Differentiation of rhabdomyosarcoma cells by the release of paused RNA polymerase

Project Summary After RNA polymerase II (Pol II) and general transcription factors are assembled into the preinitiation complex at a promoter, it synthesizes a short mRNA of 30-50 bases and then pauses (promoter-proximal pausing or PPP). The paused Pol II is released when specific transcription factors and chromatin proteins are recruited to the promoter. PPP is observed in 40-75% of the entire gene population at a time in many types of cells but its physiological functions and cancer pathogenesis remain largely elusive. The principal investigator?s group found that release of Pol II from PPP by depleting genes responsible for PPP promoted differentiation of mouse myoblasts. On the other hand, one of the pathogenic mechanisms of rhabdomyosarcoma is blockage of differentiation. The principal investigator?s group hypothesized that release of Pol II from PPP in rhabdomyosarcoma will remove the blockage and induce differentiation of the cells. To test this hypothesis, they proposed the following two aims. In Aim 1, they will deplete two components responsible for PPP in rhabdomyosarcoma cells and study whether it will induce differentiation in vitro and in vivo. PCR, immunofluorescence staining and RNA-seq will be used to assess the level of differentiation. In Aim 2, they will investigate with ChIP-seq how Pol II and histone methylations are redistributed by the depletion. These studies are expected to unravel a novel pathogenic mechanism and contribute to the treatment of rhabdomyosarcoma.