The effect of phenobarbital on the methylation level of the p16 promoter region in rat liver.

It has been suggested that non-genotoxic carcinogens (NGCs) may cause modification of the DNA methylation status. We studied the effects of phenobarbital (PB) -- a non-genotoxic rodent liver carcinogen -- on the methylation level of the promoter region of the p16 suppressor gene, as well as on hepatomegaly, DNA synthesis, and DNA-methyltransferase (DNMTs) activity in the rat liver. Male Wistar rats received PB in 1, 3 or 14 daily oral doses (at 24-h intervals), each equivalent to 1/10 of the LD(50) value. The study showed that PB has caused persistent elevation in relative liver weight (RLW) as well as a transient increase in DNA synthesis. This suggests that the PB-induced increase in RLW was due to a combination of both hyperplasia and hypertrophy of liver cells. The effect of PB on DNA synthesis corresponded to an increase in the methylation pattern of the p16 promoter sequence. Methylation of cytosine in the analyzed CpG sites of the p16 gene was found after short exposure of the animals to PB. Treatment of rats with PB for 1 and 3 days also produced an increase in nuclear DNMTs activity. After prolonged administration (14 days), DNA synthesis declined, returning to the control level. No changes in methylation of the p16 gene nor in DNMTs activity were observed. The reversibility of early induced changes in target tissues is a mark characteristic of tumor promoters. Thus, transient changes in methylation of the p16 gene, although their direct role in the mechanisms of PB toxicity, including its carcinogenic action, remains doubtful, may therefore be a significant element of such processes.

[DNA methylation--alternative mechanism of chemical carcinogenesis]. / Metylacja DNA--alternatywny mechanizm chemicznej kancerogenezy.

It is increasingly accepted that the initiation of chemical carcinogenesis should be considered as a transformation process of a normal cell caused by genetic changes, i.e. mutational DNA damage and/or epigenetic changes, which render normal gene expression impossible with the preservation of the DNA sequence intact. Epigenetic DNA methylation changes have been among the most extensively investigated processes in the recent years. Many types of cancer cells have been found to exhibit an increased or reduced level of CpG sequence methylation in promoter regions of genes, especially the genes whose protein products take part in the control of cell cycle regulation. In view of the fact that DNA methylation is thought to play a role in gene expression, its abnormal level in the genes that encode proteins participating in the control of the cell cycle and apoptosis regulation may disturb cell homeostasis resulting in pathologies that may, in turn, lead to neoplastic transformation. Although changes in the mechanisms of transcriptional activity caused by methylation of cytosine in CpG sequences have not been completely elucidated, it has been determined that this relationship is associated with a decreased level of histone acetylation, which induced a more densely-packaged chromatin structures in the methylated regions of chromosomal DNA. A large proportion of chemical carcinogens consists of chemicals whose carcinogenic activity is not related to direct damage of the genetic material. The changes in DNA methylation are being considered as one mechanism of action for non-genotoxic carcinogens (NGCs). The significance of non-genotoxic agents in the development of the carcinogenesis process indicate that their mechanisms of the action should be investigated in terms of health hazards they pose to humans exposed to this type of environmental pollutants.

Tetra-amino-acid tandem repeats are involved in HsdS complementation in type IC restriction-modification systems.

All known type I restriction and modification (R-M) systems of Escherichia coli and Salmonella enterica belong to one of four discrete families: type IA, IB, IC or ID. The classification of type I systems from a wide range of other genera is mainly based on complementation and molecular evidence derived from the comparison of the amino acid similarity of the corresponding subunits. This affiliation was seldom based on the strictest requirement for membership of a family, which depends on relatedness as demonstrated by complementation tests. This paper presents data indicating that the type I NgoAV R-M system from Neisseria gonorrhoeae, despite the very high identity of HsdM and HsdR subunits with members of the type IC family, does not show complementation with E. coli type IC R-M systems. Sequence analysis of the HsdS subunit of several different potential type IC R-M systems shows that the presence of different tetra-amino-acid sequence repeats, e.g. TAEL, LEAT, SEAL, TSEL, is characteristic for type IC R-M systems encoded by distantly related bacteria. The other regions of the HsdS subunits potentially responsible for subunit interaction are also different between a group of distantly related bacteria, but show high similarity within these bacteria. Complementation between the NgoAV R-M system and members of the EcoR124 R-M family can be restored by changing the tetra-amino-acid repeat within the HsdS subunit. The authors propose that the type IC family of R-M systems could consist of several complementation subgroups whose specificity would depend on differences in the conserved regions of the HsdS polypeptide.