Crosstalk between G-Quadruplexes and Dnmt3a-Mediated Methylation of the c-MYC Oncogene Promoter.

The methylation of cytosines at CpG sites in DNA, carried out de novo by DNA methyltransferase Dnmt3a, is a basic epigenetic modification involved in gene regulation and genome stability. Aberrant CpG methylation in gene promoters leads to oncogenesis. In oncogene promoters, CpG sites often colocalize with guanine-rich sequences capable of folding into G-quadruplexes (G4s). Our in vitro study aimed to investigate how parallel G4s formed by a sequence derived from the c-MYC oncogene promoter region affect the activity of the Dnmt3a catalytic domain (Dnmt3a-CD). For this purpose, we designed synthetic oligonucleotide constructs: a c-MYC G4-forming oligonucleotide and linear double-stranded DNA containing an embedded stable extrahelical c-MYC G4. The topology and thermal stability of G4 structures in these DNA models were analyzed using physicochemical techniques. We showed that Dnmt3a-CD specifically binds to an oligonucleotide containing c-MYC G4, resulting in inhibition of its methylation activity. c-MYC G4 formation in a double-stranded context significantly reduces Dnmt3a-CD-induced methylation of a CpG site located in close proximity to the quadruplex structure; this effect depends on the distance between the non-canonical structure and the specific CpG site. One would expect DNA hypomethylation near the G4 structure, while regions distant from this non-canonical form would maintain a regular pattern of high methylation levels. We hypothesize that the G4 structure sequesters the Dnmt3a-CD and impedes its proper binding to B-DNA, resulting in hypomethylation and activation of c-MYC transcription.

Role of Dietary Inclusion of Phytobiotics and Mineral Adsorbent Combination on Dairy Cows' Milk Production, Nutrient Digestibility, Nitrogen Utilization, and Biochemical Parameters.

Our research purpose was to study the effect of the inclusion of a combination of phytobiotics in the form of dry Fucus vesiculosus grits (FG) and a mineral adsorbent from the heat-treated mineral shungite (TMS) on milk productivity, nutrient digestibility, and biochemical parameters of the Suksun dairy cows. A total of 80 dry-hardy cows of the Suksun breed were divided into four groups (20 heads each), balanced primarily by breed, age, body weight, body condition score, and indicators of milk yield for the previous lactation. The selected cows were with an average live body weight of 512.0 ± 1.28 kg, BCS 3.0-3.5, and parities of 6250 kg milk. The control group (CON) were fed the basic ration only; the second (TMS), third (FG), and fourth (TMS + FG) groups were fed the basic ration provided by 50 g of the mineral adsorbent from heat-treated shungite, 100 g of Fucus grits (Fucus vesiculosus), 50 g of the mineral adsorbent from heat-treated shungite, and 100 g of dry grits from Fucus vesiculosus, respectively. The total protein content in milk was significantly higher in the group receiving Fucus vesiculosus by 0.05% and the group receiving a combination of mineral adsorbent and Fucus vesiculosus by 0.03%. The percentage of milk fat content recorded the highest significant value in (TMS) group when compared to the control and represented (4.37 vs. 3.95). The group of cows that received (TMS + FG) revealed a significant difference in the digestibility of both ether extract and crude fiber when compared to the control group and represented (54.74 vs. 51.71 and 60.68 vs. 55.15%), respectively. The cows supplemented with a mineral adsorbent or a combination of mineral adsorbent and Fucus vesiculosus revealed a significant difference in the digestibility of ether extract and crude fiber in the group receiving TMS + FG by 3.0% (p < 0.05) and 5.5% (p < 0.05), respectively. The intake of nitrogen with the diet increased in (FG) and (TMS + FG) groups by 11.3 g (p < 0.05) and 13.4 g (p < 0.05) of nitrogen. There was an increase (p < 0.05) in the concentration of rumen ammonia in the control group compared to the other groups. The glucose content of those cows that received FG and TMS + FG combination increased (p < 0.05) by 0.76 mmol/L and 0.90 mmol/l in relation to the control group. The globulin, albumin/globulin ratio, and the level of triglycerides revealed a significant difference between the different experimental groups. In brief, the inclusion of a combination of phytobiotics in the form of dry Fucus vesiculosus grits and a mineral adsorbent from the heat-treated mineral shungite in Suksun dairy cows' diets improved milk composition, digestibility of nutrients, utilization of nitrogen, and did not cause deleterious effects on blood biochemical indicators.