Browsing by Subject "Mutation"
Now showing 1 - 2 of 2
Results Per Page
Sort Options
Item APOBEC3 subcellular localization and genomic editing(2012-10) Lackey, Lela LynnThe APOBEC proteins are DNA cytosine deaminases with roles in immunity, including retroviral restriction and antibody maturation. Their activity theoretically makes them a danger to genomic DNA. The subfamily of APOBEC3 genes has expanded to included seven different genes in primates. Based on their subcellular localization, only a subset of these APOBEC3 proteins have access to genomic DNA, and may potentially deaminate genomic DNA. Although the nuclear envelope breaks down during mitosis, I demonstrate that none of the APOBEC3s gain access to genomic DNA during cell division. However, APOBEC3B and other APOBEC3 proteins have access to genomic DNA during interphase. I also show that APOBEC3B is actively imported into the nuclear compartment. In general, APOBEC3 nuclear localization and deaminase activity correlate with ability to affect cell cycle progression, implicating these APOBEC3s in deamination of genomic DNA. In support of these conclusions, I observed cell death, activation of the DNA damage response and DNA mutations after ectopic expression of APOBEC3A and APOBEC3B. Moreover, endogenous APOBEC3B is demonstrably nuclear and active in breast cancer cell lines where it causes genomic deamination and mutations. Endogenous APOBEC3B is highly expressed in more than half of human breast cancers compared to normal breast tissues. In addition, sequences from tumors with higher levels of APOBEC3B have more mutations, and these mutations match APOBEC3B's deamination signature. My thesis work further defines the subcellular localization of the APOBEC3 family and provides the first evidence that APOBEC3B is involved in a human cancer type.Item APOBEC3B-driven mutagenesis in breast and other human cancers(2013-08) Burns, Michael BradleyCancer is a disease that results from alteration of the cellular genome. The sources of these changes are multifarious, and in many cases unknown. This thesis focuses on the polynucleotide cytosine deaminase, APOBEC3B, as a newly discovered source of mutation in multiple human cancers. As a deaminase, APOBEC3B converts cytosines to uracils in single-stranded DNA. These uracil lesions are mutagenic as failure to properly repair them can result in a wide variety of mutation types. The initial discovery of this mutational phenomenon was described mechanistically using a variety of biochemical, genetic, and cellular assays in breast cancer cell lines. Follow-up work using publicly available next generation sequencing and clinical data indicated that this effect is operating in a large proportion of breast cancers. Expanded bioinformatic analysis that assessed APOBEC3B's potential impact was expanded to include 18 other human cancer types in addition to breast. This work shows that APOBEC3B is likely a significant contributor to the genetic heterogeneity in breast, head & neck, bladder, cervical, and lung (adeno- and squamous cell) carcinomas as evidenced by differential levels of expression in cancer tissues, increased mutation load, mutation clusters (kataegis), and an APOBEC3B mutation signature in tumors expressing high levels of this enzyme. Taken together, this thesis is a body of work describing a previously unappreciated source of genetic heterogeneity in several human cancers.