Vascular cell migration and VHL gene function in flies

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Angiogenesis is an integral part of solid tumor growth and holds promise as a target for effective therapeutics with limited deleterious side effects. Various anti-angiogenic therapies have focused on the growth factors, their cognate receptors or metalloproteinase inhibitors. These therapies, however, have not been overwhelmingly successful. Additional studies on angiogenic mechanisms can help identify new therapeutic targets. One interesting unconventional angiogenic regulator is the von Hippel- Lindau tumor suppressor gene (VHL). Mutations in the VHL tumor suppressor gene predispose individuals to highly vascularized tumors. We have recently isolated the Drosophila homolog of VHL (D-VHL) and showed that it preserved the anti-angiogenic function in the Drosophila tracheal system, which provides an accessible and relevant in vivo model for studying vascular tubule formation and cell migration. The prevailing model suggests that VHL may function as a subunit of a ubiquitin ligase, which facilitates degradation of its targets. Among its proposed targets is the alpha-subunit of hypoxia-inducible factor 1alpha (Hif-1alpha) that is a transcriptional activator of the vascular-endothelial growth factor (VEGF) and is highly unstable in normal oxygen condition (normoxia). Thus, the hyper-vascularized phenotype of VHL mutant can be explained by an up-regulation of Hif-1alpha in normoxia. Thus, the hyper-vascularized phenotype of VHL mutant can be explained by an up-regulation of Hif-1alpha in normoxia, resulting in over-expression of the potent angiogenic factor VEGF. This interesting model has not been fully validated in vivo, however. Our recent work on D-VHL further suggests that it can modulate cell migration, and perhaps cell division, by directly regulating the components of the actin filaments and microtubules. These cytoskeletal components may constitute a new battery of anti-angiogenic targets. We have also devised a promising new assay system and a provocative conjecture for analyzing the relationship between VHL and Hif-1alpha in vivo. Finally, our recent results suggest that VHL is a multi-functional and may regulate cell division. These results also indicate that VHL may be more than a ubiquitination enzyme as suggested by the current model. In order to formulate a thorough and focused research project, we need to first accomplish three short-term goals, summarized in three Specific Aims in this exploratory study (R21); (1) To determine if the Drosophila Hif-1alpha activity is regulated by hypoxia in vivo. (2) To assess the relevance of the potential D-VHL interacting proteins and complete the ongoing yeast 2-hybrid screen for D-VHL interacting proteins. (4) Generating genomic D-VHL mutant.