Wuxiang Mao

A highly conserved G-rich consensus sequence in hepatitis C virus core gene represents a new anti-hepatitis C target.

A conserved guanine-rich sequence could be a new target for anti–hepatitis C virus drug development. G-quadruplex (G4) is one of the most important secondary structures in nucleic acids. Until recently, G4 RNAs have not been reported in any ribovirus, such as the hepatitis C virus. Our bioinformatics analysis reveals highly conserved guanine-rich consensus sequences within the core gene of hepatitis C despite the high genetic variability of this ribovirus; we further show using various methods that such consensus sequences can fold into unimolecular G4 RNA structures, both in vitro and under physiological conditions. Furthermore, we provide direct evidences that small molecules specifically targeting G4 can stabilize this structure to reduce RNA replication and inhibit protein translation of intracellular hepatitis C. Ultimately, the stabilization of G4 RNA in the genome of hepatitis C represents a promising new strategy for anti–hepatitis C drug development.

Highly Selective Suppression of Melanoma Cells by Inducible DNA Cross-Linking Agents: Bis(catechol) Derivatives

A series of bis(catechol) quaternary ammonium derivatives were designed and synthesized. We investigated their ability to cross-link DNA induced by tyrosinase and found that the o-quinone is key intermediate in the process by using the nucleophile 3-methyl-2-benzothiazolinone hydrazone (MBTH) in the tyrosinase assay. Their cytotoxicities to B16F1, Hela, and CHO cells were tested by MTT assays. The specific and potent abilities to kill the tyrosinase-efficient melanoma cells kindled our interest in exploring the relationship between their abilities of cross-linking DNA and their selective cytotoxicities to cells. Through an integrated approach including intracellular imaging for detection of the dihydroxyphenyl groups, alkaline comet assays, and γ-H2AX immunofluorescence assays, the speculation was confirmed. The bis(catechol) quaternary ammonium derivatives showed notable cell selectivity because they displayed cytotoxicities after being oxidized by tyrosinase, and they were able to target the DNA efficiently in the tyrosinase-efficient melanoma cells, forming both alkylated and cross-linked species.

Fluorescent turn-on probes for the detection of fluoride ions in organic solvent and in cells

We have successfully developed two coumarin-based fluorescent probes to detect F−in vitro. Probe 1 is lipophilic for organic soluted detection. Probe 2 is an amphipathic molecule due to its TBDPS and triphenyl-phosphine groups for both mitochondrial localization and fluoride ion detection. Both of the two probes have good selectivity in F− detection; the detection limit of probe 2 was as low as 670 nM. Probe 2 has good fluorescence enhancement in response to F− in cell imaging. The probes are lowly cytotoxic to cells.

Highly Selective Detection of 5-Methylcytosine in Genomic DNA Based on Asymmetric PCR and Specific DNA Damaging Reagents

DNA methylation is a significant epigenetic modification of the genome that is involved in regulating many cellular processes. An increasing number of human diseases have been discovered to be associated with aberrant DNA methylation, and aberrant DNA methylation has been deemed to be a potential biomarker for diseases such as cancers. A safe, nontoxic, and sensitive method for accurate detection of 5-methylcytosine in genomic DNA is extremely useful for early diagnosis and therapy of cancers. In this paper, we established a novel system to detect 5-methylcytosine, which is based on bisulfite treatment, asymmetric PCR, and specific DNA damaging reagents. Our method could be used for identifying the loci of 5mC in genomic DNA and detecting the DNA methylation levels in tissues as well.

Specific recognition of guanines in non-duplex regions of nucleic acids with potassium tungstate and hydrogen peroxide

Structural features of nucleic acids have become an integral part of current biomedical research. Highly selective and readily performed methods with little toxicity that target guanosines in non-duplex nucleic acids are needed, which led us to search for an effective agent for guanosine sequencing. Treatment of DNA or RNA with potassium tungstate and hydrogen peroxide produced damaged guanosines in DNA or RNA sequences. The damaged guanosines in non-duplex DNA could be cleaved by hot piperidine. Similarly, damaged guanosines in non-duplex RNA could be cleaved by aniline acetate. We could identify structural features of nucleic acid using this strategy instead of dimethyl sulphate and Ribonuclease T1.

Application of Ammonium Persulfate for Selective Oxidation of Guanines for Nucleic Acid Sequencing

Nucleic acids can be sequenced by a chemical procedure that partially damages the nucleotide positions at their base repetition. Many methods have been reported for the selective recognition of guanine. The accurate identification of guanine in both single and double regions of DNA and RNA remains a challenging task. Herein, we present a new, non-toxic and simple method for the selective recognition of guanine in both DNA and RNA sequences via ammonium persulfate modification. This strategy can be further successfully applied to the detection of 5-methylcytosine by using PCR.