Hypoxia plays an important role in the establishment and growth of tumors by inducing angiogenesis. Hypoxia induces angiogenesis by altering gene expression at the transcriptional and post-transcriptional levels. One of the post-transcriptional effects is mediated at the level of translation. Specifically, hypoxia inhibits protein synthesis in the cell and under these conditions most mRNAs will not be translated efficiently. However, the HIF-1alpha and VEGF A mRNAs, which encode proteins that are critical players in the induction of angiogenesis, continue to be translated at this time. These mRNAs contain internal ribosome entry sites (IRESes) and can initiate translation through an alternative mechanism termed internal initiation. This proposal will test two hypotheses: 1) that the translational repression that occurs during hypoxia does not effect the translation of all mRNAs equally, and 2) that internal initiation of translation is the molecular mechanism that allows mRNAs to be translated during hypoxia. The first hypothesis will be tested using a novel approach that utilizes microarray analysis to identify the mRNAs that continue to be translated during hypoxia. The second hypothesis will be tested by determining if these translationally active mRNAs contain IRESes, using a dicistronic assay system. The trans-acting factors that interact with several of these IRESes will be detected by UV-crosslinking experiments, purified, and identified by microsequencing. The function of the cis- and trans-acting factors in IRES activity will be determined using site-directed mutagenesis and RNA interference experiments. If we find trans-acting factors that are required for IRES activity then these mRNA/protein interactions may serve as novel targets for drug development. If therapeutics could be developed that disrupt these RNA/protein complexes, then this may prevent the translation of mRNAs encoding proteins required for tumor angiogenesis and therefore prevent tumor growth.
| Thomas, Jeff D; Dias, Lizalynn M; Johannes, Gregg J (2008) Translational repression during chronic hypoxia is dependent on glucose levels. RNA 14:771-81 |
| Thomas, Jeff D; Johannes, Gregg J (2007) Identification of mRNAs that continue to associate with polysomes during hypoxia. RNA 13:1116-31 |
