부산대학교 도서관

Recently, two genes involved in amoebic liver abscess formation in a mouse model were identified by their differential expression of non-pathogenic (A1np) and pathogenic (B2p) clones of the Entamoeba histolytica isolate HM:1-IMSS. While overexpression of a gene encoding the metallopeptidase EhMP8-2 reduces the virulence of the pathogenic clone B2p, overexpression of the gene ehi_127670 (ehhp127), encoding a hypothetical protein, increases the virulence of the non-pathogenic clone A1np, while silencing this gene in the pathogenic B2p reduces virulence. To understand the role of both molecules in determining the pathogenicity of E. histolytica, silencing, and overexpression transfectants were characterized in detail. Silencing of ehmp8-2, of the homologous gene ehmp8-1, or both in non-pathogenic A1np trophozoites significantly altered the transcript levels of 347, 216, and 58 genes, respectively. This strong change in the expression profiles caused by the silencing of ehmp8-1 and ehmp8-2 implies that these peptidases regulate the expression of numerous genes. Consequently, numerous phenotypic characteristics, including cytopathic, hemolytic, and cysteine peptidase activity, were altered in response to their silencing. Silencing of ehhp127 in pathogenic B2p trophozoites did not affect the expression of other genes, whereas its overexpression in non-pathogenic A1np trophozoites results in an altered expression of approximately 140 genes. EhHP127 is important for trophozoite motility, as its silencing reduces, while its overexpression enhances movement activity. Interestingly, the specific silencing of ehhp127 also significantly affects cytopathic, cysteine peptidase, and hemolytic activities. All three molecules characterized in this study, namely EhMP8-1, EhMP8-2, and EhHP127, are present in amoeba vesicles. The results show that ehmp8-2 and ehhp127 are not only differentially expressed between pathogenic and non-pathogenic amoebae, but that they also significantly affect amoeba pathogenicity-associated phenotypes by completely different mechanisms. This observation suggests that the regulation of amoeba pathogenicity is achieved by a complex network of molecular mechanisms rather than by single factors. Author summary: The human pathogen Entamoeba histolytica can live asymptomatically in the intestine or become invasive and cause fatal liver abscesses. Approximately 15,000 people die each year as a result of an amoebic infection. Recently, two clones with different pathogenicity (A1np: non-pathogenic; B2p: pathogenic) derived from the E. histolytica isolate HM:1-IMSS were compared at transcriptome level. Two highly differentially expressed genes (ehhp127 encoding a hypothetical protein and ehmp8-2 encoding a metallopeptidase) were identified. Analysis of E. histolytica transfectants showed that silencing of ehhp127 and overexpression of ehmp8-2 in B2p trophozoites reduced amoebic liver abscess formation in the mouse model. In this study, we characterized E. histolytica silencing and overexpression transfectants of ehmp8-2, as well as of the homologous gene ehmp8-1 and of ehhp127. It was shown that the altered expression of the metallopeptidase genes has a strong influence on the expression of a large number of genes and that the phenotype is strongly altered as a result. Silencing of ehhp127 does not affect the overall expression profile. However, specific silencing impairs motility and cytopathic activity. All three molecules have been shown to be localized in trophozoite vesicles. [ABSTRACT FROM AUTHOR]