Manipulation of Gene Expression in Entamoeba histolytica
Rivka Bracha, Nancy Guillén and David Mirelman
from: Anaerobic Parasitic Protozoa: Genomics and Molecular Biology (Edited by: C. Graham Clark, Patricia J. Johnson and Rodney D. Adam). Caister Academic Press, U.K. (2010)
Our knowledge of the functions of different structural proteins and virulence factors in the cellular organization and pathogenesis of Entamoeba histolytica has significantly increased following the introduction of various molecular techniques that enable the manipulation of gene expression. Unfortunately, to date, all the attempts to integrate exogenous DNA into the parasite's genome have failed and most methods for up- and down-regulation of gene expression have been based on the transfection of stably maintained plasmids. Down-regulation has been achieved by plasmids encoding: (i) antisense RNA, (ii) truncated or mutated genes that exert dominant-negative effects, and (iii) inverted loops that generate double stranded RNA molecules. Small interfering RNA oligonucleotides incorporated directly from the culture medium also cause the down-regulation of specific genes. Recently, epigenetic silencing of the amoebapore gene, which encodes a toxic polypeptide that disrupts the membranes of bacteria and mammalian cells, was induced following transfections with a plasmid construct containing an upstream segment of the amoebapore gene. A number of additional genes have been silenced subsequently in a similar way. Amoebae in which the expression of virulence genes has been down-regulated have in many cases an altered phenotype that sheds light on the specific role of the genes and the cells display a significantly reduced pathogenic potential read more ...