TCF21 (gene)

The TCF21 gene resides on chromosome 6 at the band 6q23.2 and includes 3 exons.[7] These three exons are associated with CpG islands CGI1, CGI2, and CGI3. DNA methylation analysis revealed hypermethylation at CGI1 and CGI3, but not CGI2 in samples from various cancer tissues.[16] Luciferase reporter assays with constructs covering CGI3 sequences in sense and antisense orientation demonstrated that CGI3 harbors a promoter that directs the synthesis of a previously unknown long non-coding RNAs (lncRNAs) in antisense orientation to TCF21. This lncRNA has been named TARID (for TCF21 antisense RNA inducing demethylation).[17]

TCF21 is a member of the bHLH (basic helix-loop-helix) family of transcription factors.[18] This protein is predicted to span 179 amino acid residues and contains a bHLH domain and an arginine-rich sequence that may facilitate DNA binding.[19]

Since the identification of Tcf21 significance in various cell lineages, further research has expanded understanding of the essential roles of this gene.

TCF21 is essential for regulating properties of the mesenchyme that are critically important for several aspects of lung and kidney morphogenesis. Null TCF21 mutant mice are born but die shortly after due to severely hypoplastic lungs and kidneys that lack alveoli and mature glomeruli.[10] While TCF21 is exclusively expressed in the mesenchyme and podocytes, major defects are observed in adjacent epithelia of TCF21 mutant mice. In the kidney, TCF21 is required for conversion of condensing mesenchyme to epithelium of the nephron, branching morphogenesis and terminal differentiation of tubular epithelium. In the lung, TCF21 is required to correctly pattern the proximodistal axis of airway epithelium and for normal branching to occur.[10]

TCF21 null mice also fail to form a spleen, as TCF21 acts after splenic specification to control morphogenetic expansion of the splenic anlage and in its absence, splenic precursor cells undergo apoptosis.[11] Since this splenic phenotype resembles that of mice lacking the homeobox genes Hox11 and Bapx1, it is possible that TCF21, together with Hox11, and Bapx1 control a common essential early step in the developmental pathway for spleen organogenesis.[11]

TCF21 is essential for cardiac fibroblast cell fate, as demonstrated by the failed development of cardiac fibroblasts in mice lacking TCF21.[9] In the absence of TCF21, these fibroblast progenitor cells do not undergo epithelial-to-mesenchymal transition (EMT). While TCF21-expressing epicardial cells are initially multipotent, they become committed to the fibroblast lineage over time. Those TCF21-expressing cells that do not commit to the fibroblast lineage lose this expression and remain undifferentiated epicardial cells or coronary vascular smooth muscle cells.[9] TCF21 is expressed in mesodermal cells in the proepicardial organ that give rise to coronary artery smooth muscle cells (SMC) and loss of TCF21 results in increased expression of smooth muscle markers by cells on the heart surface consistent with premature SMC differentiation. This suggests that early expression of TCF21 is important for expansion of the SMC compartment of the coronary circulation, with persistent TCF21 expression being required for cardiac fibroblast development.[20]

Male TCF21 knockout mice, which die at birth due to respiratory failure, are reported to have feminized genitalia, implicating TCF21 in mouse gonadal development/differentiation.[21] TCF21 transcriptionally represses steroidogenic factor 1 (Sf1), a gene expression regulator that mediates sexual differentiation and is involved in coordinating cell fate decisions in gonadal progenitors.[21] Without TCF21, normal gonad development is disrupted as a result of ectopic expression of Sf1, which leads to abnormal committing of urogenital progenitor cells to steroidogenic cell fates. In the XY gonad, this disruption in organization contributes to changes in testicular structure and vasculature.[21]

In humans, TCF21 has been identified as a candidate tumor suppressor gene and is frequently epigenetically silenced in various human cancers.

Restriction landmark genomic scanning (RLGS) along a region of recurrent loss of heterozygosity (LOH) at chromosome 6q23-q24 to profile DNA methylation was used to test the hypothesis that abnormal promoter methylation could help pinpoint the location of a candidate tumor suppressor in regions of LOH. 6q23-q24 was the chosen chromosomal region due to frequently described LOH in human head and neck squamous cell carcinomas (HNSCC) and non-small-cell lung cancers (NSCLC) as well as in other tumor types, but with no identified tumor suppressor.[12] Hypermethylation was found to occur frequently in the same RLGS loci in HNSCC and NSCLC. Sodium bisulfite sequencing further identified tumor-specific methylation of TCF21 when compared to normal controls. RNA samples were isolated from tumor tissues and analyzed to correlate the amount of TCF21 mRNA and DNA methylation in the samples. Overall, tumor samples with higher levels of CpG island hypermethylation had decreased TCF21 expression than normal controls.[12] Exogenous expression of TCF21 in cells with silenced endogenous TCF21 loci resulted in a reductions in tumor properties both in vitro and in vivo. Based on these results, it was concluded that TCF21 is a tumor suppressor gene, often silenced by hypermethylation in cancer.[12]

TCF21 has also been linked to metastatic melanoma progression through the inhibition of the KISS1 metastasis-suppressor gene.[22] DNA methylation analysis in melanoma patient biopsies has demonstrated downregulation of TCF21 due to promoter hypermethylation, which also correlates with decreased survival in patients suffering from metastatic skin melanoma.[22] TCF21, together with E12 and TCF12, bind the KISS1 promoter, sustaining its activity. Without TCF21 to interact with the KISS1 promoter, KISS1 expression is lost. Melanoma cells overexpressing TCF21 have also been found to display reduced motility compared with vector-only control cells.[22]

Tobacco-induced lung cancer research has found TCF21 to be among the genes identified as highly methylated at both high and low concentrations of cigarette smoke condensate (CSC). In the presence genistein, one of the soy-derived bioactive isoflavones, methylation of TCF21 is significantly reduced.[23] Genistein has been known to affect tumorigenesis through epigenetic regulation, such as chromatin configuration and DNA methylation, activating other tumor suppressor genes that affect cancer cell survival.[24] These findings support the hypothesis that increasing hypermethylated tumor suppressor genes such as TCF21 is a potential chemopreventative pathway in tobacco-induced lung cancer.[23]

TCF21 may have also therapeutic potential for breast cancer treatment, as downregulation of TCF21 has been implicated in breast cancer tumorigenesis and proliferation.[15] TCF21 mRNA expression is very low in breast cancer cells compared with normal breast epithelial cells. This low expression is also associated with large tumor size and lymph node metastasis. Breast cancer tissues exhibit significantly downregulated expression of TCF21 mRNA and TCF21 mRNA overexpression has been found to inhibit cancerous cell proliferation.[15]

TCF21 methylation has been considered as a potential clinical marker in the diagnosis of renal cell carcinoma.[25] Accordingly, TCF21 methylation levels in urine samples may be a useful means of diagnosing renal cell carcinoma. Also TCF21 rs12190287 polymorphism can regulate TCF21 expression and may serve as a potential marker for genetic susceptibility to breast cancer.[26]

Additionally, a multi-locus genetic risk score study based on a combination of 27 loci, including the TCF21 gene, identified individuals at increased risk for both incident and recurrent coronary artery disease events, as well as an enhanced clinical benefit from statin therapy. The study was based on a community cohort study (the Malmo Diet and Cancer study) and four additional randomized controlled trials of primary prevention cohorts (JUPITER and ASCOT) and secondary prevention cohorts (CARE and PROVE IT-TIMI 22).[27]