Oncogenic KRAS/ERK/JUNB signaling suppresses differentiation regulator GATA6 in pancreatic cancer
Zheng Zhong, Xinang Cao, Pei-Ju Liao, Raman Sethi, Jeffrey A. Klomp, Clint A. Stalnecker, Jinmiao Chen, Yue Wan, Channing J. Der, David M. Virshup
Zheng Zhong, Xinang Cao, Pei-Ju Liao, Raman Sethi, Jeffrey A. Klomp, Clint A. Stalnecker, Jinmiao Chen, Yue Wan, Channing J. Der, David M. Virshup
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Research Article Cell biology Oncology

Oncogenic KRAS/ERK/JUNB signaling suppresses differentiation regulator GATA6 in pancreatic cancer

Abstract

GATA6 is a master regulator of differentiation in the pancreas, and its expression levels determine the 2 main molecular subtypes of pancreatic cancer. High GATA6 levels contribute to the classical pancreatic cancer subtype, which is associated with a higher degree of tumor differentiation and better disease prognosis. However, why GATA6 expression varies across pancreatic cancers and what regulates GATA6 expression remain elusive. Here, we report that oncogenic KRAS-activated ERK signaling suppresses GATA6 transcription in pancreatic cancers. GATA6 mRNA levels inversely correlated with KRAS/ERK activity in pancreatic tumors. A genome-wide CRISPR screen in a GATA6-EGFP reporter knockin cell line identified JUNB as the ERK-regulated transcriptional repressor for GATA6. Active ERK stabilized JUNB protein, while KRAS/ERK inhibition led to ubiquitin-independent proteasomal degradation of JUNB and increased transcription of GATA6. Upregulation of GATA6 enhanced chemosensitivity of pancreatic cancer cells, and KRAS/ERK inhibitors synergized with chemotherapy in a GATA6-dependent manner. Our study identifies how oncogenic KRAS/ERK signaling suppresses GATA6 to cause dedifferentiation in pancreatic cancer. Combining KRAS/ERK inhibitors with standard-of-care chemotherapies could be a promising therapeutic strategy for treating pancreatic cancers.

Authors

Zheng Zhong, Xinang Cao, Pei-Ju Liao, Raman Sethi, Jeffrey A. Klomp, Clint A. Stalnecker, Jinmiao Chen, Yue Wan, Channing J. Der, David M. Virshup

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Figure 5

JUNB mediates ERK’s suppressive effect on GATA6 transcription.

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JUNB mediates ERK’s suppressive effect on GATA6 transcription. (A) Knock...
(A) Knockdown of JUNB increased GATA6 transcript abundance. HPAF-II cells stably expressing dCas9-KRAB fusion protein were transduced with a lentivirus expressing a dox-inducible nontargeting control sgRNA (sgNTC#2) or sgRNAs targeting the JUNB promoter (sgJUNB#1 and sgJUNB#2). Cells were treated with dox for 6 days and analyzed by RT-qPCR. n = 2 biological replicates × 2 technical replicates/condition. (B) Overexpression of JUNB downregulated GATA6 transcript abundance. HPAF-II cells stably expressing mCherry control or JUNB were analyzed by RT-qPCR. n = 2 biological replicates × 2 technical replicates/condition. (C) Pancreatic tumors with high JUNB protein abundance showed lower levels of GATA6 transcript. JUNB protein abundance and GATA6 mRNA abundance data of human pancreatic tumors in the CPTAC cohort (39) were extracted from the cBioPortal website (85). Each dot represents an individual tumor, and mean values of each group are shown. n = 70/group. P value of 2-tailed, unpaired t test is shown. (D) Knockout of JUNB in Panc08.13 cells mimicked the effect of MEK/ERK inhibition on GATA6 expression. Panc08.13 cells were transduced with lentivirus expressing Cas9 and a nontargeting control sgRNA (sgNTC#1) or a sgRNA targeting JUNB (sgJUNB#3 or sgJUNB#4). After puromycin selection, these genome-edited cell pools were treated with DMSO or 100 nM trametinib for 24 h, followed by Western blot analysis. n = 2 biological replicates. (E) JUNB knockdown in HPAF-II cells mimicked the effect of MEK/ERK inhibition on GATA6 expression. Cells from (A) were treated with DMSO or 100 nM trametinib for 24 h, followed by Western blot analysis. n = 2 biological replicates.