Bug22p, a Conserved Centrosomal/Ciliary Protein Also Present in Higher Plants, Is Required for an Effective Ciliary Stroke in Paramecium

Abstract

Centrioles, cilia, and flagella are ancestral conserved organelles of eukaryotic cells. Among the proteins identified in the proteomics of ciliary proteins in Paramecium, we focus here on a protein, Bug22p, previously detected by cilia and basal-body high-throughput studies but never analyzed per se. Remarkably, this protein is also present in plants, which lack centrioles and cilia. Bug22p sequence alignments revealed consensus positions that distinguish species with centrioles/cilia from plants. In Paramecium, antibody and green fluorescent protein (GFP) fusion labeling localized Bug22p in basal bodies and cilia, and electron microscopy immunolabeling refined the localization to the terminal plate of the basal bodies, the transition zone, and spots along the axoneme, preferentially between the membrane and the microtubules. RNA interference (RNAi) depletion of Bug22p provoked a strong decrease in swimming speed, followed by cell death after a few days. High-speed video microscopy and morphological analysis of Bug22p-depleted cells showed that the protein plays an important role in the efficiency of ciliary movement by participating in the stroke shape and rigidity of cilia. The defects in cell swimming and growth provoked by RNAi can be complemented by expression of human Bug22p. This is the first reported case of complementation by a human gene in a ciliate.

The conserved ciliary protein Bug22 controls planar beating ofChlamydomonasflagella

“…Immunogold labeling of Bug22 in Paramecium shows that it is preferentially localized between ciliary membrane and axoneme, as well as to basal bodies. Knockdown of Bug22 expression by RNAi in this organism results in the conversion of the ciliary waveform to the flagellar waveform, which is consistent with our results (Laligné et al, 2010).…”

“…The orientation of the two basal bodies was similar in wild type and mutant cells (Fig. 3B,C), which was however altered in Bug22 RNAi strains of Paramecium (Laligné et al, 2010). In addition, the 'H' structure and Y-links in the transition zone appeared normal in the mutant compared with wild type (Fig.…”

“…It has been shown that the proximal half to two-thirds of the flagellum is more stable and resistant to drug-induced flagellar shortening (Dentler and Adams, 1992). It has also been proposed that Bug22 in Paramecium might regulate cilia rigidity (Laligné et al, 2010). It is interesting that Bug22 appears to be much more abundant than outer row dynein subunits of similar size.…”

“…This protein is highly conserved in ciliated organisms as well as in higher plants (Hodges et al, 2011;Laligné et al, 2010). Alignment of Bug22 with its human ortholog (GenBank NP_037374.1) showed 89% identity within 98% coverage.…”

“…Alignment of Bug22 with its human ortholog (GenBank NP_037374.1) showed 89% identity within 98% coverage. Bug22 has been shown to be a flagellar protein in Paramecium and Trypanosome (Hodges et al, 2011;Laligné et al, 2010). Immunogold labeling of Bug22 in Paramecium shows that it is preferentially localized between ciliary membrane and axoneme, as well as to basal bodies.…”