Desaturation of oxygenated fatty acids in Lesquerella and other oil seeds

Species of the genus Lesquerella, within the Brassicaceae family, have seed oils containing hydroxy fatty acids. In most Lesquerella species, either lesquerolic (14-hydroxy-eicosa-11-enoic), auricolic (14-hydroxy-eicosa-11,17-dienoic) or densipolic (12-hydroxy-octadeca-9,15-dienoic) acid dominates in the seed oils. Incubations of developing seed from Lesquerella species with 1-¹⁴C-fatty acids were conducted in order to study the biosynthetic pathways of these hydroxylated fatty acids. [¹⁴C]Oleic (octadeca-9-enoic) acid, but not [¹⁴C]linoleic (octadeca-9,12-dienoic) acid, was converted into the hydroxy fatty acid, ricinoleic (12-hydroxy-octadeca-9-enoic) acid, which was rapidly desaturated to densipolic (12-hydroxy-octadeca-9,15-dienoic) acid. In addition, [¹⁴C] ricinoleic acid added to Lesquerella seeds was efficiently desaturated at the 15 carbon. A pathway for the biosynthesis of the various hydroxylated fatty acids in Lesquerella seeds is proposed. The demonstration of desaturation at position 15 of a fatty acid with a hydroxy group at position 12 in Lesquerella prompted a comparison of the substrate recognition of the desaturases from Lesquerella and linseed. It was demonstrated that developing linseed also was able to desaturate ricinoleate at position 15 into densipolic acid. In addition, the linseed 15 desaturase was able to desaturate vernolic (12,13-epoxy-octadeca-9-enoic) acid and safflower microsomal 12 desaturase was able to desaturate 9-hydroxy-stearate. Thus, hydroxy and epoxy groups may substitute for double bonds in substrate recognition for oil-seed 12 and 15 desaturases.Abbreviations GLC gas-liquid chromatography - lysoPC palmitoyl-lysophosphatidylcholine - PC phosphatidylcholine This work was supported by grants from Stifteisen Svensk Oljeväxtforskning, Skanska Lantmännen Foundation, Swedish Farmers Foundation for Agricultural research, The Swedish Natural Science Research Council and The Swedish Council for Forestry and Agricultural Research. Nicki Engeseth was supported by the National Science Foundation under a grant award in 1992.     

Population-Level Scale-Up of Cervical Cancer Prevention Services in a Low-Resource Setting: Development,Implementation, and Evaluation of the Cervical Cancer Prevention Program in Zambia

Background

Very few efforts have been undertaken to scale-up low-cost approaches to cervical cancer prevention in low-resource countries.

Methods

In a public sector cervical cancer prevention program in Zambia, nurses provided visual-inspection with acetic acid (VIA) and cryotherapy in clinics co-housed with HIV/AIDS programs, and referred women with complex lesions for histopathologic evaluation. Low-cost technological adaptations were deployed for improving VIA detection, facilitating expert physician opinion, and ensuring quality assurance. Key process and outcome indicators were derived by analyzing electronic medical records to evaluate program expansion efforts.

Findings

Between 2006-2013, screening services were expanded from 2 to 12 clinics in Lusaka, the most-populous province in Zambia, through which 102,942 women were screened. The majority (71.7%) were in the target age-range of 25–49 years; 28% were HIV-positive. Out of 101,867 with evaluable data, 20,419 (20%) were VIA positive, of whom 11,508 (56.4%) were treated with cryotherapy, and 8,911 (43.6%) were referred for histopathologic evaluation. Most women (87%, 86,301 of 98,961 evaluable) received same-day services (including 5% undergoing same-visit cryotherapy and 82% screening VIA-negative). The proportion of women with cervical intraepithelial neoplasia grade 2 and worse (CIN2+) among those referred for histopathologic evaluation was 44.1% (1,735/3,938 with histopathology results). Detection rates for CIN2+ and invasive cervical cancer were 17 and 7 per 1,000 women screened, respectively. Women with HIV were more likely to screen positive, to be referred for histopathologic evaluation, and to have cervical precancer and cancer than HIV-negative women.

Interpretation

We creatively disrupted the ''no screening'' status quo prevailing in Zambia and addressed the heavy burden of cervical disease among previously unscreened women by establishing and scaling-up public-sector screening and treatment services at a population level. Key determinants for successful expansion included leveraging HIV/AIDS program investments, and context-specific information technology applications for quality assurance and filling human resource gaps.     

Transcriptional and biochemical regulation of a novel Arabidopsis thaliana bifunctional aspartate kinase-homoserine dehydrogenase gene isolated by functional complementation of a yeast hom6 mutant

An aspartate kinase-homoserine dehydrogenase (AK-HSDH) cDNA of Arabidopsis thaliana has been cloned by functional complementation of a Saccharomyces cerevisiae strain mutated in its homoserine dehydrogenase (HSDH) gene (hom6). Two of the three isolated clones were also able to complement a mutant yeast aspartate kinase (AK) gene (hom3). Sequence analysis showed that the identified gene (akthr2), located on chromosome 4, is different from the previously cloned A. thaliana AK-HSDH gene (akthr1), and corresponds to a novel bifunctional AK-HSDH gene. Expression of the isolated akthr2 cDNA in a HSDH-less hom6 yeast mutant conferred threonine and methionine prototrophy to the cells. Cell-free extracts contained a threonine-sensitive HSDH activity with feedback properties of higher plant type. Correspondingly, cDNA expression in an AK-deficient hom3 yeast mutant resulted in threonine and methionine prototrophy and a threonine-sensitive AK activity was observed in cell-free extracts. These results confirm that akthr2 encodes a threonine-sensitive bifunctional enzyme. Transgenic Arabidopsis thaliana plants (containing a construct with the promoter region of akthr2 in front of the gus reporter gene) were generated to compare the expression pattern of the akthr2 gene with the pattern of akthr1 earlier described in tobacco. The two genes are simultaneously expressed in meristematic cells, leaves and stamens. The main differences between the two genes concern the time-restricted or absent expression of the akthr2 gene in the stem, the gynoecium and during seed formation, while akthr1 is less expressed in roots.     

