Aberrant promoter methylation in human DAB2 interactive protein (hDAB2IP) gene in gastrointestinal tumour

Abstract

The human DOC-2/DAB2 interactive protein (hDAB2IP) gene is a novel member of the Ras GTPase-activating family and has been demonstrated to be a tumour-suppressor gene inactivated by methylation in several cancers. In this study, we analysed the methylation and expression status of hDAB2IP in gastrointestinal tumours. The promoter region of hDAB2IP was divided into two regions (m2a and m2b) based on our previous report, and the methylation status was determined by bisulphite DNA sequencing in gastric cancer cell lines. The gene expression was semiquantified by real-time RT -PCR, and the results indicated that the m2b promoter region might be an authentic methylation-mediated key regulator of the gene expression. Based on the sequence data, we developed a methylation-specific PCR (MSP) for the m2a and m2b regions and applied it to the samples. Methylation-specific PCR revealed aberrant methylation in the m2a region in eight of 12 gastric cancer cell lines (67%), 16 of 35 gastric cancer tissues (46%) and 29 of 60 colorectal cancer tissues (48%), and in the m2b region in eight of 12 cell lines (67%), 15 of 35 gastric cancer tissues (43%) and 28 of 60 colorectal cancer tissues (47%). On the other hand, seven (12%) and 11 (19%) of 59 gastrointestinal nonmalignant mucosal specimens showed methylation in the m2a and m2b regions, respectively, suggesting that hDAB2IP methylation might play a causative role in carcinogenesis. The 5-aza-2 0 -deoxycytidine treatment restored the gene expression in the m2b-methylated cell lines, confirming that the methylation caused gene downregulation. We also examined the relationship between hDAB2IP methylation and the clinicopathological features in patients with primary tumours, and determined that methylation in the m2b region was associated with location of the tumour in the stomach. In summary, our results demonstrated that hDAB2IP methylation is frequently present in gastrointestinal tumours and that the resulting gene silencing plays an important role in gastrointestinal carcinogenesis.

Epigenetic silencing of a Ca 2+ -regulated Ras GTPase-activating protein RASAL defines a new mechanism of Ras activation in human cancers

“…Few studies have detected mutation in the classic Ras GAP p120 GAP (29), although heterozygous mutations of the p120 GAP gene do promote abnormal angiogenic remodeling and cause the disease capillary malformationarteriovenous malformation (30). More recently, in addition to the RNAi screens that have proposed tumor suppressor functions for RASAL and CAPRI (19,20), DAB2IP (human DAB2 interactive protein), a Ras GAP that associates with the disable gene family member DAB2 (31), has been demonstrated to be silenced in prostate, breast, lung, and gastrointestinal tumors through aberrant promoter CpG methylation (32)(33)(34)(35)(36). Unfortunately, it remains to be established exactly how DAB2IP is regulated in vivo upon receptor activation.…”

Block one, unleash a hundred. Mechanisms of DAB2IP inactivation in cancer

“…[60][61][62][63] These studies focused on a CpG island partially overlapping exon 3, which contains a functional promoter and TSS. Transcripts originating at this TSS are spliced to exon 4 and are predicted to encode the shortest DAB2IP protein isoform (Figure 1).…”

“…Published studies provide compelling evidence that methylation of this region correlates with reduced DAB2IP expression in tumor tissues and cell lines, despite the presence of at least two additional TSSs located, respectively, 130 and 48 kbp upstream, and other functional TSSs further downstream. [60][61][62][63][64][65] It is possible that methylation of this region is sufficient to turn the gene off; alternatively, other sites may be coordinately methylated for efficient DAB2IP silencing. Three additional CpG islands are mapped in the hDAB2IP gene, in some cases corresponding to functional TSSs; little is currently known about the methylation status of these regions in normal or pathological conditions, or its potential impact on DAB2IP transcription.…”

Differential Regulation of RasGAPs in Cancer

“…36 DAB2IP, which is a RasGAP associated with the disabled gene family member DAB2, is also silenced in prostate, breast, lung, and gastrointestinal tumors through aberrant promoter CpG methylation. [128][129][130][131][132] Furthermore, 2 genome-wide studies identified DAB2IP as a putative tumor suppressor in aggressive prostate adenocarcinoma. 133 Unfortunately, little is known about the regulation of DAB2IP in vivo and correlation to Ras activity in the cancer samples analyzed, but together with the observations described below, epigenetic silencing of the various GAPs implicates a general theme that requires further investigation.…”