[-] Show simple item record

dc.contributor.advisorPetris, Michael J.eng
dc.contributor.authorGudekar, Nikita S.eng
dc.date.issued2020eng
dc.date.submitted2020 Falleng
dc.description.abstractCopper (Cu) is an essential trace dietary metal which is also potentially toxic. Cells have evolved an intricate system of transporters and chaperones to carefully regulate Cu homeostasis. Cu uptake is mediated by the Cu1+-specific permease, CTR1. Cytoplasmic Cu is delivered to specific compartments via a series of metallochaperones. In the trans-Golgi network (TGN), the ATP7A or ATP7B proteins are required to pump Cu into the secretory pathway to various cuproenzymes. ATP7A also plays a protective role against Cu toxicity by trafficking to the plasma membrane to export Cu from the cell. Metallothioneins (MTs) are another type of protein that plays a protective role against Cu toxicity by sequestering excess Cu in the cytoplasm. There are four isoforms of MT in mice of which MT I and MT II are ubiquitously expressed. Our preliminary data demonstrates the essentiality of ATP7A, MTI and MTII in copper tolerance. We generated a fibroblast cell line in which Atp7a, Mt-I, and Mt-II genes were deleted (Atp7a-/Mt-), which resulted in sensitivity of these cells to sub-micromolar levels of copper. Using a genome-wide CRISPR-Cas9 deletion screen, we identified a novel gene, VHL (von Hippel-Lindau), that when deleted confers Cu tolerance to Atp7a-/Mt- cells. The VHL protein is a ubiquitin-ligase responsible for the degradation of HIF transcription factors, which are the major regulators of gene expression in response to hypoxia. We show that VHL deletion confers Cu tolerance by stimulating the expression of the ATP7B copper transporter. Pharmacological inhibition of VHL-HIF1[alpha] by Roxadustat (a prolyl hydroxylase inhibitor), which is clinically used to treat anemia, conferred resistance to copper by upregulating ATP7B. Importantly, Roxadustat was found to increase hepatic ATP7B expression in vivo. These novel data identify a previously unknown link between copper homeostasis and oxygen sensing and have potential translational implications for the treatment of Wilson disease, a copper overload condition cause by insufficient expression of ATP7B.eng
dc.description.bibrefIncludes bibliographical references (pages 102-118).eng
dc.format.extentxiii, 119 pages : illustrationseng
dc.identifier.urihttps://hdl.handle.net/10355/88873
dc.identifier.urihttps://doi.org/10.32469/10355/88873eng
dc.languageEnglisheng
dc.publisherUniversity of Missouri--Columbiaeng
dc.relation.ispartofcommunityUniversity of Missouri--Columbia. Graduate School. Theses and Dissertationseng
dc.subjectAuthor-supplied keywords: KEYWORDSeng
dc.titleThe identification of novel genes involved in copper and oxygen metabolismeng
dc.typeThesiseng
thesis.degree.disciplineGenetic area program (MU)eng
thesis.degree.grantorUniversity of Missouri--Columbiaeng
thesis.degree.levelDoctoraleng
thesis.degree.namePh. D.eng


Files in this item

[PDF]

This item appears in the following Collection(s)

[-] Show simple item record