CRYO-ELECTRON TOMOGRAPHY OF GIARDIA INTESTINALIS

University Of Colorado, Boulder CO

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Abstract

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Giardia intestinalis (also known as Giardia lamblia) is a unicellular, flagellated protist with a two-stage lifecycle. Giardiasis is caused by ingestion of the cyst form of the parasite which then excysts in the duodenum into the trophozoite form. The trophozoite form has a highly specialized microtubule cytoskeleton (eight flagella, the "median body" and the "ventral disc"), which permits the parasite to evade peristaltic movement in the host's intestine. Infection with Giardia is the results of parasite attachment directly to the intestinal epithelium;deriving nutrients from the mucosal secretions of the epithelial layer. The mechanism of attachment is not fully understood, but recent evidence suggests the parasite uses suction created by conformational changes of the ventral disc. The ventral disc consists of a counterclockwise array of microtubules that originate from electron dense bands directly anterior to the caudal and posterior-lateral flagellar basal bodies. The ventral surface of the microtubules is attached to the plasma membrane. On the dorsal side of each microtubule is a microribbon that is connected to neighbor microribbons by cross-linkers. Previous studies of the Giardia cytoskeleton have focused on understanding the basic disc structure using low-resolution techniques such as light microscopy, chemically fixed plastic embedded samples, and negative stain. In our study, we are using cryo-electron tomography of isolated detergent-extracted cytoskeletons to determine the structural-functional relationships between the various components of the ventral disc. We have used our program PEET to average the microtubules of the ventral disc.