Lauwaet, T., M. Baitaluk, D.S. Reiner, E.P. Romijn, C.C.L. Wong, H. Skarin, B.J. Davids, S.R. Birkeland, M.J. Cipriano, D. Palm, S.P. Preheim, A. Gupta, S.G. Svard, A.G. McArthur, J.R. Yates, A. Ray, & F.D. Gillin. 2008. Proteome, transcriptome and interactome analyses reveal key roles of basal bodies in Giardia differentiation. Presentation at the 4th International Conference on Anaerobic Protists, Taoyuan, Taiwan.
The success of Giardia as a parasite depends on its ability to accurately interpret physiologic signals from its external environment and to respond by differentiating into a dormant infectious cyst that can awaken into a pathogenic trophozoite. Giardia is a unique model for other intestinal protozoa because its life cycle has been completed in vitro. Since excystation is rapid, we proposed that signal transduction pathways are important. We found earlier that calmodulin, PKA, and PP2A, which are crucial to and upregulated in excystation, localize to the basal bodies/centrosomes. We hypothesized that the eight flagellar basal bodies regulate and co-ordinate the cellular reorganization of excystation. We isolated basal bodies from giardial cytoskeletons, analyzed their proteome by Multidimensional Protein Identification Technology (MudPIT), and matched individual peptides to genes in the Giardia genome database. We identified a total of 369 basal body proteins of which 98 are unique to Giardia.
To evaluate their functional significance, we identified human orthologs of giardial basal body proteins and constructed an interaction network (interolog) model of Giardia basal body proteins based on known protein-protein interactions in the human interactome. To determine whether the expression of genes encoding basal body proteins changes during differentiation, we measured their mRNA expression levels during growth, encystation and excystation by SAGE (serial analysis of gene expression). Of 175 genes encoding basal body proteins with SAGE tags, 72 were upregulated in encystation and 46 in cysts and excystation. When we overlaid the interolog network with the mRNA expression data, we found that expression of most protein biosynthesis genes is upregulated during encystation, while that of signaling and cytoskeletal genes is upregulated during excystation. This is consistent with the biological roles of these pathways. We verified the authenticity of the interolog model by using calmodulin, which localizes only to the basal bodies, as bait and affinity-purified proteins from the five major functional groups in the network.
Our proteome, transcriptome, and interactome analyses reveal complex composition and dynamic expression of Giardia basal body components. We propose that these organelles are cellular control centers regulating differentiation and entry into and awakening from dormancy.
