Giardia’s ventral disc is hyperstable and composed of over 80 disc-associated proteins

Abstract

Giardia is a common protistan parasite that causes diarrheal disease worldwide. Motile trophozoites colonize the small intestine, attaching to the villi with the ventral disc, a unique and complex microtubule (MT) organelle. Attachment to the host epithelium allows Giardia to resist peristalsis during infection of the host gastrointestinal tract. Despite our emerging view of the complexity of ventral disc architecture, we are still in the very preliminary stages of understanding how specific structural elements contribute to disc stability or generate forces for attachment. The ventral disc is a large, dome-shaped, spiral MT array decorated with microribbon-crossbridge protein complexes (MR-CB) that extend upward into the cytoplasm. To find additional disc-associated proteins (DAPs), we used a modified method for disc biochemical fractionation in high salt followed by shotgun proteomic analyses and validation by GFP-tagging. Using this method in conjunction with an ongoing subcellular localization screen, we identified 54 new DAPs. Of the 87 DAPs confirmed to date, 54 localize only to the disc, and the remainder localize to additional structures including the flagella, basal bodies, or median body. Almost one third of the known DAPs lack any homology to proteins in other eukaryotes and another one third simply contain ankyrin repeat domains. Many DAPs localize to specific structural regions of the disc, including the ventral groove region and disc margin. Lastly, we show that spiral singlet MT array comprising the disc is hyperstable and lacks dynamic instability, and we attribute these unique properties to the presence of both novel DAPs as well conserved MAPs and MIPs that are known to stabilize ciliary doublet and triplet MTs.