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Hutchinson-Gilford progeria syndrome (HGPS) is a detrimental premature aging disease caused by a point mutation in human LMNA gene. This mutation results in the abnormal accumulation of a truncated pre-lamin A protein called progerin. Among the drastically accelerated signs of aging in HGPS patients, severe skin phenotypes such as alopecia and sclerotic skins always develop with the disease progression. In this dissertation, I study the HGPS molecular mechanisms focusing on early skin development by differentiating patient-derived induced pluripotent stem cells (iPSCs) to a keratinocyte lineage. Interestingly, HGPS iPSCs showed an accelerated commitment to the keratinocyte lineage than the normal control. To study potential signaling pathways that accelerated skin development in HGPS, I investigated the WNT pathway components during HGPS iPSCs-keratinocytes induction. Surprisingly, despite the unaffected β-catenin activity, the expression of a critical WNT transcription factor LEF1 was diminished from an early stage in HGPS iPSCs-keratinocytes differentiation. Chromatin immunoprecipitation (ChIP) experiment further revealed strong bindings of LEF1 to early-stage epithelial development markers K8 and K18 and that the LEF1 silencing by siRNA down-regulates the K8/K18 transcription. During the iPSCs-keratinocytes differentiation, correction of HGPS mutation by Adenine base editing (ABE), while in a partial level, rescued the phenotypes for accelerated keratinocyte lineage-commitment. ABE also reduced the cell death in HGPS iPSCs-derived keratinocytes. These findings brought new insight into the molecular basis and therapeutic application for the skin abnormalities in HGPS.

One important feature in both HGPS and normal aging is the elevated levels of Reactive Oxygen Species (ROS), which are generated from metabolic pathways to cause oxidative damage to macromolecules within the cells. Although peroxisomal bioreactions can generate free radicals as their byproducts, many metabolic enzymes within the peroxisomes play critical roles as ROS scavengers, particularly catalase. In this dissertation, I observed impaired peroxisomes-targeting protein trafficking, which suggested that the poorly assembled peroxisomes might cause high oxidative stress, contributing to the premature senescent phenotype in HGPS. I also investigated the ROS clearance efficiency by peroxisomal enzymes and found a significantly decreased catalase expression in HGPS. Furthermore, I evaluated the effects of two promising HGPS-treatment drugs Methylene Blue and RAD001 (Everolimus, a rapamycin analog), on catalase in HGPS fibroblasts. I found that both drugs effectively reduced cellular ROS levels. As a well-known antioxidant, MB did not affect catalase expression or activity. Interestingly, the RAD001 treatment significantly upregulated catalase activity in HGPS cells. This is the first characterization of peroxisomal function in HGPS and provides new insights into the cellular aspects of HGPS and the ongoing clinical trial.