Super-silencers regulated by chromatin interactions control apoptotic genes

Human silencers have been shown to exist and regulate developmental gene expression. However, the functional importance of human silencers needs to be elucidated such as the working mechanism and whether they can form "super-silencers". Here, through interrogating two putative silencer components of FGF18 gene, we found that two silencers can cooperate via compensated chromatin interactions to form a "super-silencer". Furthermore, double knock-out of two silencers exhibited synergistic upregulation of FGF18 expression and changes of cell identity. To disturb the "super-silencers", we applied combinational treatment of an EZH2 inhibitor GSK343, and a REST inhibitor, X5050 ("GR"). We found that GR led to severe loss of TADs and loops, while the use of just one inhibitor by itself only showed mild changes. Such changes of TADs and loops may due to reduced CTCF protein level observed upon GR treatment. Moreover, GSK343 and X5050 worked together synergistically to upregulate the apoptotic genes controlled by super-silencers, and thus gave rise to antitumor effects including apoptosis, cell cycle arrest and tumor growth inhibition. Overall, our data demonstrated the first example of a "super-silencer" and showed that combinational usage of GSK343 and X5050 could potentially lead to cancer ablation through disruption of "super-silencers".