Nuclear PTEN Regulates Thymidylate Biosynthesis and Cellular Sensitivity to Antifolate Treatment- [electronic resource]
Nuclear PTEN Regulates Thymidylate Biosynthesis and Cellular Sensitivity to Antifolate Treatment- [electronic resource]
- 자료유형
- 학위논문파일 국외
- 최종처리일시
- 20240214101505
- ISBN
- 9798380347693
- DDC
- 574
- 서명/저자
- Nuclear PTEN Regulates Thymidylate Biosynthesis and Cellular Sensitivity to Antifolate Treatment - [electronic resource]
- 발행사항
- [S.l.]: : Duke University., 2023
- 발행사항
- Ann Arbor : : ProQuest Dissertations & Theses,, 2023
- 형태사항
- 1 online resource(193 p.)
- 주기사항
- Source: Dissertations Abstracts International, Volume: 85-03, Section: B.
- 주기사항
- Advisor: Chen, Ming.
- 학위논문주기
- Thesis (Ph.D.)--Duke University, 2023.
- 사용제한주기
- This item must not be sold to any third party vendors.
- 초록/해제
- 요약Metabolic reprogramming contributes to tumorigenesis and holds significant promise for cancer therapy. The PTEN tumor suppressor governs a variety of biological processes, including metabolism, by acting on distinct molecular targets in different subcellular compartments. In the cytoplasm, PTEN regulates a plethora of metabolic processes through antagonizing the PI3K/AKT/mTORC1 pathway. However, the metabolic regulation of PTEN in the nucleus remain undefined. Using a gain-of-function approach to examine the metabolic consequences of PTEN targeted to different sub-cellular compartments in human prostate cancer cell lines, we reveal a nuclear function for PTEN in controlling de novo thymidylate biosynthesis and may also open novel therapeutic avenues for targeting nuclear-excluded PTEN prostate cancer cells with anti-folate cancer treatment. The first four chapters of this dissertation are introductory information that outlines the role of PTEN in cancer, the importance of metabolic compartmentalization, the fundamentals of pyrimidine biosynthesis pathways, and the novelty of anti-folate cancer treatments. Chapter 1 explains the role of PTEN as a tumor suppressor and continues on to discuss its role at the cell membrane as a metabolic regulator. The gap in knowledge in the field is understanding the role nuclear PTEN plays as a metabolic regulator. This is important as nuclear PTEN has been shown to be associated with more aggressive cancer phenotypes. Chapter 2 focuses on the background of metabolic compartmentalization and how, by strategically placing genes and metabolites spatiotemporally, the cell is able to execute key molecular mechanisms more efficiently. In this chapter, we highlight where the current field is with metabolic compartmentalization, the advantages of compartmentalization and how utilization of this knowledge can be used for definitive therapeutics. Chapter 3 focuses on pyrimidine biosynthesis and thymidylate biosynthesis. Thymidylate is synthesized de novo by thymidylate synthase (TYMS), with the enzymes dihydrofolate reductase (DHFR) and methylenetetrahydrofolate dehydrogenase 1 (MTHFD1) or serine hydroxymethyltransferase (SHMT) which are required to regenerate 5, 10-methylenetetrahydrofolate. MTHFD1 is the primary source of 5,10-methylenetetrhydrofolate generation, and therefore its proper function in the nucleus ensures the functioning of de novo thymidylate biosynthesis. Using Mass-Spectrometry, we discovered that MTHFD1 is a top candidate protein interacting with PTEN in human prostate cancer cells. This is the key focus of this paper and will be important background for Chapter 7 and 8 which delve into the thymidylate pathway and the potential role of nuclear PTEN. A deeper understanding of nuclear PTEN's regulation of MTHFD1 may, in turn, open new therapeutic avenues for anti-folate cancer treatment tailored to PTEN sub-cellular localization. This brings us to Chapter 4, which focuses on the current market of anti-folate treatment and the importance of precision medicine and combinatorial treatment. Chapter 5 is the start of our project, it is the basis of the remainder chapters and what allowed us to elucidate the importance of nuclear PTEN. To explore the role of PTEN as not just a tumor suppressor, but as a metabolic regulator, we first sought to generate overexpression cell lines with PTEN localized to various subcellular compartments. As explained in chapter 1 and 2, we discuss how in the cytosol inactive PTEN can be recruited to the plasma membrane where it functions as a lipid phosphatase to suppress the activation of the proto-oncogenic phosphoinositide 3-kinase (PI3K)-AKT-mTOR signaling pathway. In the nucleus, PTEN acts to induce cell cycle arrest and maintain genomic stability. However, the role of PTEN in metabolism is incompletely understood. It is clearly understood that each subcellular compartment harbors specific metabolic activities and PTEN is present in different subcellular locations where it performs distinct functions acting on specific effectors. To explore this, we used a gain-of-function approach to examine the metabolic consequences of PTEN targeted to different sub-cellular compartments. Plasmids were generated using site-directed mutagenesis and PCR. We used the vector pTRIPZ, which is an inducible TET-ON system. This was necessary as overexpression of a tumor suppressor in cancer cell lines, if left permanently on, leads to slow cell growth and expulsion of the plasmid by inherent cancer cell mechanisms. We generated a vector plasmid, which was used as the baseline for all experiments, a wildtype PTEN plasmid, which was "normal" PTEN and PTEN had the ability to localize to the membrane or nucleus as it pleased, a cytoplasmic membrane plasmid, which localized PTEN permanently to the membrane, nuclear PTEN, which localized PTEN permanently to the nucleus, and mutant C124S PTEN, which generates a catalytically silent PTEN variant preventing its role at the membrane and its involvement in the PI3K-AKT-mTOR pathway. All of the plasmids were transfected as a lentivirus into PC3 and C4-2 prostate cancer cells. Both cell lines are. (Abstract shortened by ProQuest).
- 일반주제명
- Pathology.
- 일반주제명
- Biochemistry.
- 일반주제명
- Molecular biology.
- 키워드
- Tumor suppressor
- 기타저자
- Duke University Pathology
- 기본자료저록
- Dissertations Abstracts International. 85-03B.
- 기본자료저록
- Dissertation Abstract International
- 전자적 위치 및 접속
- 로그인 후 원문을 볼 수 있습니다.