TCEAL7, a putative tumor suppressor gene, negatively regulates NF-kappaB pathway.
We have previously shown that a frequently downregulated gene, transcription elongation factor A-like 7 (TCEAL7), promoted anchorage-independent growth and modulated Myc activity in ovarian surface epithelial cells immortalized with temperature-sensitive large T antigen and human telomerase reverse transcriptase (OSEtsT/hTERT). Analysis of protein/DNA array showed that TCEAL7 downregulation resulted in an approximately twofold increase in nuclear factor (NF)-kappaB binding to its target DNA sequence. In this study we showed that short hairpin RNA (shRNA)-mediated downregulation of TCEAL7 in two different immortalized OSE cells showed higher NF-kappaB activity, as determined using reporter and gel-shift assays. Transient transfection of TCEAL7 inhibited the activation of NF-kappaB in TCEAL7-downregulated clones, IOSE-523 and in other ovarian cancer cell lines (OVCAR8, SKOV3ip and DOV13), suggesting that TCEAL7 negatively regulates NF-kappaB pathway. Consistent with this observation, TCEAL7-downregulated clones showed higher levels of NF-kappaB targets, such as pro-proliferative (cyclin-D1 and cMyc), pro-angiogenic (interleukin (IL)-6, IL-8 and vascular endothelial growth factor (VEGF)), inflammatory (intercellular adhesion molecule 1 (ICAM-1) and cyclooxygenase-2 (Cox-2)) and anti-apoptotic (B-cell lymphoma-extra large (Bcl-xl)) genes when compared with vector controls. Inhibition of NF-kappaB by IkappaB kinase (IKK) inhibitor (BMS 345541) attenuated cell survival and proliferation of TCEAL-knockdown clones. Although TCEAL7 inhibited p65 transcriptional activity, it did not modulate the cytoplasmic signaling of the NF-kappaB pathway, by itself or by tumor necrosis factor-alpha (TNF-alpha). Chromatin immunoprecipitation (ChIP) assays revealed increased recruitment of p65 and p300 to the promoters of IL-8 and IL-6 in TCEAL7-downregulated clones. Collectively, these results indicate a novel role for TCEAL7 in the negative regulation of NF-kappaB signaling at the basal level by modulating transcriptional activity of NF-kappaB on its target gene promoters, potentially providing a novel mechanism by which NF-kappaB activity may be deregulated in ovarian cancer cells.
Rattan
R
R
Department of Experimental Pathology, Mayo Clinic College of Medicine, Rochester, MN, USA.
Narita
K
K
Chien
J
J
Maguire
J L
JL
Shridhar
R
R
Giri
S
S
Shridhar
V
V
D045744:Cell Line, Tumor; D051546:Cyclooxygenase 2 / Q000235:genetics / Q000378:metabolism; D015536:Down-Regulation; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D016147:Genes, Tumor Suppressor / Q000502:physiology*; D006801:Humans; D020672:I-kappa B Proteins / Q000494:pharmacology*; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D010051:Ovarian Neoplasms / Q000378:metabolism / Q000473:pathology*; D014157:Transcription Factors / Q000502:physiology; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism*
D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D009687:Nuclear Proteins; C503109:TCEAL7 protein, human; D014157:Transcription Factors; D014409:Tumor Necrosis Factor-alpha; D000072000:NF-KappaB Inhibitor alpha; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human