Correct regulation of growth and proliferation is essential for normal development and, when disrupted, can lead to a number of diseases such as cancer. The interest of our laboratory is to elucidate growth control pathways by using a combination of biochemical, cellular and genetic approaches. Our research focuses on the molecular function of the mammalian DREAM complex and its target genes.
Human DREAM (LINC) complex
We have recently identified a novel multi-protein complex in human cells that is related to similar complexes in invertebrates. The human complex consists of a five-protein core module consisting of RbAp48, LIN9, LIN37, LIN52 and LIN54. The core module interacts with the retinoblastoma protein p130 and transcription factors E2F4/DP1 and B-MYB in a cell cycle dependent manner. Specifically, DREAM associates with p130 and E2F4/DP1 in quiescent cells and represses target gene transcription. In contrast, in late S-phase, the interaction of DREAM with p130/E2F4/DP1 is lost and DREAM now binds to the B-MYB transcription factor resulting in the transcriptional activation of target genes. Genome wide expression studies revealed that DREAM-B-MYB is required for activation of mitotic genes. Target genes of DREAM encode for proteins that play important roles in entry into mitosis, spindle assembly and cytokinesis, such as Plk1, Cyclin B1 and Kif20a. Consequently, cells depleted of DREAM display major defects in mitosis and cytokinesis. We have found that the complete loss of the DREAM core subunit LIN9 in conditional mouse embryonic fibroblast results in mitotic segregation defects, cytokinesis failure and senescence, whereas the partial inactivation of LIN9 contributes to tumorigenesis. This indicates that the activity of DREAM has to be tightly controlled to prevent cell lethality and genomic instability. We have also recently shown that activity of DREAM is regulated by the p53 tumor suppressor pathway. Specifically, the p53 > p21 pathway switches DREAM from a gene activator (bound to B-MYB) to a gene repressor (bound to p130/E2F4). Importantly, B-MYB fails to dissociate from DREAM in p53-mutant cells and this contributes to increased expression of mitotic genes in these cells. These data suggest that DREAM-B-MYB functions downstream of mutant p53 in tumor cells. Current studies are focused on the role of human DREAM in cell cycle regulation, differentiation and tumorigenesis.
E2F is a family of eight transcription factors that play essential roles in the regulation of cellular proliferation. E2F-responsive elements have been identified in genes that play key roles in cell cycle progression, synthesis of nucleotides, DNA replication and apoptosis. Deregulation of E2F can promote tumorigenic transformation. E2F activity is regulated by the binding to pRB, the product of the retinoblastoma tumor suppressor protein, and by binding to two related "pocket proteins", p107 and p130. E2F6 is a retinoblastoma independent transcriptional repressor. We have generated and analyzed E2f6 mutant mice and found that they display homeotic transformations of the axial skeleton that are similar to the skeletal transformations observed in polycomb mutant mice. This observation is compatible with the finding that E2F6 associates with polycomb proteins. Using cDNA micro array experiments we found that E2F6 is required to silence meiotic genes in somatic tissues. ChIP (chromatin immunoprecipitation assays) indicate that E2F6 binds to the promoters of meiotic genes through a conserved binding site. Transcriptional repression of these genes by E2F6 involves methylation of histone H3 on lysine 9 and lysine 27. We are currently analyzing the molecular mechanisms of E2F6-mediated gene repression in more detail.