Unknown,Transcriptomics,Genomics,Proteomics

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Paraoxonase 2 Serves a Proapopotic Function in Mouse and Human Cells in Response to the Pseudomonas aeruginosa Quorum-Sensing Molecule N-(3-oxododecanoyl)-Homoserine Lactone.

ABSTRACT: Pseudomonas aeruginosa use quorum-sensing molecules, including N-(3-oxododecanoyl)-homoserine lactone (C12), for intercellular communication. C12 activated apoptosis in mouse embryo fibroblasts (MEF) from both wild type (WT) and Bax/Bak double knock-out mice (WT MEF and DKO MEF that were responsive to C12, DKORM-BM- MEF): nuclei fragmented; mitochondrial membrane potential (M-bM-^HM-^FM-OM-^Hmito) depolarized; Ca2+M-BM- was released from the endoplasmic reticulum (ER), increasing cytosolic [Ca2+] (Cacyto); caspase 3/7 was activated. DKORM-BM- MEF had been isolated from a nonclonal pool of DKO MEF that were non-responsive to C12 (DKONRM-BM- MEF). RNAseq analysis, qPCR and western blots showed that WT and DKORM-BM- MEF both expressed genes associated with cancer, including paraoxonase 2 (PON2), while DKONRM-BM- MEF expressed little PON2. Adenovirus-mediated expression of human PON2 in DKONRM-BM- MEF rendered them responsive to C12: M-bM-^HM-^FM-OM-^HmitoM-BM- depolarized, CacytoM-BM- increased and caspase 3/7 activated. Human embryonic kidney 293T (HEK293T) cells expressed low levels of endogenous PON2, and these cells were also less responsive to C12. Overexpression of PON2, but not PON2-H114Q (no lactonase activity) in HEK293T cells caused them to become sensitive to C12. Because [C12] may reach high levels in biofilms in lungs of cystic fibrosis (CF) patients, PON2 lactonase activity may control M-bM-^HM-^FM-OM-^Hmito, Ca2+M-BM- release from the ER and apoptosis in CF airway epithelia. Coupled with previous data, these results also indicate that PON2 uses its lactonase activity to prevent Bax- and Bak-dependent apoptosis in response to common proapoptotic drugs like doxorubicin, staurosporine but activates Bax- and Bak-independent apoptosis in response to C12. Gene expression profiling of mouse embryo fibroblasts from WT and Bax/Bak double knock-out mice (C12 responsive and non-reponsive cell lines).

ORGANISM(S): Mus musculus

SUBMITTER: Terry Machen

PROVIDER: E-GEOD-65935 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:MUC1 is a tumor-associated antigen that is aberrantly expressed in cancer and inflammatory bowel disease (IBD). Even though immune cells express low MUC1 levels, their modulations of MUC1 are important in tumor progression. Consistent with previous clinical data that show increased myeloid-derived suppressor cells (MDSCs) in IBD, we now show that down-regulation of MUC1 on hematopoietic cells increases MDSCs in IBD, similar to our data in tumor-bearing mice. We hypothesize that MDSC expansion in IBD is critical for tumor progression. In order to mechanistically confirm the linkage between Muc1 down-regulation and MDSC expansion, we generated chimeric mice that did not express Muc1 in the hematopoietic compartment (KOM-bM-^FM-^RWT). These mice were used in 2 models of colitis and colitis-associated cancer (CAC) and their responses were compared to wildtype chimeras (WTM-bM-^FM-^RWT). KOM-bM-^FM-^RWT mice show increased levels of MDSCs during colitis that was responsible for the larger colon tumors that eventually developed in these mice. Using microarray and qRT-PCR analysis, we observed increased pro-tumorigenic signaling in the colons of KOM-bM-^FM-^RWT mice during colitis as compared to WTM-bM-^FM-^RWT mice. Our RNA (microarray and qRT-PCR analysis) and protein analysis demonstrate increased up-regulation of metalloproteinases, collagenases, defensins, complements, growth factors, cytokines and chemokines in KOM-bM-^FM-^RWT mice as compared to WTM-bM-^FM-^RWT mice. Antibody-mediated depletion of MDSCs in mice during colitis reduced colon tumor formation during CAC. Development of CAC is a serious complication of colitis and our data highlight MDSCs as a targetable link between inflammation and cancer. A total of 12 samples were analysed. RNA was made from the mucosa of the colon of WTM-bM-^FM-^RWT and KOM-bM-^FM-^RWT mice that were either untreated or treated with azoxymethane (AOM) and dextran sodium sulfate (DSS). There are 4 groups, n=3 for each group: untreated WTM-bM-^FM-^RWT mice, untreated KOM-bM-^FM-^RWT mice, AOM+DSS treated WTM-bM-^FM-^RWT mice, AOM+DSS treated KOM-bM-^FM-^RWT mice = 12 samples in total.

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Paraoxonase 2 Serves a Proapopotic Function in Mouse and Human Cells in Response to the Pseudomonas aeruginosa Quorum-Sensing Molecule N-(3-oxododecanoyl)-Homoserine Lactone.

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