Ubiquitin Pathway regulation of Ras signaling.

Ras signaling helps translate external signals into biological outputs; therefore, its precise control is crucial. We discovered that ubiquitination is a novel way to down-regulate both wild-type and oncogenic forms of Ras. We established that this negative regulation of Ras is crucial during development, and we identified Rabex-5 as the Ras ubiquitin ligase. Ras ubiquitination is a relatively recently discovered phenomenon, so there is little understanding of its biological regulation or its role in development and disease. We employ multiple approaches to address these unexplored areas: We are utilizing loss-of-function mutations in (a) E1, the most upstream enzyme in the ubiquitin pathway, and (b) Rabex-5, the Ras ubiquitin ligase, to modulate the level of Ras ubiquitination in vivo. (c) We are identifying de-ubiquitinating enzymes and other ubiquitin pathway components that regulate Ras. (d) We are establishing how Ras ubiquitination is regulated spatially and temporally throughout development, in Drosophila models of human diseases, and in mammalian cancer cell lines.

Rabex-5 coordination of Ras and Notch signaling.

Notch signaling is another important developmental signaling pathway which is also implicated in cancer. Ras and Notch have a complicated relationship which is sometimes synergistic and other times antagonistic. Surprisingly, we discovered that the Ras E3 Rabex-5 also negatively regulates Notch signaling. Ongoing projects in vivo in Drosophila and in mammalian cancer lines are addressing how Rabex-5 coordinates Ras and Notch signaling in development and in disease.

Ubiquitin Pathway regulation of longevity/Modeling XL-SMA in the fly.

In addition to utilizing loss-of-function mutations in E1 to elucidate the role of ubiquitination in Ras regulation, we also discovered that even heterozygous mutation in E1 leads to dramatic reduction in lifespan. Moreover, mutation in human E1 is associate with X-linked Infantile Spinal Muscular Atrophy (XL-SMA). This devastating disease results in tremendous suffering and affected infants typically die in their first two years. Our fly E1 mutants represent an animal model system amenable to genetic and pharmacological testing. Investigating fly E1 mutants will allow us to make general advances regarding the role of ubiquitination in aging and longevity and will serve as an animal model to advance our understanding of XL-SMA.

Hippo Pathway regulation of proliferation and organ size.

The Hippo Pathway is a master regulator of growth, cell proliferation, and apoptosis. Its loss promotes cell division, cell death resistance, overgrowth, and increased organ size. Because an organ size “checkpoint” ensures that proliferative changes do not alter organ size, how Hippo signaling integrates proliferation control within an organ size “checkpoint” remains a major open question. The Hippo Pathway inhibits the oncogene Yorkie/YAP a transcriptional co-activator. However Yki/YAP is crucial in healthy cells, to a therapeutic window of targeting YAP in cancer may be small. Identification of other post-translational targets of the Hippo Pathway may explain the role of the pathway in specific developmental contexts and may identify additional therapeutic points of entry for cancers associated with impairment of the Hippo Pathway. Through in vitro screening, we identified 100 additional post-translational pathway targets. We are focusing on characterizing a number of these in vivo with goals of (a) elucidating the complicated relationship between proliferation and organ size, (b) revealing tissue-specific and cancer-specific targets, and (c) identifying pathway targets that could serve as therapeutic points of entry.