KRAS-mediated upregulation of CIP2A promotes suppression of PP2A-B56a to initiate pancreatic cancer development [ATAC-seq]

Oncogenic mutations in KRAS are present in approximately 95% of patients diagnosed with pancreatic ductal adenocarcinoma (PDAC) and are considered the initiating event during the development of pancreatic intraepithelial neoplasia (PanIN) precursor lesions. While it is well established that KRAS mutations can drive the initiation of pancreatic oncogenesis, the effects of oncogenic KRAS signaling on regulation of phosphatases during this process is not fully appreciated. Protein Phosphatase 2A (PP2A) has been implicated in suppressing KRAS-driven cellular transformation. However, low PP2A activity is observed in PDAC cells compared to non-transformed cells, suggesting that suppression of PP2A activity is an important step in the overall development of PDAC. In the current study, we demonstrate that KRASG12D induces the expression of both Cancerous Inhibitor of PP2A (CIP2A), an endogenous inhibitor of PP2A activity, and the PP2A target, c-MYC. Consistent with these findings, KRASG12D sequestered the specific PP2A subunit responsible for c-MYC degradation, B56a, away from the active PP2A holoenzyme in a CIP2A-dependent manner. During PDAC initiation in vivo, knockout of B56a promoted KRASG12D tumorigenesis by accelerating acinar-to-ductal metaplasia (ADM) and the formation of PanIN lesions. The process of ADM was attenuated ex vivo in response to pharmacological re-activation of PP2A utilizing direct small molecule activators of PP2A (SMAPs). Together, our results suggest that suppression of PP2A-B56a through KRAS signaling can promote Myc-driven initiation of pancreatic tumorigenesis. Overall design: 5-6W old PDX1-Cre; LSL-KRASG12D (KC) mice or KC mice with B56a loss (KCB) were used for experiment. Pancreata were harvested and then isolated using the Fleming-Martinez and Storz JOVE 2019 protocol. Isolated acinar clusters were plated in a collagen matrix and cultured at 37*C and 5% O2 for 24, 48, or 72H. Structures were digested out of collagen using collagenase, washed in PBS, and then resulting cell pellets after final centrifgation were resuspended in 10%DMSO/90%FBS and stored at -80*C until used for ATAC-seq processing.