Nuclear protein 1 promotes pancreatic cancer development and protects cells from stress by inhibiting apoptosis
Tewfik Hamidi, Hana Algül, Carla Eliana Cano, Maria José Sandi, Maria Inés Molejon, Marc Riemann, Ezequiel Luis Calvo, Gwen Lomberk, Jean-Charles Dagorn, Falk Weih, Raul Urrutia, Roland Michael Schmid, Juan Lucio Iovanna
Tewfik Hamidi, Hana Algül, Carla Eliana Cano, Maria José Sandi, Maria Inés Molejon, Marc Riemann, Ezequiel Luis Calvo, Gwen Lomberk, Jean-Charles Dagorn, Falk Weih, Raul Urrutia, Roland Michael Schmid, Juan Lucio Iovanna
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Research Article Oncology

Nuclear protein 1 promotes pancreatic cancer development and protects cells from stress by inhibiting apoptosis

Abstract

Pancreatic ductal adenocarcinoma (PDAC) has the lowest survival rate of all cancers and shows remarkable resistance to cell stress. Nuclear protein 1 (Nupr1), which mediates stress response in the pancreas, is frequently upregulated in pancreatic cancer. Here, we report that Nupr1 plays an essential role in pancreatic tumorigenesis. In a mouse model of pancreatic cancer with constitutively expressed oncogenic KrasG12D, we found that loss of Nupr1 protected from the development of pancreatic intraepithelial neoplasias (PanINs). Further, in cultured pancreatic cells, nutrient deprivation activated Nupr1 expression, which we found to be required for cell survival. We found that Nupr1 protected cells from stress-induced death by inhibiting apoptosis through a pathway dependent on transcription factor RelB and immediate early response 3 (IER3). NUPR1, RELB, and IER3 proteins were coexpressed in mouse PanINs from KrasG12D-expressing pancreas. Moreover, pancreas-specific deletion of Relb in a KrasG12D background resulted in delayed in PanIN development associated with a lack of IER3 expression. Thus, efficient PanIN formation was dependent on the expression of Nupr1 and Relb, with likely involvement of IER3. Finally, in patients with PDAC, expression of NUPR1, RELB, and IER3 was significantly correlated with a poor prognosis. Cumulatively, these results reveal a NUPR1/RELB/IER3 stress-related pathway that is required for oncogenic KrasG12D-dependent transformation of the pancreas.

Authors

Tewfik Hamidi, Hana Algül, Carla Eliana Cano, Maria José Sandi, Maria Inés Molejon, Marc Riemann, Ezequiel Luis Calvo, Gwen Lomberk, Jean-Charles Dagorn, Falk Weih, Raul Urrutia, Roland Michael Schmid, Juan Lucio Iovanna

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Figure 3

Nupr1 activates RelB expression to promote cell survival upon stress.

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Nupr1 activates RelB expression to promote cell survival upon stress. Mi...
MiaPaCa2 cells were transfected with the indicated siRNAs and cultured in conventional or EBSS medium for the times indicated. (A) Heat map showing relative expression of NF-κB family genes and targets. (B and C) qRT-PCR and Western blot showing increased RelB mRNA and protein expression, respectively, in Nupr1-depleted cells compared with controls. Cyclophilin and tubulin were housekeeping controls for mRNA and protein load, respectively. (D) Cells were transfected with pCDNA3 vectors containing a Flag-tagged Nupr1, an irrelevant cytochrome c (Cyto C), or an empty vector (Empty). ChIP was performed using an anti-Flag antibody or a irrelevant IgG. Top: Occupancy of Nupr1 on the RELB promoter; bottom: DNA input (10%). (E) Cells were transfected with combinations of a RELB promoter-Luc vector, siCtrl, siNupr1, Nupr1-Flag, or Empty-Flag pCDNA3 vector, and pSV40-RL as indicated. After 9 hours, luciferase activity was determined and expressed as the ratio of specific luciferase activity to an internal standard. (F) RelB expression was measured by qRT-PCR (right) and Western blot (left). (G and H) Cell viability and caspase-3/7 activity were measured after 24 hours as described in Figure 2. (I) NUPR1 mRNA was measured after 24 hours by qRT-PCR. In E and I, values are expressed as mean ± SEM of triplicate, from 2 independent experiments. *P ≤ 0.05 relative to siCtrl-transfected cells cultured in conventional medium; †P ≤ 0.05 relative to siNupr1-transfected cells.