Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Differential gene expression in Fosl1 siRNA vs Control siRNA treated murine pancreatic cancer cells

ABSTRACT: A subset of human pancreatic ductal adenocarcinoma cells (PDACs) is characterized by high Fosl1 expression and Fosl1 is linked to the control of pro-inflammatory pathways and growth of PDAC cells. To mimick the human disease in mice (> 90% of PDAC patients harbour Kras mutations) the mutated LSL-KrasG12D allele was combined with the pancreas specific Cre recombinase Ptf1aCre (p48Cre). The two pancreatic cancer cell lines (Ptf1aCre, LSL-KrasG12D/+) were isolated from these mice and used for transcriptomics studies. The two different murine pancreatic cancer cell lines (Ptf1aCre, LSL-KrasG12D) were treated with two different Fosl1 siRNAs and one control siRNA, each. 72h after transfection a sufficient knockdown was tested by immunoblotting and qPCR. Total mRNA was isolated and checked for integrity. According to manufacture's recommendation the samples were subjected to microarray analysis using the Affymetrix Mouse Gene ST 1.0 array chip to discover differentially expressed genes.

ORGANISM(S): Mus musculus

SUBMITTER: Matthias Wirth

PROVIDER: E-MTAB-4299 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

ACCESS DATA

Similar Datasets

Project description:The objective of this study was to elucidate the role of Nupr1 in pancreatic tumorigenesis. Using the Pdx-1-cre;LSL-KrasG12D mouse as model we discovered that, in contrast to KrasG12D pancreas that develop multiple foci of pancreatic intraepithelial neoplasia (PanIN), KrasG12D;Nupr1KO pancreas were free from such lesions, indicating that Nupr1 is pivotal for PanIN formation. In vitro, MiaPaCa2 cells activated Nupr1 expression in response to nutrient deprivation and this expression was necessary for cell survival. Mechanistically, Nupr1 protected cells from stress-induced death by inhibiting apoptosis through an alternative RelBàIER3-dependent pathway and independent from activation of the classical RelA-based NF-kB pathway. In agreement with these findings, Nupr1, RelB and IER3 proteins were found co-expressed in PanINs from KrasG12D pancreas. Moreover, pancreas-specific KrasG12D;RelbDpanc mice displayed a delay in PanIN development associated with a lack of IER3 expression, further emphasizing the relevance of this pathway in vivo. Efficient PanIN formation was therefore dependent on the expression of Nupr1 and RelB, with the probable involvement of IER3. Finally, a significant correlation between expression of Nupr1, RelB and IER3 and a poor prognosis of patients with PDAC was found. Altogether, our results reveal a novel stress-related pathway that requires the functional interaction of Nupr1àRelBàIER3 in KrasG12D-dependent transformation of the pancreas and expand our understanding of the molecular machinery that mediates the early steps of pancreatic carcinogenesis. Since Nupr1 belongs to the HMG family of chromatin remodelers with transcriptional co-factor activity, Nupr1 could increase cell survival in a nutrient-deprived microenvironment by activating the expression of pro-survival genes. Moreover, considering that RelB, and not RelA/p65, is essential to the Nupr1-mediated survival mechanism that takes place upon nutrient deprivation-induced stress, we made the hypothesis that the two NF-kB transcription factors should activate different sets of genes among which a pivotal pro-survival gene would be exclusively dependent on RelB. In order to test this two hypothesis, an Affymetrix microarray analysis was performed using pancreatic cancer cells transfected with siCtrl or siNupr1 and cultured for 3, 6 or 9 hrs in EBSS; or transfected with siRelB, siRelA/p65 or siCtrl and cultured for 9 hrs in EBSS or Mock.

Project description:The maintenance of advanced malignancies relies on continued activity of driver oncogenes, although their rate-limiting role is highly context-dependent with respect to tumor types and associated genetic alterations. Oncogenic Kras mutation is the signature event in human pancreatic ductal adenocarcinoma (PDAC), serving a critical role in tumor initiation. Here, an inducible KrasG12D-driven p53 mutant PDAC mouse model establishes that advanced PDAC remains strictly dependent on continued KrasG12D expression and that KrasG12D serves a vital role in the control of tumor metabolism, through stimulation of glucose uptake and channeling of glucose intermediates through the hexosamine biosynthesis pathway (HBP) and the pentose phosphate pathway (PPP). Notably, these studies reveal that oncogenic Kras regulates ribose biogenesis. Unlike canonical models of PPP-mediated ribose biogenesis, we demonstrate that oncogenic Kras drives intermediates from enhanced glycolytic flux into the non-oxidative arm of the PPP, thereby decoupling ribose biogenesis from NADPNADPH-mediated redox control. Together, this work provides in vivo mechanistic insights into how oncogenic Kras promotes metabolic reprogramming in native tumors and illuminates potential metabolic targets that can be exploited for therapeutic benefit in Kras-driven PDAC. Primary pancreatic tumor lines were established from p48Cre tetO_LKrasG12D ROSA_rtTAL+ p53L+ mice. Five independent tumor lines (iKras1-5) were used for pancreatic injection into nude mice to generate orthotopic tumors. The mice were kept on doxycycline for 2 weeks until obvious tumor formation. Half of the animals were pulled off doxycycline for 24 hours. Tumors with over 75% cellularity were collected for total RNA prepartion. For in vitro expression profiles, the same five tumor lines were cultured in the presence or absence of doxycycline for 24 hours and total cellular RNA was prepared. For control samples, two independent tumor lines from LSL-KrasG12D p53L+ tumors were cultured in the presence or absence of doxycycline for 24 hours and total cellular RNA was prepared.

Dataset Information

Differential gene expression in Fosl1 siRNA vs Control siRNA treated murine pancreatic cancer cells

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets