Identification of genomic responses to collagen binding by trophozoites of Entamoeba histolytica

Attachment of Entamoeba histolytica trophozoites to collagen is a known stimulus for parasite activation, leading to subsequent tissue destruction and invasion. To identify cellular mechanisms of trophozoite activation, we assessed global variations in gene expression during collagen interaction with E. histolytica. A shotgun DNA microarray was constructed by use of 9600 random inserts from an E. histolytica genomic DNA library. Through differential hybridization, key differences between gene expression in collagen-activated trophozoites and that in nonactivated trophozoites were identified. Fourteen differentially regulated clones were reproducibly identified and selected for sequencing. Among the genes identified were those coding for (1) components of a signaling cascade that had been previously hypothesized to transmit responses to cell attachment, (2) adapter proteins for vesicle formation, and (3) proteins that are implicated in cytoskeletal reorganization and locomotion. Two known virulence-factor genes-those for cysteine proteinases and amebapore-also were up-regulated in response to collagen stimulation. These results provide important new clues about how a pathogen orchestrates responses to the host environment as well as a new tool for the analysis of other aspects of Entamoeba species infection and pathogenicity.