Abstract Body (Do not enter title and authors here): Background. Mutations in ACTA2, encoding alpha-2 smooth muscle actin, account for ~20% of non-syndromic familial thoracic aortic aneurysms (TAA) and confer a high risk of dissection. However, the cell-type-specific transcriptional mechanisms driving ACTA2-associated TAA remain poorly defined, particularly in patients harboring variants of uncertain significance. Objective. To delineate the transcriptional and cellular consequences of a novel ACTA2 mutation (p.Met49Thr) in human thoracic aorta using single-nucleus RNA sequencing (snRNA-seq) on archived, formalin-fixed paraffin-embedded (FFPE) tissue. Methods. A 22-year-old male with a dissected 5.3-cm ascending aortic aneurysm was found to harbor a heterozygous ACTA2 c.146T>C (p.Met49Thr) variant. Nuclei were isolated from FFPE aortic tissue and subjected to snRNA-seq (10X Genomics), generating transcriptomes from 17,938 nuclei. This dataset was integrated with a non-genetic, hypertensive TAA control (TAA-NG). Clustering, differential expression, and trajectory analyses were performed in Seurat and Monocle3. Results. The ACTA2-p.Met49Thr sample showed marked expansion of vascular smooth muscle cells (VSMCs, 70.6% vs. 39.7% in TAA-NG) with significant upregulation of proliferation-associated genes (e.g., FOSB, FOS, JUN, DEPP1). VSMC trajectory analysis identified a transcriptional progression from quiescent to pro-proliferative states, enriched for aortopathy-related GWAS traits (e.g., aortic strain and diameter). Histological analysis confirmed elastic fiber fragmentation, increased medial VSMC cellularity, and adventitial fibrosis in ACTA2-p.Met49Thr, absent in TAA-NG. A publicly accessible Shiny app and code repository provide open access to all data and analyses. Conclusion. This study defines a causal ACTA2 variant that promotes VSMC expansion and transcriptional reprogramming toward a pro-proliferative phenotype in human TAA, supporting a paradigm in which ACTA2 mutations drive disease through functional cell state transitions. Our optimized FFPE-compatible snRNA-seq protocol also enables rapid functional annotation of clinical variants that may improve precision diagnostics for familial aortopathy.