Alternative Splicing Rewiring by Neurofibromin Isoforms Through ER-alpha and RAS Dependent Mechanisms - Post-transcriptional gene regulation through alternative splicing (AS) is a critical mechanism influencing cellular proliferation, differentiation, and survival. Dysregulation of AS has been identified as a key driver in multiple cancers, including breast cancer, where RNA-binding proteins (RBPs) are amplified or overexpressed in 50% of cases. Additionally, hyperactive RAS signaling is present in 50% of breast cancers, despite RAS mutations occurring in only 4% of cases. A major negative regulator of RAS signaling, neurofibromin (NF1), is frequently altered in breast cancer through sequence mutations, structural variations, or alternatively spliced transcript variants, which have been linked to poor prognosis and disease progression. While neurofibromin is well known for its role in RAS regulation, its functions in post-transcriptional gene regulation remain largely unexplored. Notably, recent findings suggest that estrogen receptor alpha (ERα) functions as a non-canonical RBP, and our laboratory has established a critical regulatory link between NF1 and ERα in breast cancer. However, the role of neurofibromin in RNA interactions, nuclear speck formation, and AS remains unknown. This project aims to define how NF1 loss or altered isoforms contribute to breast cancer progression through dysregulation of AS, RAS, and ERα signaling. Using various cell lines that are responsive to hormone stimulation, I will investigate how NF1 isoforms influence nuclear speck formation and RBP activity to promote tumor progression. Aim 1 will determine RAS- and estrogen-dependent splicing signatures associated with NF1 isoforms, identifying their impact on nuclear speck dynamics and tumor progression. Aim 2 will define how neurofibromin localization affects post-transcriptional regulation through nuclear speck function, elucidating mechanisms by which NF1 mutant cancers develop therapeutic resistance. These studies will provide a mechanistic understanding of how NF1 regulates AS and nuclear speck formation, uncovering new therapeutic vulnerabilities and prognostic biomarkers for NF1-deficient breast cancers.
