The human T-cell factor-4 gene splicing isoforms, Wnt signal pathway, and apoptosis in renal cell carcinoma.
beta-Catenin and transcriptional factor TCF-4 (human T-cell factor-4) genes comprise the Wnt signal. The Wnt signal pathway plays an important role in malignant transformation. We hypothesize that the beta-catenin and TCF-4 gene and Wnt signal are important in the progression of renal cell carcinoma (RCC). To test this hypothesis, we investigated TCF-4 splicing isoforms, beta-catenin, and Wnt signal pathway (cyclin D1, c-myc, c-jun, and MMP7) in three RCC cell lines (A498, Caki-1, and Caki-2), 38 primary RCCs, and 29 normal kidney samples. We also analyzed the relationship between TCF-4 gene splicing isoforms, proliferation (proliferating cell nuclear antigen labeling index), and apoptosis [antiapoptotic factors (Bcl-2 and Bcl-x(L)), proapoptotic factors (Bak and Bax), and caspase-3] in RCC samples. In 38 RCC samples, four splicing isoforms of the TCF-4 gene were present in the region between exon 12 and exon 17. Thirty (79%) of 38 RCCs and all (100%) of the normal kidney samples showed mixed isoforms with both long and short reading frames in the COOH-terminal region, whereas the remaining 8 RCC samples showed only the long-form reading frame. Two COOH-terminal-binding protein sites were present only in the long-form reading frame. The eight RCCs that demonstrated only the long reading frame isoform showed early disease progression and poor prognosis. In these 8 RCC samples, down-regulation of cyclin D1, c-myc, c-jun, and MMP7 expression was observed at the mRNA level. In addition, a marked reduction of caspase-3 expression was also found at both the mRNA and the protein level. However, the beta-catenin gene was not overexpressed at the mRNA level and protein level, and mutation and deletion were not observed in exon 3. In these three renal cell lines, there was no significant difference in TCF-4 mRNA expression before and after 5-Aza-2'-deoxycytidine treatment, and there appeared to be no splicing isoforms in the region between exon 1 and exon 11. These findings suggest that alteration in beta-catenin is an infrequent event in RCC. In samples in which beta-catenin was not overexpressed, the target genes of Wnt signal were regulated through TCF-4 splicing isoforms. The imbalance between TCF-4 gene splicing isoforms with long and short reading frames is associated with RCC progression through the inhibition of the apoptotic pathway. We demonstrate for the first time that TCF-4 gene splicing isoforms and the Wnt signal pathway can induce progression of RCC by the inhibition of apoptosis and not by the induction of cell proliferation.
Shiina
Hiroaki
H
Department of Urology and Biochemistry, Shimane Medical University, Izumo 693-0085, Japan.
Igawa
Mikio
M
Breault
Julia
J
Ribeiro-Filho
Leopoldo
L
Pookot
Deepa
D
Urakami
Shinji
S
Terashima
Masaharu
M
Deguchi
Masao
M
Yamanaka
Masaki
M
Shirai
Masato
M
Kaneuchi
Masanori
M
Kane
Christopher J
CJ
Dahiya
Rajvir
R
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000595:Amino Acid Sequence; D017209:Apoptosis; D001483:Base Sequence; D002292:Carcinoma, Renal Cell / Q000473:pathology*; D002455:Cell Division; D003598:Cytoskeletal Proteins / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics*; D006801:Humans; D007680:Kidney Neoplasms / Q000473:pathology*; D008875:Middle Aged; D008969:Molecular Sequence Data; D018807:Polymorphism, Single-Stranded Conformational; D011379:Prognosis; D020033:Protein Isoforms; D011518:Proto-Oncogene Proteins / Q000502:physiology*; D012333:RNA, Messenger / Q000032:analysis; D015398:Signal Transduction / Q000502:physiology*; D051820:TCF Transcription Factors; D015534:Trans-Activators / Q000235:genetics; D058118:Transcription Factor 7-Like 2 Protein; D014157:Transcription Factors / Q000235:genetics*; D051153:Wnt Proteins; D029961:Zebrafish Proteins; D051176:beta Catenin
C495270:CTNNB1 protein, human; D003598:Cytoskeletal Proteins; D004268:DNA-Binding Proteins; D020033:Protein Isoforms; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D051820:TCF Transcription Factors; C544894:TCF7L2 protein, human; D015534:Trans-Activators; D058118:Transcription Factor 7-Like 2 Protein; D014157:Transcription Factors; D051153:Wnt Proteins; D029961:Zebrafish Proteins; D051176:beta Catenin