Inactivation of the putative suppressor gene DOK1 by promoter hypermethylation in primary human cancers

Abstract

The DOK1 gene is a putative tumour suppressor gene located on the human chromosome 2p13 which is frequently rearranged in leukemia and other human tumours. We previously reported that the DOK1 gene can be mutated and its expression down-regulated in human malignancies. However, the mechanism underlying DOK1 silencing remains largely unknown. We show here that unscheduled silencing of DOK1 expression through aberrant hypermethylation is a frequent event in a variety of human malignancies. DOK1 was found to be silenced in nine head and neck cancer (HNC) cell lines studied and DOK1 CpG hypermethylation correlated with loss of gene expression in these cells. DOK1 expression could be restored via demethylating treatment using 5-aza-2′deoxycytidine. In addition, transduction of cancer cell lines with DOK1 impaired their proliferation, consistent with the critical role of epigenetic silencing of DOK1 in the development and maintenance of malignant cells. We further observed that DOK1 hypermethylation occurs frequently in a variety of primary human neoplasm including solid tumours (93% in HNC, 81% in lung cancer) and hematopoietic malignancy (64% in Burkitt’s lymphoma). Control blood samples and exfoliated mouth epithelial cells from healthy individuals showed a low level of DOK1 methylation, suggesting that DOK1 hypermethylation is a tumour specific event. Finally, an inverse correlation was observed between the level of DOK1 gene methylation and its expression in tumour and adjacent non tumour tissues. Thus, hypermethylation of DOK1 is a potentially critical event in human carcinogenesis, and may be a potential cancer biomarker and an attractive target for epigenetic-based therapy.

Transcriptional Regulation of the Human Tumor Suppressor DOK1 by E2F1

“…DOK1 or DOK2 knockout mice show a high susceptibility to developing leukemia and hematological malignancies (19,23,33), as well as lung adenocarcinomas (2). Concomitant with these findings, we showed that DOK1 gene expression was repressed in a large proportion of head and neck cancer (HNC), lung, liver, and gastric cancers, and Burkitt's lymphoma as a result of aberrant hypermethylation of the DOK1 promoter region (1,14,24). These data firmly establish the tumor suppressor properties of DOK1.…”

“…HEK293 cells and the HNC cell lines HNC-41 (tonsil), HNC-97 (oral cavity), HNC-124 (oral cavity), and PNS-136 (paranasal sinus) and the colon cancer cell line LoVo were described previously (24). HEK293 cells were transfected using Fugene 6 (Roche Diagnostics) and analyzed 48 h after transfection.…”

“…The following antibodies were used: anti-DOK1 ab8112 (Abcam), E2F1 sc-56662, E2F1 sc-251x, E2F2 sc-633, E2F3 sc-866 (all from Santa Cruz Biotechnology), ␤-actin C4 (MP Biomedicals), p53 DO-7 (Novocastra), anti-BAX sc-526, mouse IgG, rabbit IgG (all from Santa Cruz Biotechnology), H3K9ac (Cell Signaling Technology), H3K4me3, and H3K27me3 (both from Epigentek). Immunoblotting was performed as described previously (24).…”

“…These data firmly establish the tumor suppressor properties of DOK1. Given that DOK1 is frequently altered in a variety of human cancers, it could potentially serve as a new marker and/or a therapeutic target for cancer control (1,2,14,24).…”

Dok1 and Dok2 Proteins Regulate Cell Cycle in Hematopoietic Stem and Progenitor Cells

“…Paradoxically, upon hematopoietic stress, Dok proteins oppose the quiescence of HSC (8,9,48,49). Dok1 and Dok2 dysregulations have been associated with several oncogenic processes (1,6,7,12,(50)(51)(52)(53)(54)(55)(56). Therefore, the impact of chemotherapeutic agents on HSCs may have to be considered to avoid resistance of leukemic stem cells and subsequent relapse.…”