Implications for the 8p12-11 Genomic Locus in Breast Cancer Diagnostics and Therapy: Eukaryotic Initiation Factor 4E-Binding Protein, EIF4EBP1, as an Oncogene

The goal of this research was to elucidate the individual and cooperative role of oncogenes within the frequently amplified 8p12-11 genomic locus which is known to predict poor prognosis and resistance to endocrine therapy in breast cancer. The role of specific oncogenes from the 8p11-12 amplicon in breast tumors is highlighted and the construction of expression constructs for future studies is described. Further exploration of the oncogenic role of Eukaryotic Initiation Factor 4E-Binding Protein (4EBP1, EIF4EBP1) showed 4EBP1 is often highly overexpressed in malignant tumors and is predicted as an essential driving gene in many cancer cell lines in vitro, so we hypothesized 4EBP1 was a driving oncogene in breast cancers. High 4EBP1 gene expression was associated with reduced relapse free patient survival across all breast tumor subtypes, including post treatment tumors. We found 4EBP1 was crucial to the proliferation of all breast cancer cell lines tested, but not normal cells, making it a prime therapeutic target. This is the first report showing 4EBP1 levels influence malignant estrogen receptor α (ER, ESR1) levels, and we also explored the levels of other influential cancer related genes including oncogenes within 8p11-12 which are known to influence ER. Applied data mining and a current clinical trial implied importance for 4EBP1 as a biomarker for therapeutic efficacy, so we explored mTOR inhibition in multiple cell culture models. It is likely 4EBP1 mediates upstream signaling from many known tumorigenic signaling cascades in adaptive and different ways thus it should be further explored and has much potential as a biomarker and target for breast cancer, especially endocrine resistant tumors

Characterization of the Norspermidine/Spermidine Transport Protein, PotD1, in Vibrio cholerae

Biofilm formation increases antibiotic resistance among bacteria; therefore, a thorough understanding of the proteins and molecules affecting biofilm formation is necessary. PotD1, the substrate binding protein of the PotABCD1 spermidine transport system, negatively regulates biofilm formation in the intestinal pathogen Vibrio cholerae. This work characterized the binding properties of PotD1 to its ligands and its effect on biofilm formation through genetic characterization of the binding cleft and the ability of PotD1 to interact with various polyamines, which can alter biofilm levels in this bacterium. The results demonstrated that PotD1 is also responsible for the import of norspermidine. This is the first example of a norspermidine transporter reported. Ligand competition assays showed PotD1 has a higher binding affinity for spermidine over norspermidine. This work also showed the amino acids W252 and D254 play a large role in spermidine transport, and amino acids E168 and W32 play a less important role. Lastly, mutations in the amino acids that diminished spermidine import only had a partial effect on biofilm formation, suggesting spermidine uptake and biofilm formation are partially correlated. Thus, this work demonstrates that PotD1 alters biofilm formation by both its ability to uptake spermidine and through another yet unidentified mechanism