The structural basis of transcription factor 3C recruitment by N-myc

点击翻译按钮获取中文摘要

The uncontrolled proliferation that is characteristic of cancer cells is driven by the overexpression or aberrant regulation of proteins whose normal physiological role is to drive cellular growth in response to growth signals. One such protein is the transcription factor N-myc, one member of a family of three closely related proteins (c-myc, N-myc, and L-myc) all of which alter the expression of many genes. Overexpression of myc proteins leads to increased levels of cellular proliferation and is a frequent feature of cancer cells. Myc induces senescent cells to re-enter the cell cycle, subverts key checkpoints in the cell cycle, and facilitates the rewiring of cellular metabolism to favour both cell cycle progression and biomass accumulation. N-myc was recently found to increase gene expression via an interaction with a general transcription factor complex, TFIIIC. These molecules acting together were found to activate the production of hundreds of genes. This activated set of genes disproportionally involved in functions of N-myc known to drive cancer; functions such as cell cycle progression, DNA replication, nucleotide metabolism, and telomere biology. However, it is currently unknown how N-myc and TFIIIC complex recognise each other, and how their interaction impacts on the many other proteins they interact with. The first aim of our research proposal is to characterise the molecular basis of the N-myc:TFIIIC interaction at the level of the individual amino acids in the proteins. This will be done using orthogonal methods of X-ray crystallography, cryo-electron microscopy, and cross-linking mass spectrometry. The second aim is to validate the structure in vitro and in cells by using mutations at the binding interface. The final aim is to use these mutations to begin to understand what other proteins form part of the N-myc:TFIIIC complex and to begin to understand the mechanism by which N-myc interacts with TFIIIC over other molecules which are known to bind the same, or nearby, myc sequences. A potential benefit of this research will be to help validate transcriptional activation of myc as a target for cancer drug discovery. Drug discovery against Myc has proven very difficult and we believe the problem can be solved by targeted the protein-protein interactions that underpin Myc's oncogenic activity. However, it is not clear which these are, despite many years of research. Mutations which are validated in this work can be by us and others to determine the effect of disrupting the N-myc:TFIIC interaction on cancer cell proliferation, checkpoint integrity and cancer cell metabolism. If disruption of this interaction has significant effect on cancer cell proliferation it validates this interaction as a potential approach to develop new therapeutics that target Myc and this would be a big development in the field. Moreover, an additional benefit of our research is that structures and structural knowledge generated by this work will be available as a template for the design of inhibitors.