Double-stranded RNA mediates homology-dependent gene silencing of gamma-tubulin in the human parasite Entamoeba histolytica

Approaches that eliminate mRNA are a powerful tool for reverse genetics applications in eukaryotic microbes for which gene replacement techniques have not yet been developed. Here, for the first time, we demonstrate that RNA duplexes efficiently inhibit gene expression when introduced into the human parasite Entamoeba histolytica. Chemically synthesized, small interfering RNA (siRNA) were highly specific and efficient in silencing parasite gamma-tubulin mRNA. Use of specific antibodies revealed that microtubules and gamma-tubulin were intra-nuclear in E. histolytica. The RNAi approach to modulation of gamma-tubulin mRNA resulted in loss of the highly organized microtubule array an observation that correlates with a significant reduction of gamma-tubulin as well as of the specific mRNA. Our results suggest that gamma-tubulin is essential for microtubule nucleation in E. histolytica.