The Molecular Mechanism Of Recruiting Polycomb Repressive Complex1 To Endogenous Retrovirus MLV

Endogenous Retroviruses(ERVs)are retroviral sequences integrated in the host genome,which belong to a subset of retrotransposable elements.The remaining transcriptional and retrotransposable activities of ERVs in embryonic stem cells and germ cells strongly threaten the stability and fidelity of the host genome.For self protection,higher eukaryotes have evolved a number of defensive mechanisms to repress ERVs,mainly including directly introducing mutations into ERVs and epigenetically repressing transcription of ERVs.Polycomb Group proteins(PcG)are transcriptional regulators that regulate gene expression through a series of modifications of chromatin.PcG proteins mainly function in two distinct large protein complexes,namely Polycomb Repressive Complex 1 and 2(PRC1 and PRC2).A recent report discovered that PRC1 and PRC2 acted redundantly in transcription repression of ERVs,while the recruiting mechanism of PRC1 in this process remains unclear.My thesis mainly focused on studying the recruiting mechanisms of PRC1 to a type of ERV,Murine Leukemia Virus(MLV).Unique sequence features of each individual MLV copy were characterized and the capability of various MLV copies in recruiting PRC1 was analyzed and compared.Our data confirm that the involvement of PRC1 in silencing MLV and the recruiting signals are located within MLV sequences.Methods:(1)Identify all the MLV copies within the genome of C57BL/6 mice by BLAT,and collect the information of viral sequences and incorporation site about each MLV copy;(2)Perform sequence comparison of each MLV copy using DNA STAR and further category particular MLV individuals with large deletions based on the positions of deletions;(3)Generate and verify a series stable cell lines inducibly expressing a core component of PRC1,CBX7,in mouse teratoma cells and embryonic stem cell lines with derived from various mouse strains;(4)Genotyping MLV individual copies in stable cells derived from different genetic background;(5)Characterize the enrichment of PRC1 on different MLV copies in various stable cell lines by ChIP-qPCR assays;(6)Analyze the correlation between MLV sequences and PRC1 recruitment.Results:(1)73 MLV copies were identified in the C57BL/6 mouse genome;(2)34 out of 73 MLV copies having large deletions in the C57BL/6 mouse genome were further classified into several categories;(3)Series stable cell lines expressing CBX7-Venus were generated and verified;(4)A great number of MLV individual copies were genotyped in stable cell lines with distinct genetics background;(5,6)ChIP-qPCR data confirmed that the recruiting signals were inside of the MLV sequences and implied that CBX7/PRC1 was selectively targeted to MLV copies with large deletions.