Effect of acyl-CoA oxidase activity on the accumulation of gamma-decalactone by the yeast Yarrowia lipolytica: a factorial approach

beta-Oxidation is a cyclic pathway involved in the degradation of lipids. In yeast, it occurs in peroxisomes and the first step is catalyzed by an acyl-CoA oxidase (Aoxp). The yeast Yarrowia lipolytica possesses several genes (POX) coding for Aoxps. This study is based on the factorial analysis of results obtained with the many POX derivative strains that have been constructed previously. The effect of interactions between Aoxps on the acyl-CoA oxidase (Aox) activity was important even at the second order. We then investigated the effect of Aox activity on growth and lactone production. Aox activity was correlated with acidification of the medium by cells and with cellular growth but not with lactone production, although Aox activity on short chains was inversely correlated with lactone accumulation. Due to the poor correlation between Aox activity and lactone production, the modeling of this parameter gave no satisfactory results but growth depending on Aox activity was modeled.     

Tecia solanivora infestation increases tuber starch accumulation in Pastusa Suprema potatoes

In response to infestation with larvae of the Guatemalan tuber moth(Tecia solanivora), some Solanum tuberosum(potato) varieties exhibit an overcompensation response, whereby the total dry mass of uninfested tubers is increased. Here, we describe early responses,within the first few days, of T. solanivora feeding, in the Colombian potato variety Pastusa Suprema. Nontargeted metabolite profiling showed significant secondary metabolism changes in T. solanivora-infested tubers,but not in uninfested systemic tubers. In contrast,changes in primary metabolism were greater in uninfested systemic tubers than in the infested tubers, with a notable 80% decline in systemic tuber sucrose levels within 1 d of T. solanivora infestation. This suggested either decreased sucrose transport from the leaves orincreased sink strength, i.e., more rapid sucrose to starch conversion in the tubers. Increased sucrose synthesis was indicated by higher rubisco activase and lower starch synthase gene expression in the leaves of infested plants.Elevated sink strength was demonstrated by 45% more total starch deposition in systemic tubers of T. solanivorainfested plants compared to uninfested control plants.Thus, rather than investing in increased defense of uninfested tubers, Pastusa Suprema promotes deposition of photoassimilates in the form of starch as a response to T. solanivora infestation.     

Performance of 11 tree species under different management treatments in restoration plantings in a tropical dry forest

Ecological restoration in tropical dry forests urgently needs to incorporate experimental evidence to increase effectiveness. The main barriers for tree establishment are adverse microenvironmental conditions and competition with exotic grasses. Therefore, management should address such barriers in order to enhance tree performance. We evaluated the effect of plastic mulching, grass removal, and no management on survival after 2 months and stem volume and canopy size after 2 years and integrated response index (IRI) in plantings of 11 native tree species with different growth rates in pastures near the tropical dry forest of Chamela, Mexico. Results revealed that: (1) initial seedling mortality was minimal in all treatments (8%) and lowest under no management (2%); (2) plastic mulching, but not grass removal, leads to increased size for most species, irrespective of their growth rank; (3) a trade‐off between initial plant survival and size after 2 years occurred due to plastic mulching; and (4) most species showed similar values of the IRI because of high survival, stem volume, or canopy cover. Grass removal decreased early survival of all species and increased stem volume only for one slow‐growing species. The use of plastic mulching increased stem volume for slow‐growing species, whereas fast‐growing species developed larger canopies with that treatment. Effects of grass removal and mulching seem to be very species‐specific and not dependent in growth rank of species, although overall mulching seems to provide better conditions for seedling performance than grass removal alone.     

Determinants of sequence-specificity within human AID and APOBEC3G

Human APOBEC3G (A3G) and activation-induced deaminase (AID) belong to a family of DNA-cytosine deaminases. While A3G targets the last C in a run of C's, AID targets C in the consensus sequence WRC (W is A or T and R is a purine). Guided by the structures of the A3G carboxyl-terminal catalytic domain (A3G-CTD), we identified two potential regions (region 1 and region 2) that may interact with DNA and swapped the corresponding regions between a variant of A3G-CTD and AID. The resulting hybrids were expressed in Escherichia coli and two different genetic assays and a biochemical assay were used to determine the sequence selectivity of the hybrids in promoting C to T mutations. The results show that while the 10 amino acid region 2 of A3G was its principal sequence-specificity determinant, region 1 of A3G enhanced the target cytosine preference conferred by region 2. In contrast, neither of the two regions in AID individually or in combination were sufficient to confer the DNA sequence preference of this protein upon A3G. Instead, introduction of AID sequences in A3G relaxed the sequence-specificity of the latter protein. Our results show that the sequence selectivity of APOBEC family of enzymes is determined by at least two separate sequence segments and there may be additional regions of the protein involved in DNA sequence recognition.     

