Construction of an RNAi vector for knockdown of GM-ACS genes in the cotyledonary nodes of soybean

Background: Ethylene plays an important role in the regulation of floral organ development in soybean, and 1-aminocyclopropane-1-carboxylate synthase (ACS) is a rate-limiting enzyme for ethylene biosynthesis. However, whether ACS also regulates floral organ differentiation in soybean remains unknown. To address this, we constructed an RNAi vector to inhibit ACS expression in cotyledonary nodes. Linear DNA cassettes of RNAi-ACS obtained by PCR were used to transform soybean cotyledonary nodes. Results: In total, 131 of 139 transiently transformed plants acquired herbicide resistance and displayed GUS activities in the new buds. In comparison to untransformed seedling controls, a greater number of flower buds were differentiated at the cotyledonary node; GM-ACS1 mRNA expression levels and ethylene emission in the transformed buds were reduced. Conclusion: These results indicate that the cotyledonary node transient transformation system may be suitable for stable transformation and that the inhibition of ACS expression may be an effective strategy for promoting floral organ differentiation in soybean.

The effects of ethylene on the HCl-extractability of trace elements during soybean seed germination

Background Ethylene is capable of promoting seed germination in some plant species. Mobilization of metals such as Fe, Cu, Mn, and Zn in mature seeds takes place when seeds are germinating. However, whether ethylene is involved in the regulation of soybean seed germination and metal element mobilization during early seed germination stage remains unknown. In the present study, seeds were treated with ethylene synthesis inhibitor aminoethoxyvinylglycine (AVG) and ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC), and double distilled H2O (ddH(2)0) treatment was used as control. Ethylene emission, ACC synthase (ACS) expression, ACS enzyme activity and Ca, Zn, Mn, Cu and Fe content in hypocotyls were qualified to analyze the relationship between ethylene and mobilization of these elements. Results The results showed that ACS expression, ACS enzyme activity and ethylene emission peaked at 1 and 7 d after sowing. AVG inhibited ethylene production, promoted the hypocotyls length, ACS expression and its activity, concentrations of total and HCl-extractable Zn, and HCl-extractable Fe in hypocotyls, while ACC caused opposite effects. AVG and ACC treatment had no significantly effects on total and HCl-extractable Ca, Cu and HCl-extractable Mn. Total Mn concentration was promoted by AVG at 1, 3, and 5 d significantly, while ACC treatment tended to have no significantly effects on Mn concentration. Conclusion These findings suggested that ethylene is at least partly involved in the regulation of soybean seed germination. Remobilization of Zn and Fe may be negatively regulated by ethylene.

New insights into steroidogenesis in normo- and hyperandrogenic polycystic ovary syndrome patients / Novos aspectos na esteroidogênese em pacientes normo ou hiperandrogênicas com síndrome de ovários policísticos

OBJECTIVE: This study sought to examine corticosteroidogenic enzyme activities in normo- and hyperandrogenic polycystic ovary syndrome (PCOS) patients. SUBJECTS AND METHODS: This cohort study included 81 patients with biochemical hyperandrogenism and 41 patients with normal androgen levels. Enzyme activities were assessed according to the serum steroid product/precursor ratios at baseline and after adrenal stimulation. RESULTS: At baseline, in the delta 4 (Δ4) pathway, hyperandrogenic patients showed greater 17-hydroxylase and 17,20 lyase activities in converting progesterone (P4) into 17-hydroxyprogesterone (17-OHP4) and 17-hydroxypregnenolone (17-OHPE) into androstenedione (A) (p = 0.0005 and p = 0.047, respectively) compared to normoandrogenic patients. In the delta 5 (Δ5) pathway, the 17-hydroxylase and 17,20 lyase enzymes showed similar activities in both groups. Hyperandrogenic patients presented lower 21-hydroxylase, lower 11β-hydroxylase (p = 0.0001), and statistically significant increases in 3β-hydroxysteroid dehydrogenase II (3β-HSDII) activities (p < 0.0001). Following tetracosactrin stimulation, only the 17,20 lyase activity remained up-regulated in the Δ4 pathway (p < 0.0001). CONCLUSION: Hyperandrogenic patients had higher 17,20 lyase activity, both at baseline and after adrenal stimulation. Greater conversion of dehydroepiandrosterone (DHEA) into A with normal conversion of 17-OHPE to 17-OHP4 in hyperandrogenic PCOS patients indicated different levels of 3β-HSDII activity in adrenal cells, and hyperandrogenic patients had lower 11β-hydroxylase and 21-hydroxylase activities.

