Abstract Oncogenic KRAS is a central driver of pancreatic ductal adenocarcinoma (PDAC), primarily through maintenance of ERK signaling. However, activation of the KRAS-MEK-ERK through a KRAS-independent manner is not fully understood. Here, we identify one alternative mechanism of ERK activation mediated by the RNA-binding protein survival motor neuron domain containing protein 1 (SMNDC1), which regulates splicing of ERK1 (MAPK3). SMNDC1 is amplified or overexpressed in ∼30% of PDACs and is associated with decreased survival in both human and murine models. Gain- and loss-of-function studies revealed that SMNDC1 promotes tumor growth in part by driving inclusion of MAPK3-E4, which encodes the kinase-activating phosphorylation sites Thr202/Tyr204. Use of an ERK kinase biosensor (ERK-KTR), which translocates to the nucleus upon ERK inactivation, confirmed that E4 inclusion is critical for maintaining downstream ERK activity. Conversely, forced exclusion of MAPK3-E4 using antisense oligonucleotides (ASOs) diminished ERK signaling, despite the continued presence of wild-type ERK2, underscoring the functional significance of the ERK1 splice isoform. To examine this regulation at the tissue and cell level, we performed BaseScope in situ hybridization on a tissue microarray comprising over 200 PDAC patients. While expression of MAPK3 transcripts with and without E4 was broadly correlated, we observed a consistent shift toward E4 inclusion in tumor samples, indicating selective pressure for the active isoform in PDAC. Mechanistically, high-throughput RNA Bind-n-Seq (RBNS) and enhanced CLIP (eCLIP) in PDAC cells revealed that SMNDC1 binds a C-rich polypyrimidine tract at the distal 3′ splice site of MAPK3-E4. Mutational analysis demonstrated that the conserved Tudor domain of SMNDC1 is required for this RNA interaction and for E4 splicing. Additionally, we identified a feedback mechanism whereby inclusion of a conserved poison exon in SMNDC1 triggers its own degradation, suggesting tight post-transcriptional control of this splicing regulator. Altogether, these findings define a sequence- and domain-specific splicing switch that enhances ERK activity through MAPK3-E4 inclusion, providing a KRAS-independent mechanism of MAPK pathway activation and a potential therapeutic vulnerability in PDAC. Citation Format: Deanne E. Yugawa, Md Afjalus Siraj, Muaz Faruque, Gilbert Giri, Grant Goda, Daniel Dominguez, Pablo Perez-Pinera, Resham Bhattacharya, Priyabrata Mukherjee, Luisa F. Escobar-Hoyos. Tightly regulated alternative RNA splicing of MAPK3 (ERK1) as a Kras-independent mechanism of oncogenesis in PDAC abstract. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research—Emerging Science Driving Transformative Solutions; Boston, MA; 2025 Sep 28-Oct 1; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2025;85 (18Suppl₃): Abstract nr A060.
Yugawa et al. (Sun,) studied this question.