Abstract
Cytoplasmic assembly of ciliary dyneins, a process known as preassembly, requires numerous non-dynein proteins, but the identities and functions of these proteins are not fully elucidated. Here, we show that the classical Chlamydomonas motility mutant pf23 is defective in the Chlamydomonas homolog of DYX1C1. The pf23 mutant has a 494 bp deletion in the DYX1C1 gene and expresses a shorter DYX1C1 protein in the cytoplasm. Structural analyses, using cryo-ET, reveal that pf23 axonemes lack most of the inner dynein arms. Spectral counting confirms that DYX1C1 is essential for the assembly of the majority of ciliary inner dynein arms (IDA) as well as a fraction of the outer dynein arms (ODA). A C-terminal truncation of DYX1C1 shows a reduction in a subset of these ciliary IDAs. Sucrose gradients of cytoplasmic extracts show that preassembled ciliary dyneins are reduced compared to wild-type, which suggests an important role in dynein complex stability. The role of PF23/DYX1C1 remains unknown, but we suggest that DYX1C1 could provide a scaffold for macromolecular assembly.
Highlights
Motile cilia are antenna-like organelles protruding from many types of cells and required for motility and cell signaling [1, 2]
The movement of motile cilia is driven by the ciliary dynein motors, which are composed of outer (ODA) and inner dynein arms (IDA) [8,9,10]
The ciliary lengths of the rescued strains are near wild-type in the SG (Sager and Granick) media. These results strongly suggest that DYX1C1 is the gene responsible for the pf23 phenotype, and that an intact DYX domain is essential for ciliary dynein assembly in pf23 [37]
Summary
Motile cilia ( known as flagella) are antenna-like organelles protruding from many types of cells and required for motility and cell signaling [1, 2]. The movement of motile cilia is driven by the ciliary dynein motors, which are composed of outer (ODA) and inner dynein arms (IDA) [8,9,10]. Multiple non-dynein cytoplasmic proteins have been identified that are required for preassembly of the axonemal dyneins (reviewed in [15, 16]). These include ODA7/LRRC50/DNAAF1, PF13/KTU/DNAAF2, PF22/DNAAF3, HEATR2, IDA10/MOT48, ODA16, ZYMD10, PIH1D3 and LRRC6 [17,18,19,20,21,22,23,24] (see S1 Table). Neither the molecular mechanism of the preassembly process, nor the complete genetic/functional relationship between these factors is known
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