Functional structure analyses of the DNA-alkaloid complex promoting myogenic differentiation
Sayaka Shinji, Koji Umezawa, Takeshi Shimosato, Tomohide Takaya.
Graduate School of Science and Technology, Shinshu University, Nagano, Japan.
2018 International Symposium on Animal Science and Technology (Ina, Japan), 2018/08/10 (Poster).
Abstract
Summary: We identified that a telomeric DNA aptamer, myoDN, potently promotes differentiation of myogenic precursor cells into myotubes. The activity of myoDN is markedly enhanced by forming complex with berberine (Ber), an alkaloid from Phellodendron amurense. myoDN-Ber complex will be useful to promote skeletal muscle development for meat production. In this study, we investigated how myoDN interacts with Ber through analyses of myoDN mutants and Ber derivatives.
Materials & Methods: The mixtures of myoDN mutants and Ber derivatives (berberine, palmatine, coptisine, and jatrorrhizine) were subjected to agarose gel electrophoresis and ethidium bromide staining. Since Ber derivatives strongly fluoresce yellow under ultra violet illumination, the myoDN-bound derivative can be detected as an yellow band at the same position to that of myoDN alone. To examine the effects of cations on myoDN-Ber complex formation, myoDN and Ber were mixed in HCl, NaCl, MgCl2, KCl, CaCl2, MnCl2, FeSO4, CuSO4, or ZnSO4 solution.
Results & Discussion: Coptisine as well as Ber tightly formed complex with myoDN and enhanced its activity. Palmatine weakly bound to myoDN but jatrorrhizine did not at all. Mutant analyses of myoDN showed that the 5th thymidine and 15th guanine are independently required for interaction with Ber. myoDN-Ber complex was formed strongly in CaCl2 and weakly in MgCl2 solution but not detected in other cationic solutions. These results provide useful information to understand the functional structure of myoDN-Ber complex.