Journal
CELLULAR SIGNALLING
Volume 25, Issue 5, Pages 1136-1148Publisher
ELSEVIER SCIENCE INC
DOI: 10.1016/j.cellsig.2013.01.026
Keywords
mTOR; PMT; CTGF; TGF beta; G alpha(q/11)
Categories
Funding
- NIH, NHLBI
- NIH/NIAID [A1038396]
- NATIONAL HEART, LUNG, AND BLOOD INSTITUTE [ZIAHL006069] Funding Source: NIH RePORTER
- NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES [R01AI038396] Funding Source: NIH RePORTER
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Pasteurella multocida toxin (PMT) is a mitogenic protein that hijacks cellular signal transduction pathways via deamidation of heterotrimeric G proteins. We previously showed that rPMT activates mTOR signaling via a G alpha(q/11)/PLC beta/PKC mediated pathway, leading in part to cell proliferation and migration. Herein, we show that mTOR and MAPK, but not membrane-associated tyrosine kinases, are activated in serum-starved 3T3 cells by an autocrine/paracrine substance(s) secreted into the conditioned medium following rPMT treatment. Surprisingly, this diffusible factor(s) is capable of activating mTOR and MAPK pathways even in MEF G alpha(q/11) double knockout cells. Microarray analysis identified connective tissue growth factor (CTGF) mRNA as the most upregulated gene in rPMT-treated serum-starved 3T3 cells relative to untreated cells. These results were further confirmed using RT-PCR and Western blot analyses. In accord with rPMT-induced mTOR activation, upregulation of CTGF protein was observed in WT MEF, but not in G alpha(q/11) double knockout MEF cells. Although CTGF expression is regulated by TGF beta, rPMT did not activate TGF beta pathway. In addition, MEK inhibitors U0126 or PD98059, but not mTOR specific inhibitors, rapamycin and Torin 1, inhibited rPMT-induced upregulation of CTGF. Importantly, CTGF overexpression in serum-starved 3T3 cells using adenovirus led to phosphorylation of ribosomal protein S6, a downstream target of mTOR. However, despite the ability of CTGF to activate the mTOR pathway, upregulation of CTGF alone could not induce morphological changes as those observed in rPMT-treated cells. Our findings reveal that CTGF plays an important role, but there are additional factors involved in the mitogenic action of PMT.
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