Repair of mitomycin C mono- and interstrand cross-linked DNA adducts by UvrABC: a new model

Mitomycin C induces both MC-mono-dG and cross-linked dG-adducts in vivo. Interstrand cross-linked (ICL) dG-MC-dG-DNA adducts can prevent strand separation. In Escherichia coli cells, UvrABC repairs ICL lesions that cause DNA bending. The mechanisms and consequences of NER of ICL dG-MC-dG lesions that do not induce DNA bending remain unclear. Using DNA fragments containing a MC-mono-dG or an ICL dG-MC-dG adduct, we found (i) UvrABC incises only at the strand containing MC-mono-dG adducts; (ii) UvrABC makes three types of incisions on an ICL dG-MC-dG adduct: type 1, a single 5′ incision on 1 strand and a 3′ incision on the other; type 2, dual incisions on 1 strand and a single incision on the other; and type 3, dual incisions on both strands; and (iii) the cutting kinetics of type 3 is significantly faster than type 1 and type 2, and all of 3 types of cutting result in producing DSB. We found that UvrA, UvrA + UvrB and UvrA + UvrB + UvrC bind to MC-modified DNA specifically, and we did not detect any UvrB- and UvrB + UvrC–DNA complexes. Our findings challenge the current UvrABC incision model. We propose that DSBs resulted from NER of ICL dG-MC-dG adducts contribute to MC antitumor activity and mutations

Increased viability of fibroblasts when pretreated with ceria nanoparticles during serum deprivation

Francielli S Genier,1 Maximilian Bizanek,1 Thomas J Webster,1,2 Amit K Roy1,2 1Department of Chemical Engineering, Northeastern University, Boston, MA, USA; 2Wenzhou Institute of Biomaterials and Engineering (WIBE), Wenzhou University, Wenzhou, People’s Republic of China Abstract: Conditions of cellular stress are often the cause of cell death or dysfunction. Sustained cell stress can lead to several health complications, such as extensive inflammatory responses, tumor growth, and necrosis. To prevent disease and protect human tissue during these conditions and to avoid medication side effects, nanomaterials with unique characteristics have been applied to biological systems. This paper introduces the pretreatment in human dermal fibroblasts with cerium oxide nanoparticles during nutritional stress. For this purpose, human dermal fibroblast cells received cell culture media with concentrations of 250 µg/mL and 500 µg/mL of nano-cerium oxide before being exposed to 24, 48, and 72 hours of serum starvation. Contrast images demonstrated higher cell confluence and cell integrity in cells pretreated with ceria nanoparticles compared to untreated cells. It was confirmed by MTS assay after 72 hours of serum starvation that higher cell viability was achieved with ceria nanoparticles. The results demonstrate the potential of cerium oxide nanoparticles as protective agents during cellular starvation. Keywords: cerium oxide, nanoparticles, serum starvation, human dermal fibroblast

In vitro characterization of MitE and MitB: Formation of N-acetylglucosaminyl-3-amino-5-hydroxybenzoyl-MmcB as a key intermediate in the biosynthesis of antitumor antibiotic mitomycins

Mitomycins, produced by several Streptomyces strains, are potent anticancer antibiotics that comprise an aziridine ring fused to a tricyclic mitosane core. Mitomycins have remarkable ability to crosslink DNA with high efficiency. Despite long clinical history of mitomycin C, the biosynthesis of mitomycins, especially mitosane core formation, remains unknown. Here, we report in vitro characterization of three proteins, MmcB (acyl carrier protein), MitE (acyl AMP ligase), and MitB (glycosyltransferase) involved in mitosane core formation. We show that 3-amino-5-hydroxybenzoic acid (AHBA) is first loaded onto MmcB by MitE at the expense of ATP. MitB then catalyzes glycosylation of AHBA-MmcB with uridine diphosphate-N-acetylglucosamine (UDP-GlcNAc) to generate a key intermediate, GlcNAc-AHBA-MmcB, which contains all carbon and nitrogen atoms of the mitosane core. These results provide important insight into mitomycin biosynthesis