Chemical and photochemical properties of chloroharmine derivatives in aqueous solutions.

Thermal and photochemical stability (Φ(R)), room temperature UV-vis absorption and fluorescence spectra, fluorescence quantum yields (Φ(F)) and lifetimes (τ(F)), quantum yields of hydrogen peroxide (Φ(H2O2)) and singlet oxygen (Φ(Δ)) production, and triplet lifetimes (τ(T)) have been obtained for the neutral and protonated forms of 6-chloroharmine, 8-chloroharmine and 6,8-dichloroharmine, in aqueous media. When it was possible, the effect of pH and oxygen concentration was evaluated. The nature of electronic transitions of protonated and neutral species of the three investigated chloroharmines was established using Time-Dependent Density Functional Theory (TD-DFT) calculations. The impact of all the foregoing observations on the biological role of the studied compounds is discussed.

Oxidatively generated base modifications in DNA: Not only carcinogenic risk factor but also regulatory mark?

The generation of DNA modifications in cells is in most cases accidental and associated with detrimental consequences such as increased mutation rates and an elevated risk of malignant transformation. Accordingly, repair enzymes involved in the removal of the modifications have primarily a protective function. Among the well-established exceptions of this rule are 5-methylcytosine and uracil, which are generated in DNA enzymatically under controlled conditions and fulfill important regulatory functions in DNA as epigenetic marks and in antibody diversification, respectively. More recently, considerable evidence has been obtained that also 8-oxo-7,8-dihydroguanine (8-oxoG), a frequent pro-mutagenic DNA modification generated by endogenous or exogenous reactive oxygen species (ROS), has distinct roles in the regulation of both transcription and signal transduction. Thus, the activation of transcription by the estrogen receptor, NF-κB, MYC and other transcription factors was shown to depend on the presence of 8-oxoG in the promoter regions and its recognition by the DNA repair glycosylase OGG1. The lysine-specific histone demethylase LSD1, which produces H2O2 as a by-product, was indentified as a local generator of 8-oxoG in some of these cases. In addition, a complex of OGG1 with the excised free substrate base was demonstrated to act as a guanine nucleotide exchange factor (GEF) for small GTPases such as Ras, Rac and Rho, thus stimulating signal transduction. The various findings and intriguing novel mechanisms suggested will be described and compared in this review.

Role of the DNA repair glycosylase OGG1 in the activation of murine splenocytes.

OGG1 (8-oxoguanine-DNA glycosylase) is the major DNA repair glycosylase removing the premutagenic DNA base modification 8-oxo-7,8-dihydroguanine (8-oxoG) from the genome of mammalian cells. In addition, there is accumulating evidence that OGG1 and its substrate 8-oxoG might function in the regulation of certain genes, which could account for an attenuated immune response observed in Ogg1-/- mice in several settings. Indications for at least two different mechanisms have been obtained. Thus, OGG1 could either act as an ancillary transcription factor cooperating with the lysine-specific demethylase LSD1 or as an activator of small GTPases. Here, we analysed the activation by lipopolysaccaride (LPS) of primary splenocytes obtained from two different Ogg1-/- mouse strains. We found that the induction of TNF-α expression was reduced in splenocytes (in particular macrophages) of both Ogg1-/- strains. Notably, an inhibitor of LSD1, OG-L002, reduced the induction of TNF-α mRNA in splenocytes from wild-type mice to the level observed in splenocytes from Ogg1-/- mice and had no influence in the latter cells. In contrast, inhibitors of the MAP kinases p38 and JNK as well as the antioxidant N-acetylcysteine attenuated the LPS-stimulated TNF-α expression both in the absence and presence of OGG1. The free base 8-oxo-7,8-dihydroguanine had no influence on the TNF-α expression in the splenocytes. The data demonstrate that OGG1 plays a role in an LSD1-dependent pathway of LPS-induced macrophage activation in mice.

The repair of oxidized purines in the DNA of human lymphocytes requires an activation involving NF-YA-mediated upregulation of OGG1.

8-Oxoguanine DNA glycosylase (OGG1), which initiates the repair of DNA purine modifications such as 8-oxo-7,8-dihydroguanine (8-oxoG), is often regarded as a house keeping protein ubiquitously active in mammalian cells. We have analysed the repair rates of oxidized purines generated by photosensitization in peripheral human lymphocytes and observed that the cells were virtually unable to remove these lesions (less than 10% removal within 24h). However, stimulation of the lymphocytes with phytohemagglutinin (PHA) strongly accelerated the repair so that ∼30% of the lesions were repaired within 4h. Within 24h following PHA stimulation and preceding the induction of cell proliferation, Western blots revealed an approximately 4-fold up-regulation of OGG1. The levels of OGG1 mRNA were 4-fold increased already after 6h. Chromatin immunoprecipitation analysis indicated that the up-regulation of OGG1 was associated with increased binding of the transcription factor NF-YA to the promoter of the OGG1 gene. The binding of NF-YA and subsequent induction of OGG1 was inhibited in the presence of an inhibitor of Jun kinase, indicating an activation of the corresponding signalling pathway as the mechanism underlying this transcriptional up-regulation. Our results reveal a strict control of base excision repair in cells of the human immune system.