Mitchell, John L. A.
Ph.D. (Doctor of Philosophy)
Department of Biological Sciences
Cancer cells--Growth--Regulation||\ODC antizyme inhibitor
The polyamines putrescine, spermidine and spermine are ubiquitous organic cations essential for normal cell growth and proliferation. Since abnormalities in polyamine levels have been implicated in stimulating cell growth, tumor development and cytotoxicity, polyamine metabolism and transport must be tightly regulated to maintain optimal levels. Antizyme (AZ) is a key regulatory protein that controls polyamine levels by binding to and targeting ornithine decarboxylase (ODC), the first and rate-limiting enzyme in polyamine biosynthesis, for degradation by 26S proteasome and inhibiting polyamine incorporation into cells. Antizyme inhibitor (AZI) was first identified as a protein that specifically binds to AZ with higher affinity than that of ODC and releases ODC from inactive complexes with AZ. This study was aimed at understanding the physiological functions of AZI in regulating polyamine homeostasis in mammalian cells. A cell line with controlled gene expression was created and physiological effects of AZI overexpression were investigated. AZI was determined to be a very labile protein that colocalized with AZ in various organelles and cell compartments. Overexpression of AZI diminished the inhibitory effects of AZ on both ODC activity and polyamine transport. When AZI was overexpressed, it bound AZ, preventing ODC degradation, and thereby increased ODC activity and polyamine levels. Furthermore, AZI overexpression prevented AZ-mediated feedback control of polyamine uptake. Consequently, inward transport of natural polyamines as well as structural analogs of polyamines were increased. Because polyamine analogs have great potential as chemotherapeutic agents, the effects of AZI on uptake and sensitivity to selected analogs were investigated. Similar overexpression of AZI may be a common trait of certain human tumors. Since cells overexpressing AZI have diminished feedback regulation of the polyamine transporter, higher analog concentrations were seen to accumulate. Although these cells were generally more sensitive to the treated analogs, the amplitude of the effect varied with the different analogs. This study reveals that AZI overexpression diminishes the effects of AZ and increases ODC activity and polyamine levels, which may play a role in carcinogenesis. Conversely, AZI overexpression increases uptake and sensitivity of certain polyamine analogs, which can be exploited further for the design of more effective cancer therapies.
Thane, Thynne K., "Investigation of the physiological functions of antizyme inhibitor in polyamine regulation and control of cancer cell growth" (2006). Graduate Research Theses & Dissertations. 3658.
xiv, 205, pages (some color pages)
Northern Illinois University
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