Silencing of metallothionein 1A gene in melanoma

Abstract

Background: When a CpG island (CGI; a dense cluster of CpGs) located in the 5′ region of a gene is methylated, its transcription is suppressed. Tumorigenesis of melanoma is associated with trace elements. Metallothionein 1A is closely associated with the metabolism of trace elements. However, little is known about the metallothionein 1A gene (MT1A) in melanoma. Objective: The purpose is to reveal the methylation and expression status of MT1A in melanoma. Methods: Quantitative real-time methylation-specific PCR (RT-MSP) and bisulfite sequencing were performed to examine MT1A methylation status. Quantitative real-time reverse transcription-PCR (RT-PCR) was performed to examine MT1A expression. Results: Some melanoma cell lines exhibited high methylation levels of the CGI located in the 5′ region of MT1A (5′ MT1A CGI) with suppression of MT1A. Other melanoma cell lines and normal cultured melanocytes exhibited low methylation levels of 5′ MT1A CGI with expression of MT1A. Treatment with a demethylating agent resulted in transcriptional induction of MT1A in the melanoma cell lines SK-MEL-5 and G-361 with high methylation levels prior to treatment. The methylation levels of 5′ MT1A CGI ranged widely from 0.0% to 91.4% in 21 clinical melanoma samples but showed a narrow, low range from 0.0% to 6.4% in 23 clinical melanocytic nevus samples. Data of bisulfite sequencing was generally compatible with those of RT-MSP. The methylation levels ranged according to the types of melanoma (Kruskal-Wallis test, P = 0.047). Conclusion: MT1A is aberrantly silenced by DNA methylation of 5′ MT1A CGI in melanoma.