TCF21 hypermethylation regulates renal tumor cell clonogenic proliferation and migration

Abstract

We recently identified hypermethylation at the gene promoter of transcription factor 21 (TCF21) in clear cell sarcoma of the kidney (CCSK), a rare pediatric renal tumor. TCF21 is a transcription factor involved in tubular epithelial development of the kidney and is a candidate tumor suppressor. As there are no in vitro models of CCSK, we employed a well‐established clear cell renal cell carcinoma (ccRCC) cell line, 786‐O, which also manifests high methylation at the TCF21 promoter, with consequent low TCF21 expression. The tumor suppressor function of TCF21 has not been functionally addressed in ccRCC cells; we aimed to explore the functional potential of TCF21 expression in ccRCC cells in vitro. 786‐O clones stably transfected with either pBABE‐TCF21‐HA construct or pBABE vector alone were functionally analyzed. We found that ectopic expression of TCF21 in 786‐O cells results in a trend toward decreased cell proliferation (not significant) and significantly decreased migration compared with mock‐transfected 786‐O cells. Although the number of colonies established in colony formation assays was not different between 786‐O clones, colony size was significantly reduced in 786‐O cells expressing TCF21. To investigate whether the changes in migration were due to epithelial‐to‐mesenchymal transition changes, we interrogated the expression of selected epithelial and mesenchymal markers. Although we observed upregulation of mRNA and protein levels of epithelial marker E‐cadherin in clones overexpressing TCF21, this did not result in surface expression of E‐cadherin as measured by fluorescence‐activated cell sorting and immunofluorescence. Furthermore, mRNA expression of the mesenchymal markers vimentin (VIM) and SNAI1 was not significantly decreased in TCF21‐expressing 786‐O cells, while protein levels of VIM were markedly decreased. We conclude that re‐expression of TCF21 in renal cancer cells that have silenced their endogenous TCF21 locus through hypermethylation results in reduced clonogenic proliferation, reduced migration, and reduced mesenchymal‐like characteristics, suggesting a tumor suppressor function for transcription factor 21.

Multiple biological functions of transcription factor 21 in the development of various cancers

“…However, another study found no changes in mRNA levels of the mesenchymal markers such as VIM and SNAI1, but there was a significant upregulation of the epithelial marker E-cadherin at both the mRNA and the protein levels in renal tumor cells expressing ectopic TCF21 . 61 These results also suggested that E-cadherin might be a proximate downstream target of TCF21. E-box elements of the E-cadherin promoter are involved in silencing E-cadherin promoter activity in cancer cells.…”

Transcription Factor 21 Is Required for Branching Morphogenesis and Regulates the Gdnf-Axis in Kidney Development

“…[15][16][17][18] In disease, Tcf21 is reported to function as a tumor suppressor and loss of Tcf21 expression and/or methylation of its promoter have been identified in a variety of lung and urogenital tumors. 19,20 In other studies, Tcf21 has been linked to coronary artery disease, 21,22 podocyte differentiation, diabetic kidney disease, 23 and changes in adipose phenotype. 24 In the kidney, previous work has shown that Tcf21 null mutant mice are born with severely hypoplastic kidneys, and that although Tcf21 is exclusively expressed in the mesenchyme, in its absence, major defects in UB epithelial differentiation and branching are observed.…”

CircRNA hsa_circ_100395 regulates miR-1228/TCF21 pathway to inhibit lung cancer progression

“…Moreover, we showed hsa_-circ_100395 promoted TCF21 expression via sponging miR-1228 in lung cancer. TCF21 has been acknowledged as a tumor suppressor in renal tumor [30], ovarian cancer [31], colorectal cancer [32] and lung cancer [33]. However, how TCF21 expression was regulated in lung cancer remains elusive.…”