[40 Analysis of Promoter Methylation and Its Role in Silencing Metallothionein I Gene Expression in Tumor Cells

By Kalpana Ghoshal, Sarmila Majumder, and Samson T. Jacob Introduction

Metallothioneins (MTs) are a group of evolutionarily conserved, cysteine-rich proteins implicated in a variety of biochemical reactions that include scavenging of oxygen-free radicals.1 Cysteine moieties, mostly arranged in Cys-X-Cys clusters, are involved in the formation of metal-thiolate bonds that impart antioxidant properties to these molecules.2 In fact, MT is the most potent scavenger of highly reactive hydroxyl radicals. Four different isoforms of MTs, encoded by different genes, have been identified in mammals.3 Among these forms, MT-I and MT-II are ubiquitously expressed and highly induced in response to various environmental toxicants, various pathologic conditions, and stressors. MT-III is primarily expressed in the glutaminergic neurons of the brain, whereas the expression of MT-IV is confined to the stratified squamous epithelium of skin and tongue.

2 P. J. Thornalley and M. Vasak, Biochim. Biophys. Acta 827, 36 (1985).

3 C. J. Quaife, S. D. Findley, J. C. Erickson, G. J. Froelic, E. J. Kelly, B. P. Zambrowicz, and R. D.

Palmiter, Biochemistry 33, 7250 (1994).

factors also was shown. Importantly, various combinations of these factors mediate cell type-specific activation of a particular chemokine.

In this chapter we have focused mainly on analysis of the activation of different chemokines and characterization of factors directly affecting this activation. The next objective will be the study of upstream mediators, such as Rac proteins, stress-and mitogen-activated kinases, and ceramide and sphingomyelinase pathways. All of them are activated during ischemia-reperfusion and are sensitive to ox/redox regulation, and have been implicated in the activation of transcription factors interacting with chemokine promoters. Methods to study those mechanisms have been described.34

The possibility that modulation of chemoattractant activation via antioxidants can significantly affect tissue injury warrants further investigation and will lead to the development of new techniques.

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