Targeting TUT1 Depletes Tri-snRNP Pools to Suppress Splicing and Inhibit Pancreatic Cancer Cell Survival

Abstract Pancreatic ductal adenocarcinoma (PDAC) is highly aggressive and lacks effective therapeutic options. Cancer cells frequently become more dependent on splicing factors than normal cells due to increased rates of transcription. Terminal uridylyltransferase 1 (TUT1) is a specific terminal uridylyltransferase for U6 small nuclear RNA (snRNA), which plays a catalytic role in the spliceosome. In this study, we found that TUT1 was required for the survival of PDAC cells but not for normal pancreatic cells. In PDAC cells, the uridylylation activity of TUT1 promoted U4/U6.U5 tri-small nuclear ribonucleoprotein particles (snRNP) assembly by facilitating the binding of LSM proteins to U6 snRNA and subsequent tri-snRNP assembly. PDAC cells required higher amounts of U4/U6.U5 tri-snRNP to efficiently splice pre-mRNA with weak splice sites to support the high transcriptional output. Depletion of TUT1 in PDAC cells resulted in inefficient splicing of exons in a group of highly expressed RNAs containing weak splice sites, thereby resulting in the collapse of an mRNA processing circuit and consequently dysregulating splicing required by PDAC cells. Overall, this study unveiled an interesting function of TUT1 in regulating splicing by modulating U4/U6.U5 tri-snRNP levels and demonstrated a distinct mechanism underlying splicing addiction in pancreatic cancer cells. Significance: The higher amounts of U6 snRNA in tri-snRNP pools in pancreatic cancer cells compared with normal cells confers sensitivity to TUT1 inhibition, which mimics tri-snRNP inhibition and causes pancreatic cancer cell senescence.