(486c) Deep Profiling of Heterochromatin Associated Proteins and Their Role in Liver Cell Reprogramming
In order to screen for proteins that are responsible for the formation of heterochromatin, we treated fibroblast cells with siRNAs targeting each of the candidate protein, and examined the change in expression level of key hepatic genes such as DSC2, NR1H4, and CRP. Using RNA-seq, we identified 90 proteins that led to overexpression of the hepatic genes. To further understand how each candidate protein affects the formation of heterochromatin, we stained the siRNA treated cells with H3K9me3 antibody which works as a marker for heterochromatin domain and analyzed the change in heterochromatin structure in a single cell resolution. We examined how diffuse and moderately compacted heterochromatin region and highly compacted focal region of heterochromatin are affected upon different siRNA treatment. Some siRNA-treated cells exhibited both fewer densely packed and moderately packed chromatin regions. Some other siRNA-treated cells showed changes in either focal heterochromatin regions or diffuse regions, but not both. Our results imply that heterochromatin proteins regulate heterochromatin formation and subsequent gene silencing in a differential manner. This study will provide better understanding on the mechanism of gene silencing via heterochromatin formation, as well as to suggest better stem cell reprogramming strategies.