YcbX and yiiM, two novel determinants for resistance of Escherichia coli to N-hydroxylated base analogues

We have shown previously that lack of molybdenum cofactor (MoCo) in Escherichia coli leads to hypersensitivity to the mutagenic and toxic effects of N -hydroxylated base analogues, such as 6- N -hydroxylaminopurine (HAP). However, the nature of the MoCo-dependent mechanism is unknown, as inactivation of all known and putative E. coli molybdoenzymes does not produce any sensitivity. Presently, we report on the isolation and characterization of two novel HAP-hypersensitive mutants carrying defects in the ycbX or yiiM open reading frames. Genetic analysis suggests that the two genes operate within the MoCo-dependent pathway. In the absence of the ycbX - and yiiM -dependent pathways, biotin sulfoxide reductase plays also a role in the detoxification pathway. YcbX and YiiM are hypothetical members of the MOSC protein superfamily, which contain the C-terminal domain (MOSC) of the eukaryotic MoCo sulphurases. However, deletion of ycbX or yiiM did not affect the activity of human xanthine dehydrogenase expressed in E. coli , suggesting that the role of YcbX and YiiM proteins is not related to MoCo sulphuration. Instead, YcbX and YiiM may represent novel MoCo-dependent enzymatic activities. We also demonstrate that the MoCo/YcbX/YiiM-dependent detoxification of HAP proceeds by reduction to adenine.     

A perinatal nitric oxide donor increases renal vascular resistance and ameliorates hypertension and glomerular injury in adult fawn-hooded hypertensive rats

Enhancing perinatal nitric oxide (NO) availability persistently reduces blood pressure in spontaneously hypertensive rats. We hypothesize that this approach can be generalized to other models of genetic hypertension, for instance those associated with renal injury. Perinatal exposure to the NO donor molsidomine was studied in fawn-hooded hypertensive (FHH) rats, a model of mild hypertension, impaired preglomerular resistance, and progressive renal injury. Perinatal molsidomine increased urinary NO metabolite excretion at 8 wk of age, i.e., 4 wk after treatment was stopped (P < 0.05). Systolic blood pressure was persistently reduced after molsidomine (42-wk females: 118 +/- 3 vs. 141 +/- 5 and 36-wk males: 139 +/- 4 vs. 158 +/- 4 mmHg; both P < 0.001). Perinatal treatment decreased glomerular filtration rate (P < 0.05) and renal blood flow (P < 0.01) and increased renal vascular resistance (P < 0.05), without affecting filtration fraction, suggesting persistently increased preglomerular resistance. At 4 wk of age natriuresis was transiently increased by molsidomine (P < 0.05). Molsidomine decreased glomerulosclerosis (P < 0.05). Renal blood flow correlated positively with glomerulosclerosis in control (P < 0.001) but not in perinatally treated FHH rats. NO dependency of renal vascular resistance was increased by perinatal molsidomine. Perinatal enhancement of NO availability can ameliorate development of hypertension and renal injury in FHH rats. Paradoxically, glomerular protection by perinatal exposure to the NO donor molsidomine may be due to persistently increased preglomerular resistance. The mechanisms by which increased perinatal NO availability can persistently reprogram kidney function and ameliorate hypertension deserve further study.     

Characteristics of active transport of thyroid hormone into rat hepatocytes

Thyroid hormone uptake into primary cultured rat hepatocytes was studied using 1-min incubations with radio-iodine-labelled iodothyronines. (1) Uptake of thyroxine indicates two saturable sites apparent K_m values of 1.2 nM and 1.0 μM, and non-saturable uptake. Similar kinetics of triiodothyronine uptake have been observed. (2) The high-affinity systems of both hormones are energy-dependent (i.e., inhibited by KCN and oligomycin). It is postulated that these systems represent active transport of thyroid hormone into the cell. (3) Analysis of mutual inhibition by the substrates for the triiodothyronine and thyroxine transport systems indicates that triiodothyromine and thyroxine cross the cell membrane via separate transport systems. (4) Preincubation with ouabain resulted in a decrease in uptake of both triiodothyronine and thyroxine, suggesting that a sodium gradient is essential for this transport.     