Methane oxidation coupled to oxygenic photosynthesis in anoxic waters

Freshwater lakes represent large methane sources that, in contrast to the Ocean, significantly contribute to non-anthropogenic methane emissions to the atmosphere. Particularly mixed lakes are major methane emitters, while permanently and seasonally stratified lakes with anoxic bottom waters are often characterized by strongly reduced methane emissions. The causes for this reduced methane flux from anoxic lake waters are not fully understood. Here we identified the microorganisms and processes responsible for the near complete consumption of methane in the anoxic waters of a permanently stratified lake, Lago di Cadagno. Interestingly, known anaerobic methanotrophs could not be detected in these waters. Instead, we found abundant gamma-proteobacterial aerobic methane-oxidizing bacteria active in the anoxic waters. In vitro incubations revealed that, among all the tested potential electron acceptors, only the addition of oxygen enhanced the rates of methane oxidation. An equally pronounced stimulation was also observed when the anoxic water samples were incubated in the light. Our combined results from molecular, biogeochemical and single-cell analyses indicate that methane removal at the anoxic chemocline of Lago di Cadagno is due to true aerobic oxidation of methane fuelled by in situ oxygen production by photosynthetic algae. A similar mechanism could be active in seasonally stratified lakes and marine basins such as the Black Sea, where light penetrates to the anoxic chemocline. Given the widespread occurrence of seasonally stratified anoxic lakes, aerobic methane oxidation coupled to oxygenic photosynthesis might have an important but so far neglected role in methane emissions from lakes.     

The distribution of the NADPH regenerating mannitol cycle among fungal species

The mannitol cycle is an important NADPH regenerating system in Alternaria alternata. The cycle is built up of the following enzymes: mannitol 1-phosphate dehydrogenase, mannitol 1-phosphatase, mannitol dehydrogenase and hexokinase. The net reaction of one cycle turn is: NADH+NADP⁺+ATP NAD⁺+NADPH+ADP+P_i. The enzymes needed for an operating cycle were found in Aspergillus, Botrytis, Penicillium, Pyricularia, Trichothecium, Cladosporium and Thermomyces all genera belonging to Fungi Imperfecti. The only genus of this class lacking the cycle was Candida. No genera from the classes Basidiomycetes and Phycomycetes showed any mannitol 1-phosphate dehydrogenase or mannitol 1-phosphatase activities. The genera investigated, belonging to Ascomycetes, Gibberella, Ceratocystis and Neurospora all lacked mannitol 1-phosphate dehydrogenase. It was concluded that the mannitol cycle is an important and widespread pathway for NADH oxidation and NADP⁺ reduction in the organisms belonging to the class Fungi Imperfecti.     

From swimming to walking: a single basic network for two different behaviors

 In this paper we consider the hypothesis that the spinal locomotor network controlling trunk movements has remained essentially unchanged during the evolutionary transition from aquatic to terrestrial locomotion. The wider repertoire of axial motor patterns expressed by amphibians would then be explained by the influence from separate limb pattern generators, added during this evolution. This study is based on EMG data recorded in vivo from epaxial musculature in the newt Pleurodeles waltl during unrestrained swimming and walking, and on a simplified model of the lamprey spinal pattern generator for swimming. Using computer simulations, we have examined the output generated by the lamprey model network for different input drives. Two distinct inputs were identified which reproduced the main features of the swimming and walking motor patterns in the newt. The swimming pattern is generated when the network receives tonic excitation with local intensity gradients near the neck and girdle regions. To produce the walking pattern, the network must receive (in addition to a tonic excitation at the girdles) a phasic drive which is out of phase in the neck and tail regions in relation to the middle part of the body. To fit the symmetry of the walking pattern, however, the intersegmental connectivity of the network had to be modified by reversing the direction of the crossed inhibitory pathways in the rostral part of the spinal cord. This study suggests that the input drive required for the generation of the distinct walking pattern could, at least partly, be attributed to mechanosensory feedback received by the network directly from the intraspinal stretch-receptor system. Indeed, the input drive required resembles the pattern of activity of stretch receptors sensing the lateral bending of the trunk, as expressed during walking in urodeles. Moreover, our results indicate that a nonuniform distribution of these stretch receptors along the trunk can explain the discontinuities exhibited in the swimming pattern of the newt. Thus, separate limb pattern generators can influence the original network controlling axial movements not only through a direct coupling at the central level but also via a mechanical coupling between trunk and limbs, which in turn influences the sensory signals sent back to the network. Taken together, our findings support the hypothesis of a phylogenetic conservatism of the spinal locomotor networks generating axial motor patterns from agnathans to amphibians. Received: 12 October 2001 / Accepted in revised form: 16 May 2002 Correspondence to: T. Bem (e-mail: tiaza.bem@ibib.waw.pl)