Abstract
The Ras-association domain family 1 (RASSF1) gene has seven different isoforms; isoform A is a tumor-suppressor gene (RASSF1A). The promoter of RASSF1A is inactivated in many cancers, whereas the expression of another major isoform, RASSF1C, is not affected. Here, we show that RASSF1C, but not RASSF1A, interacts with betaTrCP. Binding of RASSF1C to betaTrCP involves serine 18 and serine 19 of the SS(18)GYXS(19) motif present in RASSF1C but not in RASSF1A. This motif is reminiscent of the canonical phosphorylation motif recognized by betaTrCP; however, surprisingly, the association between RASSF1C and betaTrCP does not occur via the betaTrCP substrate binding domain, the WD40 repeats. Overexpression of RASSF1C, but not of RASSF1A, resulted in accumulation and transcriptional activation of the beta-catenin oncogene, due to inhibition of its betaTrCP-mediated degradation. Silencing of RASSF1A by small interfering RNA was sufficient for beta-catenin to accumulate, whereas silencing of both RASSF1A and RASSF1C had no effect. Thus, RASSF1A and RASSF1C have opposite effects on beta-catenin degradation. Our results suggest that RASSF1C expression in the absence of RASSF1A could play a role in tumorigenesis.
Highlights
Since the discovery of RASSF1A epigenetic inactivation in lung tumors [1, 2], numerous reports have implicated the inactivation of the RASSF1A promoter by hypermethylation in tumorigenesis across a wide spectrum of human tumors [3, 4]
We did two-hybrid screening using a hTrCP fragment from residues 1 to 291 as bait, and we found that hTrCP interacts with RASSF1C
We studied the effect of RASSF1C overexpression on two other known substrates of hTrCP, InBa and ATF4, and we found that the degradation rate of these hTrCP substrates was inhibited by overexpression of RASSF1C, whereas RASSF1A had no effect (Supplementary Fig. S2 for InBa; data not shown for ATF4)
Summary
Since the discovery of RASSF1A epigenetic inactivation in lung tumors [1, 2], numerous reports have implicated the inactivation of the RASSF1A promoter by hypermethylation in tumorigenesis across a wide spectrum of human tumors [3, 4]. The RASSF1 gene has eight exons and is located within a 120-kb region of chromosome 3p21.3 that frequently undergoes allele loss in human tumors [3, 4]. Differential promoter usage and alternative splicing generate seven different transcripts (RASSF1A–RASSF1G). Isoforms A and C are ubiquitously expressed, whereas the expression of the other isoforms is tissue specific [4]. A direct correlation between promoter methylation and loss of RASSF1A expression has been shown in many tumor cell lines, including small-cell lung, non–.
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