Acute promyelocytic leukemia (APL) is associated with reciprocal translocations that always involve the RAR locus on chromosome 17, which variably translocates and fuses to the PML, PLZF, NPM1 (for brevity referred to as X genes/proteins). We originally hypothesized that X-RAR and RAR-X would act to interfere with both the transcriptional function of RARand the biological function of X proteins, and tat X proteins would thus play a key role in leukemogenesis and oncogenesis. Fundamental to this proposal, we also hypothesized, that the function of the X proteins of APL may be perturbed or lost in malignancies other than APL, and that the X proteins could share common activities and biological roles, as APL is ultimately the invariable outcome of their perturbed functions. On thisbasis, we proposed to study the various X genes comparatively and systematically in their physiologic and developmental roles as well as in the pathogenesis of human cancer through a direct genetic approach in the mouse as well as in human primary cancers of various histology. With this approach we have already accrued a wealth of information through the analysis of complete knock-out (KO) mouse mutant strains and primary cells/tissues from these mutants and human cancers. This analysis led to fundamental groundbreaking discoveries and strongly supported the hypothesis that X genes are not only critical cancer genes, but in fact tumor suppressor genes (TSGs) whose function is broadly perturbed in human cancer, well beyond the pathogenesis of APL. These discoveries have established me as an outstanding investigator in the field of TSG biology and regulation, and have been recognized at multiple levels with more than 20 national and international awards, numerous invited Keynote lectures and other accolades. We now propose to continue to study these critical genes as paradigms for tumor suppression, through the development of a second generation of models and tools in order to explore how they function in leukemia and other cancers, and, importantly, to develop and test new cancer therapies. To this end, we will adopt three main strategies: (1) To generate a second generation of conditional-tissue specific, cancer relevant, mutants for each of the genes in question to address cell specific and non-adaptive functions of these TSGs; (2) To characterize new regulatory mechanisms of these TSGs, including the non-coding RNAs dimension, to develop a set of criteria and novel concepts that defines how TSG function can be perturbed towards cancer initiation and progression; and (3) To develop and test novel therapeutic approaches that target TSG deficiency, targeting key pathways as well as signaling and regulatory nodes identified throughout our studies.