TRF2 is an essential telomeric protein involved in preventing the telomere ends from being recognized as DNA breaks. I have shown that TRF2 does not appear to play a major role in the UV -induced DNA damage response in IMR90, Cockayne syndrome or XPC deficient cells. TRF2 binds telomeric DNA via its Myb domain and also contains an N-terminal basic domain. Expression of TRF2MMM causes telomere fusions, whereas TRF2^(ΔB) causes rapid deletion of telomeric DNA, as both phenotypes result in senescence. These phenotypes are dependant upon recombination events. Thus, the basic domain of TRF2 may be essential to suppress recombination events at telomeres. However, it is not fully understood what amino acid residues in the basic domain of TRF2 are indispensable to maintain its function. By creating mutations in the arginine residues in the basic domain of TRF2, I have shown that the positive charge of the basic domain alone is not sufficient to maintain its protective function. By expressing these TRF2 mutants in the presence or absence of the Myb domain in HT1080 and BJ/hTERT cells, I have been able to recapitulate the TRF2^(ΔB) and TRF2^(ΔBΔM) decreased proliferation and senescence phenotypes. Furthermore, by analyzing anaphase and metaphase chromosomes and performing Southern blotting, I have shed light on the molecular mechanisms responsible for the deleterious phenotypes observed in the TRF2 mutants. Amino acid changes from arginines to lysines introduced into the basic domain of TRF2 results in a significant increase in telomere doublets. However, when these TRF2 mutants are expressed in the absence of the Myb domain, a significant increase in telomere fusions events occur. Collectively, my results indicate that more than one arginine residue in the basic domain is essential to maintain the protective function of TRF2, as these arginine residues may act as substrates for protein arginine methyltransferases. / Thesis / Master of Science (MSc)

Skin cancer is the most common type of cancer in the US. The American Academy of Dermatology estimated that more than 3.5 million new cases of skin cancer are diagnosed in the US each year and 1 in 5 Americans will likely to develop skin cancer in their life time. Most cases of skin cancer are caused by exposure to ultraviolet (UV) radiation from the sun. Some of the most common sunscreen ingredients are unstable and can form harmful radicals upon exposure to UV radiation. There is a strong clinical need for a more stable and effective sunscreen ingredient such as bovine milk phospholipids (MP). Phospholipids were shown to have beneficial health effects such as regulation of the inflammatory reactions, protective effects against colon cancer, and reducing cardiovascular risk factors. Previous histology and MTT tissue viability research studies suggested that MP act upon skin cells in a protective manner against UV radiation. This thesis aims to further investigate the protective effects of bovine milk phospholipids by evaluating the expression of a UV-induced DNA damage marker, cyclin-dependent kinase inhibitor, p21WAF1/CIP1. Western blots were used to quantify p21 expression in human keratinocytes in four categories of samples: No-UV, UV, UV+MP, MP and in HeLa (p21 positive control). In the No-UV samples, cells were not irradiated by UV light. However, in the UV samples, keratinocytes were exposed to a UV dosage of 10 mJ/cm2. In the UV+MP samples, keratinocytes were first treated with 1% MP solution (w/v) in their culture media for 24 hours then exposed to a UV dosage of 10 mJ/cm2. In MP, keratinocytes were treated with 1% MP solution in their culture media for 24 hours. Total cell proteins were extracted 24 hours post-UV irradiation. The same amount of protein from each sample (determined by BCA assay) was loaded into a 4-12% Bis-Tris SDS-PAGE gel, run under denaturing, non-reducing conditions then blotted and treated with antibodies for the quantitative detection of p21 proteins. Finally, intensities of p21 protein bands were analyzed by using ImageJ software. Under non-reducing conditions, three p21 proteins covalently bonded with each other showed up as 63 KDa molecules on the PVDF membrane. The UV, and HeLa samples showed a 2.28 fold, and 1.23 fold increase in p21 expression, respectively, compared with the No-UV samples control. The MP samples showed a 0.948 fold decrease in p21 compared with the No-UV samples, and the UV+MP samples showed only a 1.13 fold increase in p21. When comparing with the UV sample, the UV+MP sample has 50.4% less p21 expression. Less p21 expression in the UV+MP sample compare with the UV sample suggested that less DNA damage occurred in the sample that was treated with milk phospholipids. This result strongly suggests that 1% bovine milk phospholipids can protect skin cells from UV induced DNA damage.

Distributed temperature and strain sensing is demanded for a wide range of applications including real-time monitoring of industrial processes, health monitoring of civil infrastructures, etc. Optical fiber distributed sensors have attracted tremendous research interests in the past decade to meet the requirements of such applications. This research presents a multiplexed sensor array for distributed temperature and strain sensing that can multiplex a large number of UV-induced sensors along a single fiber. The objective of this research is to develop a quasi-distributed sensing technology that will greatly increase the multiplexing capacity of a sensor network and can measure temperature and strain with a high accuracy and high resolution. UV-induced intrinsic Fabry-Perot interferometric (IFPI) optical fiber sensors, which have low reflectance and low power loss, are good candidates for multiplexed sensors networks. Partial reflectors are constructed by irradiating photosensitive fiber with a UV laser beam. A pair of reflectors will form a Fabry-Perot interferometer that can be used for temperature and strain sensing. A sensor fabrication system based on a pulsed excimer laser and a shadow mask is developed. A spectrum-based measurement system is presented to measure the interference fringes of IFPI sensors. A swept coherent light source is used as the light source. The spectral responses of the IFPI sensors at different wavelengths are measured. A frequency division multiplexing (FDM) scheme is proposed. Multiple sensors with different optical path differences (OPD) have different sub-carrier frequencies in the measured spectrum of the IFPI sensors. The multiplexing capacity of the sensor system and the crosstalk between sensors are analyzed. Frequency estimation based digital signal processing algorithms are developed to determine the absolute OPDs of the IFPI sensors. Digital filters are used to select individual frequency components and filter out the noise. The frequency and phase of the filtered signal are estimated by means of peak finding and phase linear regression methods. The performance of the signal processing algorithms is analyzed. Experimental results for temperature and strain measurement are demonstrated. The discrimination of the temperature and strain cross sensitivity is investigated. Experimental results show that UV-induced IFPI sensors in a FDM scheme have good measurement accuracy for temperature and strain sensing and potentially have a large multiplexing capacity. / Ph. D.

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