Comparison Of H3K4Trimethylation Patterns Preimplantation Embryos And Analysis Of Methyltransferase Promoter Of Mouse

The trimethylation on lysine4of histone3(H3K4me3) is considered as a marker ofactive transcription, and it plays important roles in the transcription reprogrammingefficiency. In this study, In order to investigate the impact of the patterns of H3K4me3ofpreimplantation embryos in mouse by the vitro operation and culture environment. Whichreveals the problem of low efficiency of nuclear transfer reprogramming.In this study, thelevels of H3K4me3in MII stage oocytes, in vivo and in vitro fertilization (IVF) mousepreimplantation embryos were examined using immunofluorescence histochemistry. TheIVF embryos were further treated by Trichostatin A (TSA, a histione deacetylase inhibitor)to investigate the effect of histone acetylation on H3K4me3. We found that the levels ofH3K4me3in MII stage oocytes are intensive on metaphase chromosomes. The patterns ofH3K4trimethylation of in vivo embryos from zygotes to blastocysts stages are similar tothose of IVF embryos, it Immunofluorescence intensity decreased, In4-cell of in vivo andIVF embryos, only very weak signal was detected, the levels of H3K4me3were increased.At the pronuclear stage, showing an asymmetric pattern in these two kinds of embryos.This asymmetric methylation of H3K4me3was maintained until the2-cellstage. yet themajor difference is that the intensity of H3K4me3is higher in in vivo embryos. Moreover,the levels of H3K4me3in TSA-treated groups were increased compared to the IVFembryos. Our results suggested that the culture condition and environmental change mightbe one of the most significant reasons to cause the alteration of histone modification. Wemust pay more attention to the functions of maternal hormones, local factors on theepigenetic modification in the future studies. In the this study, we cloned the Mll2、Mll4、Mll5gene promoter of mouse and constructed Mll2、Mll4and Mll5promoter5’deletionfragments(-pGL3),the transformation of Nanog-pcDNA3.1， Sox2-pcDNA3.1，Oct4-pcDNA3.1，c-Myc-pcDNA3.1，Klf4-pcDNA3.1to analysis the activity of themethyltransferase promoter in HEK293cells Mll2、Mll4and Mll5promoter5’deletionfragments.The results show that: through the transformation of HEK293cell by Mll2、Mll4and Mll5promoter5’deletion fragments(-pGL3) separately, we found that the potentialactive region of the Mll2promoter may be located upstream away from the transcriptionstart site about-1679bp;the potential active region of the Mll4promoter may be locatedupstream away from the transcription start site about-1575bp~-1852bp;the potential activeregion of the Mll5promoter may be located upstream away from the transcription start site about-1666bp~-1958bp.By the co-transfection of promoter and transcription factors,Wefound that Sox2，Oct4have certain advancement to Mll2、Mll4and Mll5promoter activity.