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SMAD proteins directly suppress PAX2 transcription downstream of transforming growth factor-beta 1 (TGF-?1) signalling in renal cell carcinoma.

ABSTRACT: Canonical TGF-?1 signalling promotes tumor progression by facilitating invasion and metastasis, whereby release of TGF-?1, by (for example) infiltrating immune cells, induces epithelial to mesenchymal transition (EMT). PAX2, a member of the Paired box family of transcriptional regulators, is normally expressed during embryonic development, including in the kidney, where it promotes mesenchymal to epithelial transition (MET). PAX2 expression is silenced in many normal adult tissues. However, in contrast, PAX2 is expressed in several cancer types, including kidney, prostate, breast, and ovarian cancer. While multiple studies have implicated TGF-? superfamily members in modulating expression of Pax genes during embryonic development, few have investigated direct regulation of Pax gene expression by TGF-?1. Here we have investigated direct regulation of PAX2 expression by TGF-?1 in clear cell renal cell carcinoma (CC-RCC) cell lines. Treatment of PAX2-expressing 786-O and A498 CC-RCC cell lines with TGF-?1 resulted in inhibition of endogenous PAX2 mRNA and protein expression, as well as expression from transiently transfected PAX2 promoter constructs; this inhibition was abolished in the presence of expression of the inhibitory SMAD, SMAD7. Using ChIP-PCR we showed TGF-?1 treatment induced SMAD3 protein phosphorylation in 786-O cells, and direct SMAD3 binding to the human PAX2 promoter, which was inhibited by SMAD7 over-expression. Overall, these data suggest that canonical TGF-? signalling suppresses PAX2 transcription in CC-RCC cells due to the direct binding of SMAD proteins to the PAX2 promoter. These studies improve our understanding of tumor progression and epithelial to mesenchyme transition (EMT) in CC-RCC and in other PAX2-expressing cancer types.

SUBMITTER: Kaur G

PROVIDER: S-EPMC6003550 | biostudies-literature | 2018 Jun

REPOSITORIES: biostudies-literature

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SMAD proteins directly suppress <i>PAX2</i> transcription downstream of transforming growth factor-beta 1 (TGF-β1) signalling in renal cell carcinoma.

Kaur Gagandeep G Li Caiyun Grace CG Chantry Andrew A Stayner Cherie C Horsfield Julia J Eccles Michael R MR

Oncotarget 20180601 42

Canonical TGF-β1 signalling promotes tumor progression by facilitating invasion and metastasis, whereby release of TGF-β1, by (for example) infiltrating immune cells, induces epithelial to mesenchymal transition (EMT). PAX2, a member of the Paired box family of transcriptional regulators, is normally expressed during embryonic development, including in the kidney, where it promotes mesenchymal to epithelial transition (MET). PAX2 expression is silenced in many normal adult tissues. However, in c ...[more]

PMID: 29928489

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Project description:Caffeine administration is an important part of the therapeutic treatment of bronchopulmonary dysplasia (BPD) in preterm infants. However, caffeine mediated effects on airway remodelling are still undefined. The TGF-β/Smad signalling pathway is one of the key pathways involved in airway remodelling. Connective tissue growth factor (CTGF), a downstream mediator of TGF-β, and transgelin, a binding and stabilising protein of the cytoskeleton, are both regulated by TGF-β1 and play an important role in airway remodelling. Both have also been implicated in the pathogenesis of BPD. The aim of the present study was to clarify whether caffeine, an unspecific phosphodiesterase (PDE) inhibitor, and rolipram, a prototypical PDE-4 selective inhibitor, were both able to affect TGF-β1-induced Smad signalling and CTGF/transgelin expression in lung epithelial cells. Furthermore, the effect of transgelin knock-down on Smad signalling was studied. The pharmacological effect of caffeine and rolipram on Smad signalling was investigated by means of a luciferase assay via transfection of a TGF-β1-inducible reporter plasmid in A549 cells. The regulation of CTGF and transgelin expression by caffeine and rolipram were studied by promoter analysis, real-time PCR and Western blot. Endogenous transgelin expression was down-regulated by lentiviral transduction mediating transgelin-specific shRNA expression. The addition of caffeine and rolipram inhibited TGF-β1 induced reporter gene activity in a concentration-related manner. They also antagonized the TGF-β1 induced up-regulation of CTGF and transgelin on the promoter-, the mRNA-, and the protein-level. Functional analysis showed that transgelin silencing reduced TGF-β1 induced Smad-signalling and CTGF induction in lung epithelial cells. The present study highlights possible new molecular mechanisms of caffeine and rolipram including an inhibition of Smad signalling and of TGF-β1 regulated genes involved in airway remodelling. An understanding of these mechanisms might help to explain the protective effects of caffeine in prevention of BPD and suggests rolipram to be a potent replacement for caffeine.

Dataset Information

SMAD proteins directly suppress PAX2 transcription downstream of transforming growth factor-beta 1 (TGF-?1) signalling in renal cell carcinoma.

Publications

SMAD proteins directly suppress <i>PAX2</i> transcription downstream of transforming growth factor-beta 1 (TGF-β1) signalling in renal cell carcinoma.

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