Format

Send to

Choose Destination
Cell Mol Gastroenterol Hepatol. 2018 May 16;6(2):181-198. doi: 10.1016/j.jcmgh.2018.05.002. eCollection 2018.

E47 Governs the MYC-CDKN1B/p27KIP1-RB Network to Growth Arrest PDA Cells Independent of CDKN2A/p16INK4A and Wild-Type p53.

Author information

1
Development, Aging and Regeneration Program, Sanford Burnham Prebys Medical Discovery Institute, La Jolla, California.
2
Center for Computational Biology and Bioinformatics, School of Medicine.
3
Department of Surgery, Division of Surgical Oncology, Moores Cancer Center.
4
Genomics Core, Sanford Burnham Prebys Medical Discovery Institute, La Jolla, California.
5
Department of Pediatrics, University of California San Diego, La Jolla, California.

Abstract

Background & Aims:

Oncogenic mutations in KRAS, coupled with inactivation of p53, CDKN2A/p16INK4A, and SMAD4, drive progression of pancreatic ductal adenocarcinoma (PDA). Overexpression of MYC and deregulation of retinoblastoma (RB) further promote cell proliferation and make identifying a means to therapeutically alter cell-cycle control pathways in PDA a significant challenge. We previously showed that the basic helix-loop-helix transcription factor E47 induced stable growth arrest in PDA cells in vitro and in vivo. Here, we identified molecular mechanisms that underlie E47-induced growth arrest in low-passage, patient-derived primary and established PDA cell lines.

Methods:

RNA sequencing was used to profile E47-dependent transcriptomes in 5 PDA cell lines. Gene Ontology analysis identified cell-cycle control as the most altered pathway. Small interfering RNA/short hairpin RNA knockdown, small-molecule inhibitors, and viral expression were used to examine the function of E47-dependent genes in cell-cycle arrest. Cell morphology, expression of molecular markers, and senescence-associated β-galactosidase activity assays identified cellular senescence.

Results:

E47 uniformly inhibited PDA cell-cycle progression by decreasing expression of MYC, increasing the level of CDKN1B/p27KIP1, and restoring RB tumor-suppressor function. The molecular mechanisms by which E47 elicited these changes included altering both RNA transcript levels and protein stability of MYC and CDKN1B/p27KIP1. At the cellular level, E47 elicited a senescence-like phenotype characterized by increased senescence-associated β-galactosidase activity and altered expression of senescence markers.

Conclusions:

E47 governs a highly conserved network of cell-cycle control genes, including MYC, CDKN1B/p27KIP1, and RB, which can induce a senescence-like program in PDA cells that lack CDKN2A/p16INK4A and wild-type p53. RNA sequencing data are available at the National Center for Biotechnology Information GEO at https://www.ncbi.nlm.nih.gov/geo/; accession number: GSE100327.

KEYWORDS:

CDK, cyclin-dependent kinase; CDKN1B/p27KIP1, CDKN1B/p27Kinase Inhibitory Protein 1; CDKN2A/p16INK4A, CDKN2A/p16Inhibitor of CDK 4A; CEBP-α, CCAAT/enhancer binding protein alpha; CENP-A, centromere protein A; CIP, Cyclin-Dependent Kinase Inhibitor 1; Cell Cycle; DDR, DNA damage response; ERK, extracellular signal–regulated kinase; GO, Gene Ontology; INK, Inhibitor of CDK; KIP, Kinase Inhibitory Protein; MSCV, murine stem cell virus; OIS, oncogene-induced senescence; PCR, polymerase chain reaction; PDA, pancreatic ductal adenocarcinoma; Pancreatic Ductal Adenocarcinoma; RB, retinoblastoma; RNA-seq, RNA sequencing; SA-βgal, senescence-associated β-galactosidase; SKP, S-phase Kinase-associated; Senescence; bHLH; bHLH, basic helix-loop-helix; lfdr, local false discovery rate; mRNA, messenger RNA; shRB, short hairpin RNA directed against RB; shRNA, short hairpin RNA; si-p27, small interfering RNA directed against p27

Comment in

Supplemental Content

Full text links

Icon for Elsevier Science Icon for PubMed Central
Loading ...
Support Center