Study On The Effect Of Nspc1 On Proliferation And Differentiation Of Neural Stem Cells In Rats

Polycomb Group(PcG) protein was firstly found in Drosophila which is conserved from C. elegans to human. It plays an important role during the progress of embryonic development, cell cycle, tumor genesis and stem cell self-renew. NSPc1 (Nervous System Polycomb 1, also named as Polycomb group RING finger 1, PCGF1) is a novel member of PcG, and it shares high homology with Bmi1 and Mel18. Previous work on this gene showed that NSPc1 has the classic transcriptional repression activity and also affects cell proliferation.It was popular that study the function of PcG in stem cells these years. Since NSPc1 is highly expressed in mouse nervous system, we have addressed ourselves to the function of NSPc1 during development. We first established mouse neural stem cell(mNSC) primary culture, proliferation, differentiation, electroporation and NCFC assay platform. This is the fundamental work for the later research.Based on this, we made the lentiviral expression construct for either over-expression or knockdown NSPc1. Then we packaged viral particles and concentrated by ultracentrifugation. The final titer of virus could be up to 2×10~7TU/ml.We overexpressed and knocked down NSPc1 by electroperation in mNSC, respectively. Then we observed the phenotype of the transfected mNSC when they were exposured to proliferation or differentiation condition. We found knocking down NSPc1 can inhibit mNSC's proliferation while overexpression of NSPc1 can maintain or even increase the inNSC's activity. We also noticed that the mNSC have much tendency to differentiate into neurons when we knocked down NSPc1. According to the timing of neuron genesis in mouse's brain, we speculate NSPc1 could play essential role in mouse embryonic brain development.In conclusion, we have established mNSC culture as well as lentivirus package and concentration methodology. We also investigated the phenotype of NSPc1 using gain of function and loss of function methods in mNSC which would be the clue for our following research of this gene.