Algorithmic co-optimization of genetic constructs and growth conditions: application to 6-ACA,a potential nylon-6 precursor

Optimizing bio-production involves strain and process improvements performed as discrete steps. However, environment impacts genotype and a strain that is optimal under one set of conditions may not be under different conditions. We present a methodology to simultaneously vary genetic and process factors, so that both can be guided by design of experiments (DOE). Advances in DNA assembly and gene insulation facilitate this approach by accelerating multi-gene pathway construction and the statistical interpretation of screening data. This is applied to a 6-aminocaproic acid (6-ACA) pathway in Escherichia coli consisting of six heterologous enzymes. A 32-member fraction factorial library is designed that simultaneously perturbs expression and media composition. This is compared to a 64-member full factorial library just varying expression (0.64 Mb of DNA assembly). Statistical analysis of the screening data from these libraries leads to different predictions as to whether the expression of enzymes needs to increase or decrease. Therefore, if genotype and media were varied separately this would lead to a suboptimal combination. This is applied to the design of a strain and media composition that increases 6-ACA from 9 to 48 mg/l in a single optimization step. This work introduces a generalizable platform to co-optimize genetic and non-genetic factors.     

Analysis of the demersal community of fish and cephalopods on the Agulhas Bank, South Africa

The demersal fish and cephalopod communities of the continental shelf and upper slope from 17 to 395m deep were studied during five annual cruises between Cape Agulhas and Port Alfred, South Africa. The cruises showed a consistent pattern of an inshore community (<100m), a shelf community ( c . 90–190m) and a shelf-edge/upper slope fauna (>200m). These groups were identified by dendrograms and multidimensional scaling cluster analysis, which supported on-board observations of catch variation with depth. Although the boundaries are not clearly defined, examination of physical features at the clustered stations suggests that depth, temperature and, to a lesser extent, oxygen concentration are important in the grouping. Occasional, apparently anomalous associations of inshore stations suggested that water temperature and oxygen may over-ride the normal depth distributions of the species groups. This intimates that patterns offish and cephalopod distribution may be dynamic and in part related to the physical parameters of the water body.     

Effect of flower traits and hosts on the abundance of parasitoids in perennial multiple species wildflower strips sown within oilseed rape (<Emphasis Type="Italic">Brassica napus</Emphasis>) crops

Reducing the use of insecticides is an important issue for agriculture today. Sowing wildflower strips along field margins or within crops represents a promising tool to support natural enemy populations in agricultural landscapes and, thus, enhance conservation biological control. However, it is important to sow appropriate flower species that attract natural enemies efficiently. The presence of prey and hosts may also guide natural enemies to wildflower strips, potentially preventing them from migrating into adjacent crops. Here, we assessed how seven flower traits, along with the abundance of pollen beetles (Meligethes spp., Coleoptera: Nitidulidae) and true weevils (Ceutorhynchus spp., Coleoptera: Curculionidae), affect the density of parasitoids of these two coleopterans in wildflower strips sown in an oilseed rape field in Gembloux (Belgium). Only flower traits, not host (i.e. pollen beetles and true weevils) abundance, significantly affected the density of parasitoids. Flower colour, ultraviolet reflectance and nectar availability were the main drivers affecting parasitoids. These results demonstrate how parasitoids of oilseed rape pests react to flower cues under field conditions. Similar analyses on the pests and natural enemies of other crops are expected to help to develop perennial flower mixtures able to enhance biological control throughout a rotation system.     

A solution approach for deriving alternative fuel station infrastructure requirements

When an alternative fuel is introduced, the infrastructure through which that fuel is made available to the market is often underdeveloped. Transportation service providers relying on such infrastructures are unlikely to adopt alternative fuel vehicles as it may impose long detours for refueling. In this paper, we design and apply a new solution approach to derive minimum infrastructure requirements, in terms of the number of alternative fuel stations. The effectiveness of our approach is demonstrated by applying it to the case of introducing liquefied natural gas (LNG) as a transportation fuel in The Netherlands. From this case, we learn that, depending on the driving range of the LNG trucks and the size of area on which those trucks operate, a minimum of 5–12 LNG fuel stations is necessary to render LNG trucks economically and environmentally beneficial.