Methylation of WTH3, a possible drug resistant gene, inhibits p53 regulated expression

<p>Abstract</p> <p>Background</p> <p>Previous results showed that over-expression of the <it>WTH3 </it>gene in MDR cells reduced <it>MDR1 </it>gene expression and converted their resistance to sensitivity to various anticancer drugs. In addition, the <it>WTH3 </it>gene promoter was hypermethylated in the MCF7/AdrR cell line and primary drug resistant breast cancer epithelial cells. <it>WTH3 </it>was also found to be directly targeted and up regulated by the <it>p53 </it>gene. Furthermore, over expression of the <it>WTH3 </it>gene promoted the apoptotic phenotype in various host cells.</p> <p>Methods</p> <p>To further confirm <it>WTH3</it>'s drug resistant related characteristics, we recently employed the small hairpin RNA (shRNA) strategy to knockdown its expression in HEK293 cells. In addition, since the <it>WTH3 </it>promoter's p53-binding site was located in a CpG island that was targeted by methylation, we were interested in testing the possible effect this epigenetic modification had on the <it>p53 </it>transcription factor relative to <it>WTH3 </it>expression. To do so, the <it>in vitro </it>methylation method was utilized to examine the <it>p53 </it>transgene's influence on either the methylated or non-methylated <it>WTH3 </it>promoter.</p> <p>Results</p> <p>The results generated from the gene knockdown strategy showed that reduction of <it>WTH3 </it>expression increased <it>MDR1 </it>expression and elevated resistance to Doxorubicin as compared to the original control cells. Data produced from the methylation studies demonstrated that DNA methylation adversely affected the positive impact of <it>p53 </it>on <it>WTH3 </it>promoter activity.</p> <p>Conclusion</p> <p>Taken together, our studies provided further evidence that <it>WTH3 </it>played an important role in MDR development and revealed one of its transcription regulatory mechanisms, DNA methylation, which antagonized <it>p53</it>'s positive impact on <it>WTH3 </it>expression.</p

Stereoselective multi-step reduction of dehydrocholic acid with hydroxysteroid dehydrogenases

Die zweistufige Reduktion von Dehydrocholsäure zu 12-Keto-Ursodesoxycholsäure ist der biokatalytische Schritt in der chemo-enzymatischen Synthese von Ursodesoxycholsäure. Hydroxysteroiddehydrogenasen exprimierende E. coli-Ganzzellbiokatalysatoren wurden für ein einfaches Eintopfverfahren entwickelt. Durch Modellierung wurde der Prozess optimiert. Im Litermaßstab konnten dadurch selbst bei geringen Biokatalysatormengen 28 g L-1 Dehydrocholsäure bei hohen Raum-Zeit-Ausbeuten von 120 g L-1 d-1 vollständig zum reinen chiralen Produkt umgewandelt werden.The two-step reduction of dehydrocholic acid to 12-keto-ursodeoxycholic acid is the biocatalytic step in the chemo-enzymatic synthesis of ursodeoxycholic acid. E. coli whole-cell biocatalysts expressing hydroxysteroid dehydrogenases were created to enable a simple biocatalytic one-pot process. The process was analyzed and optimized through modeling. When applying even low amounts of the whole cell biocatalyst on a liter-scale, 28 g L-1 dehydrocholic acid could be completely converted to the desired pure chiral product and high space-time yields of 120 g L-1 d-1 were achieved

A novel heterogeneous particle addition method based on laser cladding hybrid wire arc additive manufacturing: improvement performance of stainless steel components

The direct preparation of large components by laser cladding (LC) is not economical, but if it is used as a means of trace element addition and combined with the high efficiency advantages of wire arc additive manufacturing (WAAM), efficient manufacturing and high degree of freedom design of components can be achieved. In this study, high toughness and high plasticity stainless steel components were successfully prepared by combining WAAM and LC. The microstructure in the hybrid deposited specimens evolved into a single austenite, and the proportion of cubic and {001} textures increased. The mechanical properties obtained were significantly greater than those recommended for use by ASTM A479. The toughness of the hybrid deposited specimens was significantly improved, with a maximum increase in yield strength of 32%, exceeding 350 MPa. The elongation increased by a maximum of 12.7%, reaching 89.2% in the horizontal direction