Abstract
Tumour suppressor miR-34c deficiency resulted from hyper-methylation in its promoter is believed to be one of the main causes of colorectal cancer (CRC). Till date, miR-34c has been validated as a direct target of p53; but previous evidence suggested other transcription factor(s) must be involved in miR-34c transcription. In the present study, we in the first place identified a core promoter region (-1118 to -883bp) of pre-miR-34c which was embedded within a hyper-methylated CpG island. Secondly, E2F1 promoted miR-34c transcription by physical interaction with the miR-34c promoter at site -897 to -889bp. The transcriptional activating effect of E2F1 on miR-34c was in a p53 independent manner but profoundly promoted in the presence of p53 with exposure to 5-aza-2'-deoxycytidine (DAC). Thirdly, stem cell factor (SCF), a miR-34c target, was specifically reduced upon an introduction of E2F1 which lead to suppression of CRC cell proliferation. The E2F1-suppressed cell proliferation was partially abrogated by additional miR-34c inhibitor, indicating that the anti-proliferation effect of E2F1 was probably through activating miR-34c-SCF axis. Finally, SCF/KIT signalling increased E2F1 production by reducing its proteosomal degradation dependent on PI3K/Akt-GSK3β pathway. In conclusion, our results suggested the existence of E2F1-miR-34c-SCF negative feedback loop which was interrupted by the hyper-methylation of miR-34c promoter in CRC cells and increased cell proliferation.
Highlights
Colorectal cancer (CRC) ranks the top five most malignant neoplasms both in China and western countries
Little was known about the molecules that mediated miR-34c expression in CRC, except p53
We demonstrated that, besides p53, E2F1 was a potent activator for miR-34c transcription in CRC cells
Summary
Colorectal cancer (CRC) ranks the top five most malignant neoplasms both in China and western countries. The tumorigenesis and development of CRC are resulted from a network of multiple aberrant molecules including miRNAs. miRNAs posttranscriptionally regulate their target genes via either translational repression or mRNA degradation. Depending on their targets, miRNAs could serve as either oncogenes or tumour suppressing genes [1,2]. MiR-34 family including miR-34a, miR-34b and miR-34c have been suggested to be candidates of tumour suppressing genes that could induce apoptosis and cell-cycle arrest and inhibit proliferation and colony formation in soft agar [3]; whereas in CRC tissues miR-34s were silenced [4,5,6]. We have demonstrated that over-expression of miR-34c induced apoptosis and inhibited proliferation in CRC cells by silencing its target, stem cell factor (SCF, known as KITLG) [12], suggesting miR-34c as a prominent target for the treatment of CRC patients
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