Deep EST profiling of developing fenugreek endosperm to investigate galactomannan biosynthesis and its regulation

Galactomannans are hemicellulosic polysaccharides composed of a (1 → 4)-linked β-D-mannan backbone substituted with single-unit (1 → 6)-α-linked D-galactosyl residues. Developing fenugreek (Trigonella foenum-graecum) seeds are known to accumulate large quantities of galactomannans in the endosperm, and were thus used here as a model system to better understand galactomannan biosynthesis and its regulation. We first verified the specific deposition of galactomannans in developing endosperms and determined that active accumulation occurred from 25 to 38 days post anthesis (DPA) under our growth conditions. We then examined the expression levels during seed development of ManS and GMGT, two genes encoding backbone and side chain synthetic enzymes. Based on transcript accumulation dynamics for ManS and GMGT, cDNA libraries were constructed using RNA isolated from endosperms at four ages corresponding to before, at the beginning of, and during active galactomannan deposition. DNA from these libraries was sequenced using the 454 sequencing technology to yield a total of 1.5 million expressed sequence tags (ESTs). Through analysis of the EST profiling data, we identified genes known to be involved in galactomannan biosynthesis, as well as new genes that may be involved in this process, and proposed a model for the flow of carbon from sucrose to galactomannans. Measurement of in vitro ManS and GMGT activities and analysis of sugar phosphate and nucleotide sugar levels in the endosperms of developing fenugreek seeds provided data consistent with this model. In vitro enzymatic assays also revealed that the ManS enzyme from fenugreek endosperm preferentially used GDP-mannose as the substrate for the backbone synthesis

Sugar-starvation-induced changes of carbon metabolism in excised maize root tips

International audienc

Qualité des fruits : approches métabolique, biotechnologique et génomique

National audienc

Identification and characterization of the 2-enoyl-CoA hydratases involved in peroxisomal beta-oxidation in rat liver.

In this study we attempted to determine the number of 2-enoyl-CoA hydratases involved in peroxisomal beta-oxidation. We therefore separated peroxisomal proteins from rat liver on several chromatographic columns and measured hydratase activities on the eluates with different substrates. The results indicate that rat liver peroxisomes contain two hydratase activities: (1) a hydratase activity associated with multifunctional protein 1 (MFP-1) (2-enoyl-CoA hydratase/delta 3, delta 2-enoyl-CoA isomerase/L-3-hydroxyacyl-CoA dehydrogenase) and (2) a hydratase activity associated with MFP-2 (17 beta-hydroxysteroid dehydrogenase/D-3-hydroxyacyl-CoA dehydrogenase/2-enoyl-CoA hydratase). MFP-1 forms and dehydrogenates L-3-hydroxyacyl-CoA species, whereas MFP-2 forms and dehydrogenates D-3-hydroxyacyl-CoA species. A portion of MFP-2 is proteolytically cleaved, most probably in the peroxisome, into a 34 kDa 17 beta-hydroxysteroid dehydrogenase/D-3-hydroxyacyl-CoA dehydrogenase and a 45 kDa D-specific 2-enoyl-CoA hydratase. Finally, the results confirm that MFP-1 is involved in the degradation of straight-chain fatty acids, whereas MFP-2 and its cleavage products seem to be involved in the degradation of the side chain of cholesterol (bile acid synthesis)

Risk factors for liver injury in patients with acquired immunodeficiency syndrome treated with nevirapine

ObjectiveTo investigate the risk factors for hypersensitivity-associated liver injury induced by the combined antiretroviral therapy (c-ART) including nevirapine (NVP) in patients with acquired immunodeficiency syndrome. MethodsThe clinical data and blood samples of 132 patients who received the combined therapy including NVP in Zhongnan Hospital of Wuhan University from June 2008 to October 2015 were collected, and PCR-SSP was used to determine the genotypes of human leukocyte antigen (HLA) DRB1 and HLA-Cw. The patients who experienced hypersensitivity-associated liver injury induced by NVP within 6 weeks of c-ART were enrolled in the liver injury group (41 patients), and those who did not experience liver injury were enrolled in the control group (91 patients). The risk factors for liver injury induced by NVP hypersensitivity were analyzed. The t-test was used for comparison of continuous data between groups; the chi-square test was used for comparison of categorical data between groups. The univariate logistic regression method was used to analyze the risk factors for liver injury associated with NVP hypersensitivity, and the variables with P＜0.10 were included in the multivariate logistic regression model to perform stepwise regression analysis. The Spearman correlation coefficient was used to analyze the correlation between the number of CD4 cells and alanine aminotransferase (ALT) level in patients experiencing hypersensitivity-associated liver injury. ResultsThe results of the multivariate logistic regression analysis showed that male sex (OR=12.297, 95%CI: 2.467-61.300, P=0002), a high CD4 cell count at baseline (OR=1.010, 95%CI: 1.001-1.018, P=0.022), HCV co-infection(OR=10.598, 95%CI: 1.411-79.613, P=0.022), and a HLA-Cw*03 carrier (OR=34.119, 95%CI: 5.543-210.023, P＜0001) were risk factors for liver injury associated with NVP hypersensitivity. In the patients with HCV co-infection or a high CD4 cell count (≥200/μl) or carrying HLA-Cw*03 allele, male patients had a significantly higher incidence rate of liver injury than female patients (63.9% vs 11.6%, χ2=23.390, P＜0.001). Baseline CD4 cell count was positively correlated with ALT level (r=0.583, P＜0001). ConclusionMale patients infected with human immunodeficiency virus who are co-infected with HCV and have a high CD4 cell count at baseline should avoid using NVP. The value of HLA-Cw*03 gene screening in predicting hepatotoxicity associated with NVP hypersensitivity awaits further investigation