Michaelis, SusanAlkhamis, Danyah2025-06-192025https://hdl.handle.net/20.500.14154/75616Lamin A is a nuclear scaffold protein, synthesized as a precursor called prelamin A, which undergoes several steps of posttranslational processing. This study explores the impact of mutations that affect the final proteolytic processing step, which is mediated by the zinc metalloprotease ZMPSTE24. We assessed the effects of these LMNA mutations, specifically at the L647 residue, on nuclear morphology and protein processing. After establishing a HeLa T-REx cell culture system, we expressed wild-type GFP-tagged LMNA and 11 GFP-tagged LMNA mutants. These mutations were examined against the abnormalities seen in cells from patients with Hutchinson-Gilford Progeria Syndrome (HGPS) and other disorders characterized by the accumulation of unprocessed prelamin A. Results in mammalian cells were compared to previous work performed in a humanized yeast system. Our findings indicate that diminished processing of prelamin seen in the yeast system is recapitulated in mammalian cells. Our results also indicate significant nuclear morphological abnormalities. However, there were no specific correlations between mutations with low cleavage efficiency and higher abnormal nuclei count. This research marks a significant step in understanding the impact of mutations at L647 in prelamin A processing and associated cellular defects. Overall, this study lays the foundation for future investigations into understanding the molecular mechanisms of how LMNA mutations may result in diseases.43deLamin AProgeriaHGPSAgingLaminCellPrelamin AZMPSTE24ProgerinThesis