DYF-1 Is Required for Assembly of the Axoneme in Tetrahymena thermophila

Abstract

In most cilia, the axoneme can be subdivided into three segments: proximal (the transition zone), middle (with outer doublet microtubules), and distal (with singlet extensions of outer doublet microtubules). How the functionally distinct segments of the axoneme are assembled and maintained is not well understood. DYF-1 is a highly conserved ciliary protein containing tetratricopeptide repeats. In Caenorhabditis elegans, DYF-1 is specifically needed for assembly of the distal segment (G. Ou, O. E. Blacque, J. J. Snow, M. R. Leroux, and J. M. Scholey. Nature. 436:583-587, 2005). We show that Tetrahymena cells lacking an ortholog of DYF-1, Dyf1p, can assemble only extremely short axoneme remnants that have structural defects of diverse natures, including the absence of central pair and outer doublet microtubules and incomplete or absent B tubules on the outer microtubules. Thus, in Tetrahymena, DYF-1 is needed for either assembly or stability of the entire axoneme. Our observations support the conserved function for DYF-1 in axoneme assembly or stability but also show that the consequences of loss of DYF-1 for axoneme segments are organism specific.Cilia are microtubule-rich cellular extensions that arise from basal bodies near the surfaces of most eukaryotic cell types. Defective cilia cause a wide variety of diseases, including polycystic kidney disease, primary ciliary dyskinesia, and retinal degeneration (3). A typical motile cilium has a microtubulebased framework, the axoneme, which contains nine outer (mostly doublet) microtubules and two central (singlet) microtubules. In most cilia, the axoneme can be subdivided into three segments: proximal (transition zone), middle (containing outer doublet microtubules), and distal (containing singlet extensions of peripheral microtubules). The outer doublet microtubules of the middle segment have a complete tubule A made of 13 protofilaments and an incomplete tubule B made of 11 protofilaments that is fused to the wall of the A tubule (36, 57). The outer microtubules in the distal segment lack the B tubule (32, 49). The distal segment also lacks dynein arms and radial spokes, and its microtubules are terminated by caps that are associated with the plasma membranes at the tips of cilia (11, 50). The distal segments are characterized by a high level of microtubule turnover, which could play a role in the regulation of the length of cilia (31).The mechanisms that establish the segmental subdivision of the axoneme are not well understood. Studies of Caenorhabditis elegans indicate that the distal segment is assembled using a mechanism that differs from the one utilized in the middle and proximal segments (54). In most cell types, ciliogenesis is dependent on the intraflagellar transport (IFT) pathway, a bidirectional motility of protein aggregates, known as IFT particles, that occurs along outer microtubules (10,28,29,42).IFT particles are believed to provide platforms for transport of axonemal precursors (23, 44). The anterograde component of IFT that delivers cargo from ...

Tubulin Tyrosine Ligase-like Genes ttll3 and ttll6 Maintain Zebrafish Cilia Structure and Motility

“…Also, because cilia length was not affected by ttll3/6 knockdown, whereas fleer mutants show shorter axonemes and defects in multiciliated cell development, it is likely that fleer has additional functions in ciliogenesis. Strong ciliogenesis defects in Tetrahymena and Trypanosome Dyf-1 mutants also suggest additional roles for Fleer/Dyf-1 in IFT processes underlying ciliogenesis (48,49).…”

Ciliary and Flagellar Structure and Function—Their Regulations by Posttranslational Modifications of Axonemal Tubulin

“…Zebrafish fleer is also an ortholog of IFT70/DYF-1 and is involved in proper establishment of B tubules (Pathak et al, 2007). In Tetrahymena, the deletion mutant of IFT70/DYF-1 causes hyperglutamylation of axonemal tubulin, rather than decreasing the level of glutamylation (Dave et al, 2009). A battery of reports indicates that IFT70/DYF-1/Fleer is involved in the functional connection between IFT machineries and glutamylation-performing enzymes.…”

ChlamydomonasIFT70/CrDYF-1 Is a Core Component of IFT Particle Complex B and Is Required for Flagellar Assembly

“…The defects include the absence of a central pair and outer doublet microtubules, and incomplete or absent B tubules on the outer microtubules (Dave et al , 2009). However, in contrast to the results with the fleer zebrafish, the level of tubulin glutamylation was increased in the axonemal remnants of the DYF1p knockout Tetrahymena cells (Dave et al , 2009).…”