OBJETIVO: O objetivo deste estudo foi examinar a atividade de enzimas responsáveis pela produção de corticosteroides em pacientes normo e hiperandrogênicas com síndrome de ovários policísticos (SOP). SUJEITOS E MÉTODOS: A coorte estudada incluiu 81 pacientes com hiperandrogenismo bioquímico e 41 pacientes com níveis normais de androgênio. A atividade enzimática foi avaliada de acordo com as proporções de produto/precursor do esteroide sérico, no momento inicial do estudo e depois de estimulação adrenal. RESULTADOS: No momento inicial, na via delta 4 (Δ4), as pacientes hiperandrogênicas mostraram maior atividade da 17-hidroxilase e 17,20 liase na conversão da progesterona (P4) em 17-hidroxiprogesterona (17-OHP4) e na conversão da 17-hidroxipregnenolona (17-OHPE) em androstenediona (A) (p = 0,0005 e p = 0,047, respectivamente) em comparação com pacientes normoandrogênicas. Na via delta 5 (Δ5), a 17-hidroxilase e a 17,20 liase mostraram atividades similares nos dois grupos. As pacientes hiperandrogênicas mostraram menor atividade da 21-hidroxilase, menor atividade da 11β-hidroxilase (p = 0,0001) e aumento estatisticamente significativo na atividade da 3β-hidroxiesteroide desidrogenase II (3β-HSDII) (p < 0.0001). Após a estimulação com tetracosactrin, apenas a atividade da 17,20 liase permaneceu regulada para cima na via Δ4 (p < 0.0001). CONCLUSÃO: As pacientes hiperandrogênicas apresentaram atividade mais alta da 17,20 liase, tanto no momento inicial quanto depois da estimulação adrenal. Maior conversão da desidroepiandrosterona (DHEA) em A com conversão normal da 17-OHPE em 17-OHP4 em pacientes hiperandrogênicas com SOP indica níveis diferentes de atividade da 3β-HSDII em células da adrenal, e pacientes hiperandrogênicas apresentaram menores atividades da 11β-hidroxilase e da 21-hidroxilase.

Correlation of glutathione S-transferase and DDT dehydrochlorinase activities with DDT susceptibility in Anopheles and Culex mosquitos from northern Thailand.

Comparative DDT-susceptibility status as well as glutathione S-transferase activity and DDTase activity of Anopheles minimus (A). An. annularis and Culex quinquefasciatus were investigated to ascertain the role of these enzymes in DDT-resistance. The standard WHO susceptibility test kits was used to discriminate between resistant and susceptible populations. GST activity was measured in microtiter plates whereas DDTase activity was determined by HPLC quantitation of DDT metabolites. This is the first report of DDT-resistance in the Thai malaria vector, An. minimus species A. A positive correlation of DDT-resistance and DDTase activity was observed in this species as well as in the suspected vector, An. annularis. However, GST activity was not correlated to DDT-resistance in either species. Statistical analysis and scatter plots demonstrated the non-correlation between DDTase and GST activity in An. annularis. Studies in Culex quinquefisciatus revealed difference in GST/ DDTase and the relationship to DDT-resistance compared to the anopheline species. The Culex GST activity is correlated to DDTase activity. These results suggested that a positive correlation of GST and DDTase activity might be species dependent.

A PMA degrading constitutive organomercurial lyase in a broad-spectrum mercury resistant Bacillus pasteurii strain DR2.

A broad-spectrum Hg-resistant strain of B. pasteurii DR2 utilized phenylmercuric acetate (PMA) as sole source of carbon. This bacterial strain contained a constitutive organomercurial lyase which specifically degraded PMA but not other organo-mercurials. This PMA-lyase activity was also stimulated to different extents when this bacterial strain was grown in presence of different organic compounds as sole source of carbon.