This project aims to determine the mechanisms underlying the regulation of a newly identified iron importer, ZIP14, and to examine its function in establishing iron homeostasis in the body. Elucidating the mechanisms of ZIP14's regulation offers opportunities to enhance our understanding of both the normal iron metabolism and the pathogenesis of iron-overload disorders. Therefore, this proposed research is directly relevant to public health and the NIH's mission. I have three and a half years of postdoctoral research experience when submitting this application. My long term career goal is to become an independent investigator at a major academic institution and to contribute to biomedical knowledge and general health improvement. I am determined to devote my career to examine the molecular basis of iron metabolism disorders and put this knowledge to use for improving the life quality of patients with these diseases. The central theme of my postdoctoral research has been the investigation of the mechanisms underlying iron-metabolism and the identification of therapeutic gene targets such that gene- specific therapies can be applied on a personalized basis. This will remain my primary objective for the foreseeable future. My central goal during the five-year span of this K99/R00 award is to achieve research independence. The mentored phase of this award offers the perfect opportunity to receive the necessary training and develop the tools needed for validation and further characterization of ZIP14. I will take advantage of the two-year mentored phase to gain additional methodological training, improve my communication skills, and enhance my management and teaching skills. I will start looking for university faculty positions towards the end of the first year of the K99 stage to ensure timely and smooth transition to the R00 phase. I will continue to work toward establishing research independence by preparing an application for R01 support during the second to the third year's transition of the independent R00 phase. The projects that I have accomplished to this point as well as my future goals regarding this subject suggest a career path that is ideal for the K99/R00 funding mechanism. In my preliminary studies, I have identified an amino acid residue that is involved in regulating the iron sensitivity of ZIP14. This residue is closely adjacent to a mutated amino acid that was recently identified in patients with brain iron overload, suggesting the existence of a potential iron sensing motif near this region. I have also found a novel proteasome-mediated pathway for the degradation of ZIP14. This pathway involves dislocation of ZIP14 from the membrane into the cell cytosol. Of note, iron prevents the extraction of ZIP14 from the membrane for proteasomal processing. Further, I found that ZIP14 ubiquitination is positively regulated by p53. However, the region that is critical in determining ZIP14's iron sensitivity, the mechanism of ZIP14's membrane dislocation, and the role of p53 in the iron- dependent regulation of ZIP14 are not known. Therefore, a more complete examination of the mechanisms underlying the iron regulation and the degradation process of ZIP14 is required. Importantly, further exploration of the contribution of ZIP14 to the pathogenesis of disorders in iron metabolism represents an ideal project from which to launch an independent research career. There are a number of preliminary experiments, assays, and reagents that need to be created and performed during the mentored phase before a full analysis of ZIP14 can be undertaken. Upon completion of these steps, the independent phase of the award will allow for complete follow-up analysis of the hypotheses generated by data during the mentored phase. Moreover, this K99/R00 award will enable me to establish a structured system for validation and characterization of the in vitro and in vivo roles of ZIP14 with which my eventual laboratory can explore new topics in iron biology.
Iron content inside cells needs to be tightly controlled to meet the dual challenge of avoiding iron deficiency and iron overload. Iron importers play fundamental roles in the maintenance of cellular iron homeostasis and systemic iron homeostasis within the body. This project seeks to understand the mechanism underlying the regulation of an iron importer, ZIP14.