Cancer is one of the deadliest ailments in the United States, killing hundreds of thousands of people per year. One group of proteins that have been implicated in human cancers are the Phosphatase of Regenerating Livers, or PRLs. PRL-1 has been shown to have differing effects on cancer. Some research has found this protein to be an oncogene while others have found it to be a tumor suppressor. In our lab, past research has shown that for dPRL-1 to function as a tumor suppressor, it must localize to the adherens junction. The adherens junction helps glue epithelial cells together; malfunctioning adherens junctions, found in some cancers, no longer glue cells together thereby allowing them to metastasize. To better understand how dPRL-1 can function as a tumor suppressor, I performed a co-immunoprecipitation on the homolog, dPRL-1 in Drosophila melanogaster to discover the proteins that dPRL-1 interacts with in vivo at the adherens junction. To do this, I overexpressed dPRL-1 in the developing wing epithelium of the organisms and used an antibody to dPRL-1 for co-immunoprecipitation. This approach is an unbiased way to identify proteins that dPRL-1 interacts with while functioning as a tumor suppressor. My initial attempts at co-immunoprecipitation revealed one unknown protein of possible significance, but I have been unable to identify it yet. I am also testing my protocol to see if E-cadherin, a protein component of the adherens junction that contributes to cell adhesion, and dPRL-1 co-immunoprecipitate. We expect this interaction since our lab found that the two proteins colocalize via fluorescence microscopy. The proteins that dPRL-1 interacts with when functioning as a tumor suppressor could be mutated or missing in cancer cells thereby preventing dPRL-1 from functioning as a tumor suppressor. Hopefully, this knowledge will help clinicians make more effective chemotherapies to target cancers affected by PRLs.

It is not fully known what controls signal preference in branched signal transduction pathways. One hypothesis is that the configuration and orientation of receptors determines signal preference. We are investigating this hypothesis in the context of the Tie2 receptor pathway which has downstream signaling involved in cell survival via Akt, migration via Dok-R, and sprouting via FAK, among other branches. We are using computer designed protein tools, short-armed claw trimers and nanocages, conjugated to F domains of the canonical Tie2 ligand angiopoietin 1 in our investigation. We have shown that short-armed claw trimers with three conjugated F domains do not yield increased phosphorylation of Akt, suggesting that more than three receptors must cluster together for signal activation. The nanocages are multivalent, for they have multiple F domains conjugated to them. We have different valency nanocages with different percentages of F domain conjugation. We are using these different valency nanocages to investigate which valency is optimal for receptor activation. We have shown that higher valency nanocages yield increased phosphorylation of Akt. We predict that higher valency nanocages cluster more Tie2 receptors together in a specific conformation. Thus, we show a positive correlation between Tie2 clustering and the phosphorylation of Akt due to Tie2 activation. Now we will investigate combination conjugations of F domain and integrin binders to the nanocages as well as other branches of the Tie2 pathway: Dok-R corresponding to wound healing ability and FAK corresponding to tube formation ability. This work has potential to help generate therapeutic compounds for wound healing.