Telomerase activity in response to mild oxidative stress

We have analysed telomerase activity to determine whether it can be modified when BCL-2 is endogenously overexpressed in response to a mild oxidative stress treatment as part of a survival mechanism, in contrast with an exogenous bcl-2 overexpression due to a retroviral infection. Endogenous bcl-2 overexpression was induced after a low oxidative insult of H2O2 in mice primary lung fibroblasts and L929 cell, whereas bcl-2 exogenous overexpression was performed using a retroviral infection in L929 cells. Telomerase activity was quantified in Bcl-2 overexpressing cells by the TRAP assay. When the cells were treated with different H2O2 concentrations, only those exposed to 50 μM showed increased telomerase activity. This correlates with BCL-2 expression as part of the endogenous response to mild oxidative stress. Oxidative stress generated during the toxic mechanism of chemotherapeutic drugs might induce BCL-2 increment, enhancing telomerase activity and reactivating the oncogenic process. Clinical trials should take into consideration the possibility of telomerase activation following increased BCL-2 expression when treating patients with ROS (reactive oxygen species) generation by anti-cancer drugs.     

Production of 3-hydroxy-γ-decalactone, the precursor of two decenolides with flavouring properties, by the yeast Yarrowia lipolytica

3-Hydroxy-γ-decalactone is the precursor of dec-2 and dec-3-en-4-olides which are valuable aroma compounds not yet produced. To promote the accumulation of this lactone, the yeast Yarrowia lipolytica was placed in different environmental conditions aiming at altering β-oxidation fluxes. The concentration of substrate, pH, aeration and dissolved oxygen level were modified. We observed an important accumulation at low aeration (0.40 molar yields) and, to a lesser extent, at lower pH (0.15). As oxygen played a key-role, we evaluated its effect at fixed dissolved oxygen and at the pH which was the most favourable to the biotransformation (pH 4.5). At 5% and 30% dissolved oxygen, yields reached 0.50. β-Oxidation fluxes are very dependent on the presence of oxygen and conditions of accumulation of 3-hydroxy-γ-decalactone with very high yields were identified. These results are an important step in the production of the two decenolides. Moreover, they show the high dependence of β-oxidation fluxes on environmental conditions and relate these conditions to the accumulation of intermediates, results that are of interest to all the processes using yeast on lipids or alkanes.     

Use of dietary diversity score as a proxy indicator of nutrient adequacy of rural elderly people in Sri Lanka

ABSTRACT: BACKGROUND: Macro and micro nutrient deficiencies are public health concerns in most developing countries including Sri Lanka, partly due to monotonous, cereal-based diet that lacks diversity. The objective of the study was to assess validity of food variety score (FVS), dietary diversity score (DDS) and dietary serving score (DSS) as indicators of nutrient adequacy of rural elderly people in Sri Lanka. FINDINGS: A sample of 200 apparently healthy elderly people >60y of age were studied. A single 24 h recall was performed to compute dietary diversity indicators. Pearson's correlation was used to assess the utility of FVS, DDS and DSS as indicators of nutrient adequacy. Sensitivity (Se) and specificity (Spe) analysis were done to determine the most appropriate cut-off points for using FVS and DDS to categorize elderly people with adequate nutrient intake. The average (standard deviation) of the food variety score, dietary diversity score and dietary serving score was 8.4 (2), 4.4 (0.9) and 11.4 (2.5), respectively. Mean adequacy ratio (MAR) of 12 nutrients was 0.39 (39 %). Pearson's correlation coefficients between MAR and FVS was 0.45 (P < 0.01), for DDS it was 0.48 (P < 0.01) and for DSS it was 0.58 (P < 0.01). When maximizing sensitivity and specificity, the best cut-off point for achieving 50 % of MAR was about 9 and 4.5 for FVS and DDS, respectively. CONCLUSION: In conclusion, FVS, DDS and DSS were useful proxy indicators of nutrient adequacy of rural elderly people in Sri Lanka. Indeed, the performance of the indicators is improved when considering the quantities of food consumed.