Project Summary

The multidrug resistance-associated protein (MRP) subfamily of the ATP-binding cassette (ABC) transporters play a key role in protecting cells from drugs and environmental toxins such as heavy metals (mercury, arsenic, lead, and cadmium). The MRPs (also called ABCCs) are distinguished from other ABC transporters by two striking hallmarks: 1) they contain an additional "N-terminal extension" with a conserved region called LO, and 2) they transport substrates in the form of glutathione (GSH)-conjugates or complexes. The overall goal of this project is to determine how MRP function is regulated in vivo. Specifically, we will define the role of MRP-protein kinase interactors in regulating MRP function. Our experimental approach is directed at identifying the mechanism by which an MRP-protein kinase interactor, the yeast casein kinase 2 (CKII) alpha subunit, Ckalp, negatively regulates the function of the yeast MRP, yeast cadmium factor 1 (Ycflp). Cka1p has been shown to be catalytically active as both a homodimer and a heterodimer with the CKII alpha' subunit, Cka2. Therefore this study will determine if Cka1p regulation of Ycf1p function requires Cka2p. In addition we will examine the role of the homologous human kinase, CKalpka in regulating human MRP1 and MRP6 function. The studies described within this proposal will provide useful insight into the role of kinases in the regulation of MRP function, the role of the MRPs in "metabolic quality control," and have the potential to establish new and more effective treatments for MRP related diseases such as cystic fibrosis, multidrug resistance in cancer, and pseudoxanthoma elasticum. We will use the powerful genetic, molecular, and cellular tools available in Saccharomyces cerevisiae to address several key issues of the MRPs. The specific aims of this project are: 1) To determine the mechanism by which Ckalp negatively regulates Ycflp function, 2) To determine if Cka1p regulation of Ycf1p function requires Cka2p, and 3) To determine the role of CKII in regulating human MRP1 and MRP6 physiological function. Support for this COBRE grant will provide Dr. Paumi with the necessary funds to generate preliminary data and valuable mentorship that will ultimately result in an NIH funded R01 grant.

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