Homo-FRET assay shows that the inducible multimerization of mGBP2 is dependent on a functional GTPase domain. The consistent results indicate that GTP binding, self-assembly, and stimulated hydrolysis activity are required for physiological localization of the protein in infected and uninfected cells. Ultimately, we show that the GTPase domain regulates efficient recruitment to T. gondii in response to IFN-γ.     

The CpG island methylator phenotype in colorectal cancer: progress and problems

In recent years, attention has focused on the biology and potential clinical importance of the CpG island methylator phenotype (CIMP) in colorectal cancer (CRC). While it is generally well accepted that etiologically and clinically distinct subgroups exist in this disease, a precise definition of CIMP remains to be established. Here, we summarize existing literature that documents the prevalence of CIMP in CRC, with particular attention to the various methods and definitions used to classify a tumor as CIMP positive. Through a systematic review on both case-series and population based studies, we examined only original research articles reporting on sporadic CRC and/or adenomas in unselected cases. Forty-eight papers published between January 1999 and August 2011 met the inclusion criteria. We describe the use of multiple gene panels, marker threshold values, and laboratory techniques which results in a wide range in the prevalence of CIMP. Because there is no universal standard or consensus on quantifying the phenotype, establishing its true prevalence is a challenge. This bottleneck is becoming increasingly evident as molecular pathological epidemiology continues to offer possibilities for clear answers regarding environmental risk factors and disease trends. For the first time, large, unselected series of cases are available for analysis, but comparing populations and pooling data will remain a challenge unless a universal definition of CIMP and a consensus on analysis can be reached, and the primary cause of CIMP identified.     

Plant-driven weathering of apatite - the role of an ectomycorrhizal fungus

Ectomycorrhizal (EcM) fungi are increasingly recognized as important agents of mineral weathering and soil development, with far‐reaching impacts on biogeochemical cycles. Because EcM fungi live in a symbiotic relationship with trees and in close contact with bacteria and archaea, it is difficult to distinguish between the weathering effects of the fungus, host tree and other micro‐organisms. Here, we quantified mineral weathering by the fungus Paxillus involutus, growing in symbiosis with Pinus sylvestris under sterile conditions. The mycorrhizal trees were grown in specially designed sterile microcosms in which the supply of soluble phosphorus (P) in the bulk media was varied and grains of the calcium phosphate mineral apatite mixed with quartz, or quartz alone, were provided in plastic wells that were only accessed by their fungal partner. Under P limitation, pulse labelling of plants with ¹⁴CO₂ revealed plant‐to‐fungus allocation of photosynthates, with 17 times more ¹⁴C transferred into the apatite wells compared with wells with only quartz. Fungal colonization increased the release of P from apatite by almost a factor of three, from 7.5 (±1.1) × 10^?10 mol m^?2 s^?1 to 2.2 (±0.52) × 10^?9 mol m^?2 s^?1. On increasing the P supply in the microcosms from no added P, through apatite alone, to both apatite and orthophosphate, the proportion of biomass in roots progressively increased at the expense of the fungus. These three observations, (i) proportionately more plant energy investment in the fungal partner under P limitation, (ii) preferential fungal transport of photosynthate‐derived carbon towards patches of apatite grains and (iii) fungal enhancement of weathering rate, reveal the tightly coupled plant–fungal interactions underpinning enhanced EcM weathering of apatite and its utilization as P source.     

Regulatory T cells in human lymphatic filariasis: stronger functional activity in microfilaremics

Infection with filarial parasites is associated with T cell hyporesponsiveness, which is thought to be partly mediated by their ability to induce regulatory T cells (Tregs) during human infections. This study investigates the functional capacity of Tregs from different groups of filarial patients to suppress filaria-specific immune responses during human filariasis. Microfilaremic (MF), chronic pathology (CP) and uninfected endemic normal (EN) individuals were selected in an area endemic for Brugia timori in Flores island, Indonesia. PBMC were isolated, CD4CD25(hi) cells were magnetically depleted and in vitro cytokine production and proliferation in response to B. malayi adult worm antigen (BmA) were determined in total and Treg-depleted PBMC. In MF subjects BmA-specific T and B lymphocyte proliferation as well as IFN-gamma, IL-13 and IL-17 responses were lower compared to EN and CP groups. Depletion of Tregs restored T cell as well as B cell proliferation in MF-positives, while proliferative responses in the other groups were not enhanced. BmA-induced IL-13 production was increased after Treg removal in MF-positives only. Thus, filaria-associated Tregs were demonstrated to be functional in suppressing proliferation and possibly Th2 cytokine responses to BmA. These suppressive effects were only observed in the MF group and not in EN or CP. These findings may be important when considering strategies for filarial treatment and the targeted prevention of filaria-induced lymphedema.