Metallothionein III (MT3) is a putative tumor suppressor gene that is frequently inactivated in pediatric acute myeloid
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RESEARCH
Open Access
Metallothionein III (MT3) is a putative tumor suppressor gene that is frequently inactivated in pediatric acute myeloid leukemia by promoter hypermethylation Yan-Fang Tao1†, Li-Xiao Xu1†, Jun Lu1, Lan Cao1, Zhi-Heng Li1, Shao-Yan Hu1, Na-Na Wang1, Xiao-Juan Du2, Li-Chao Sun3, Wen-Li Zhao1, Pei-Fang Xiao1, Fang Fang1, Yan-Hong Li1, Gang Li1, He Zhao1, Yi-Ping Li1, Yun-Yun Xu1, Jian Ni4, Jian Wang1, Xing Feng1* and Jian Pan1*
Abstract Background: Acute myeloid leukemia (AML) is the second most common form of leukemia in children. Aberrant DNA methylation patterns are a characteristic feature in various tumors, including AML. Metallothionein III (MT3) is a tumor suppresser reported to show promoter hypermethylated in various cancers. However, the expression and molecular function of MT3 in pediatric AML is unclear. Methods: Eleven human leukemia cell lines and 41 pediatric AML samples and 20 NBM/ITP (Norma bone marrow/ Idiopathic thrombocytopenic purpura) control samples were analyzed. Transcription levels of MT3 were evaluated by semi-quantitative and real-time PCR. MT3 methylation status was determined by methylation specific PCR (MSP) and bisulfite genomic sequencing (BSG). The molecular mechanism of MT3 was investigated by apoptosis assays and PCR array analysis. Results: The MT3 promoter was hypermethylated in leukemia cell lines. More CpG’s methylated of MT3 was observed 39.0% pediatric AML samples compared to 10.0% NBM controls. Transcription of MT3 was also significantly decreased in AML samples compared to NBM/ITP controls (P < 0.001); patients with methylated MT3 exhibited lower levels of MT3 expression compared to those with unmethylated MT3 (P = 0.049). After transfection with MT3 lentivirus, proliferation was significantly inhibited in AML cells in a dose-dependent manner (P < 0.05). Annexin V assay showed that apoptosis was significantly upregulated MT3-overexpressing AML cells compared to controls. Real-time PCR array analysis revealed 34 dysregulated genes that may be implicated in MT3 overexpression and apoptosis in AML, including FOXO1. Conclusion: MT3 may be a putative tumor suppressor gene in pediatric AML. Epigenetic inactivation of MT3 via promoter hypermethylation was observed in both AML cell lines and pediatric AML samples. Overexpression of MT3 may inhibit proliferation and induce apoptosis in AML cells. FOXO1 was dysregulated in MT3-overexpressing cells, offering an insight into the mechanism of MT3-induced apoptosis. However, further research is required to determine the underlying molecular details. Keywords: Metallothionein III, Pediatric acute myeloid leukemia, Methylation, Tumor suppressor
* Correspondence: [email protected]; [email protected] † Equal contributors 1 Department of Hematology and Oncology, Children’s Hospital of Soochow University, Suzhou, China Full list of author information is available at the end of the article © 2014 Tao et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Com
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