Functional Study Of LSD1in Mouse Embryo Fibroblast Cells Reprogramming

Fibroblasts can be reprogrammed to induced pluripotent stem (iPS) cells upon expression of Oct4, Sox2, Klf4and c-myc. The generation of iPS cells is a nuclear reprogramming process with alteration and rebuilding of chromatin structures. LSD1is a histone lysine specific demethylase and may play a critical role in cellular reprogramming as one of the chromatin regulators. This study is focused on the function of LSD1in the process of iPSCs generation. The expression levels of LSD1protein in MEFs and ES cells were compared at first. Then LSD1was overexpressed and knocked down in the iPSCs generation system. The inhibitor of LSD1was also used to explore the function of LSD1. Next, Co-immunoprecipitation analysis and quantitative PCR were carried out to find the mechanisms of LSD1function. Western blot analysis showed that the expression level of LSD1in ES cells was higher than in MEFs. Overexpression of LSD1in the reprogramming system had no effect on the generation of iPS cells. However, more iPS cell colonies were formed with LSD1knockdown by RNAi or with LSD1inhibitor tranylcypromine. Further experiments found that the inhibitor functioned at the early stage of cellular reprogramming. LSD1may regulate and be regulated by MET related factors to function in this process because the results showed the period of LSD1function was consistent with MET (Mesenchymal-to-Epithelia Transition). Co-immunoprecipitation analysis indicated that there are physical interaction between LSD1and Oct4/Nanog. The results of quantitative PCR showed LSD1regulated the downstream genes of Oct4. In all, these data suggests that LSD1regulates the generation of iPS cells via interaction with Oct4/Nanog.