Final Report Summary - SPLICE (Molecular mechanisms of transcriptional regulation of lymphocyte development by the E2A splice variants E12 and E47)
In this project, we transferred cellular models, such as pro B cell cultures and E12/E47 deficient mice, as well as different methods, such as E2A chromatin immunoprecipitations (ChIP), which had been established before, to the host lab in Germany. Using these tools, we identified the E2A splice variant specific DNA binding properties by using ChIP-Sequencing in combination with E12- and E47-deficient pro B cells. Currently, we further elucidate the specific gene regulatory functions of E12 and E47 during B cell development, illuminating the need for two highly similar proteins, E12 and E47, for B cell development. In order to identify novel regulators of E12 or E47 splicing, we generated fluorescence-based splicing reporter constructs. These studies will for the first time investigate E2A splicing and might also identify splicing regulators, which are important for the splicing of other mRNAs. Finally, using mass spectroscopy of E47 containing protein complexes, we identified various transcriptional cofactors, such as CoREST1 or Dnmt1, which interact with E47. Taken together, these studies will lead to a better understanding of the regulation of E12/E47 splicing and of the molecular mechanisms used by the two highly similar proteins E12 and E47 in regulating B cell development.