Composition and function of the C1b/C1f region in the ciliary central apparatus

Ewa Joachimiak,Bianka Świderska,Hanna Fabczak,Anna Osinka,Dorota Wloga,Martyna Poprzeczko,Hanan Farahat,Ewa Sitkiewicz

doi:10.1038/s41598-021-90996-9

Abstract

Motile cilia are ultrastructurally complex cell organelles with the ability to actively move. The highly conserved central apparatus of motile 9 × 2 + 2 cilia is composed of two microtubules and several large microtubule-bound projections, including the C1b/C1f supercomplex. The composition and function of C1b/C1f subunits has only recently started to emerge. We show that in the model ciliate Tetrahymena thermophila, C1b/C1f contains several evolutionarily conserved proteins: Spef2A, Cfap69, Cfap246/LRGUK, Adgb/androglobin, and a ciliate-specific protein Tt170/TTHERM_00205170. Deletion of genes encoding either Spef2A or Cfap69 led to a loss of the entire C1b projection and resulted in an abnormal vortex motion of cilia. Loss of either Cfap246 or Adgb caused only minor alterations in ciliary motility. Comparative analyses of wild-type and C1b-deficient mutant ciliomes revealed that the levels of subunits forming the adjacent C2b projection but not C1d projection are greatly reduced, indicating that C1b stabilizes C2b. Moreover, the levels of several IFT and BBS proteins, HSP70, and enzymes that catalyze the final steps of the glycolytic pathway: enolase ENO1 and pyruvate kinase PYK1, are also reduced in the C1b-less mutants.

Highlights

Motile cilia are ultrastructurally complex cell organelles with the ability to actively move
When expressed as C-terminal 2V5 or 3HA fusions under control of the native promoter, Spef2A localized in cilia, along their entire length with exception of the distal tip (Fig. 1A-A”)
To better understand the outcome of these mutations, we investigated the effect of corresponding mutations in CFAP69 (Ser249X) and SPEF2A (Lys304X) on Tetrahymena cells motility

Summary

Introduction

Motile cilia are ultrastructurally complex cell organelles with the ability to actively move. Both the outer doublets and the central microtubules serve as scaffolds for periodically attached multi-protein complexes. Smaller structural defects within the C1a-c-e supercomplex such as a loss of a part of the C1c (fap[76] mutant), small parts of C1c and C1e (fap216), entire C1c-e (fap81), or minor defects of C1a (fap92) only reduce the beat frequency and lead to flagella asynchrony[10]. In fap[46] and fap[74] mutants, flagella lack the entire C1d projection and the sheath between C1d and C1b (recently described as C1f3) which causes a range of defects including reduced beat frequency, twitching, and even paralysis.

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Results

Conclusion