The intranuclear disposition of plasmid DNA is extremely important for transgene expression. Exogenous histones have been used as carriers of plasmid DNA in histone-mediated gene delivery. In this study, the effects of exogenous histone H3 complexed with plasmid DNA on transgene expression efficiency were examined. The plasmid−histone complexes in various ratios were transfected into HeLa cells by osmotic pressure.Histone H3 suppressed transgene expression in the nucleus in a dose-dependent manner. Our results suggest that the histone-mediated gene delivery is unlikely to be useful, from the viewpoint of the intranuclear disposition.

Running title: Effects of non-B DNA sequences on transgene expression DNA conformation may be an important factor affecting gene transcription.In this study, we examined how DNA sequences with unusual conformations affect transgene expression. A 30 and (CG) 15 sequences that can adopt the B' and Z conformations, respectively, were introduced into a β-actin promoter. Luciferase plasmids containing the manipulated promoter were transfected into NIH3T3 cells by electroporation and were delivered into mouse livers with a hydrodynamics-based injection. Expression from plasmid with the (CG) 15 sequence was multiple times higher than expression from control plasmid DNA. The A 30 sequence also tended to enhance expression. These results suggest that non-B DNA sequences could improve transgene expression in cells.

Plasmid DNA was chromatinized with core histones (H2A, H2B, H3, and H4) in vitro and was delivered into mouse liver by hydrodynamics-based administration. Transgene expression from the chromatinized plasmid DNA was more efficient than that from plasmid DNA delivered in the naked form. The use of acetylation-enriched histones isolated from cells treated with a histone deacetylase inhibitor (trichostatin A) seemed to be more effective. These results indicated that chromatinized plasmid DNA is useful for efficient transgene expression in vivo.