mRNA Sequencing Reveals Upregulation of Glutathione S-Transferase Genes during Acanthamoeba Encystation

Abstract

Some members of the genus Acanthamoeba are facultative pathogens typically with a biphasic lifestyle: trophozoites and cysts. Acanthamoeba is capable of infecting the cornea, resulting in Acanthamoeba keratitis. The cyst is one of the key components for the persistence of infection. Gene expression during Acanthamoeba encystation showed an upregulation of glutathione S-transferase (GST) genes and other closely related proteins. mRNA sequencing showed GST, and five genes with similar sequences were upregulated after 24 h of inducing encystation. GST overexpression was verified with qPCR using the HPRT and the cyst-specific protein 21 genes as controls. The GST inhibitor ethacrynic acid was found to decrease cell viability by 70%. These results indicate a role of GST in successful encystation, possibly by maintaining redox balance. GST and associated processes could be targets for potential treatments alongside regular therapies to reduce relapses of Acanthamoeba infection.

Keywords: Acanthamoeba; cyst; encystation; glutathione S-transferase.

Figure 1

Differential expression of GST (left) and CSP21 (right) after 24 h of encystment stimulus. Each graph compares treatments at 0- and 24-h, representing fold change in expression; error bars represent standard error of the mean.

Figure 2

Cell viability of cultures encysting with GST inhibitors. Values represent percentage of cell viability. Negative control was used as a regular culture with AX2. The positive control is the encysting culture with NEM. EA25: Ethacrynic acid 25 µM. EA250: Ethacrynic acid 250 µM. SS100: Sulfosalazine 100 µM. SS1: Sulfosalazine 1 mM. Error bars represent the standard error of the mean.