Dataset Information

Structural organization of the intermediate and light chain complex of Chlamydomonas ciliary I1 dynein.

ABSTRACT: Axonemal I1 dynein (dynein f) is the largest inner dynein arm in cilia and a key regulator of ciliary beating. It consists of two dynein heavy chains, and an intermediate chain/light chain (ICLC) complex. However, the structural organization of the nine ICLC subunits remains largely unknown. Here, we used biochemical and genetic approaches, and cryo-electron tomography imaging in Chlamydomonas to dissect the molecular architecture of the I1 dynein ICLC complex. Using a strain expressing SNAP-tagged IC140, tomography revealed the location of the IC140 N-terminus at the proximal apex of the ICLC structure. Mass spectrometry of a tctex2b mutant showed that TCTEX2B dynein light chain is required for the stable assembly of TCTEX1 and inner dynein arm interacting proteins IC97 and FAP120. The structural defects observed in tctex2b located these 4 subunits in the center and bottom regions of the ICLC structure, which overlaps with the location of the IC138 regulatory subcomplex, which contains IC138, IC97, FAP120, and LC7b. These results reveal the three-dimensional organization of the native ICLC complex and indicate potential protein-protein interactions that are involved in the pathway by which I1 regulates ciliary motility.

SUBMITTER: Fu G

PROVIDER: S-EPMC8820979 | biostudies-literature | 2021 Jun

REPOSITORIES: biostudies-literature

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Structural organization of the intermediate and light chain complex of Chlamydomonas ciliary I1 dynein.

Fu Gang G Scarbrough Chasity C Song Kangkang K Phan Nhan N Wirschell Maureen M Nicastro Daniela D

FASEB journal : official publication of the Federation of American Societies for Experimental Biology 20210601 6

Axonemal I1 dynein (dynein f) is the largest inner dynein arm in cilia and a key regulator of ciliary beating. It consists of two dynein heavy chains, and an intermediate chain/light chain (ICLC) complex. However, the structural organization of the nine ICLC subunits remains largely unknown. Here, we used biochemical and genetic approaches, and cryo-electron tomography imaging in Chlamydomonas to dissect the molecular architecture of the I1 dynein ICLC complex. Using a strain expressing SNAP-tag ...[more]

PMID: 33993568

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Project description:Motile flagella (also called "motile cilia") play a variety of important roles in lower and higher eukaryotes, including cellular motility and fertility. Flagellar motility is driven by several species of the gigantic motor-protein complexes, flagellar dyneins, that reside within these organelles. Among the flagellar-dynein species, a hetero-dimeric dynein called "IDA f/I1" has been shown to be particularly important in controlling the flagellar waveform, and defects in this dynein species in humans cause ciliopathies such as multiple morphological abnormalities of the flagella and asthenoteratozoospermia. IDA f/I1 is composed of many subunits, including two HCs (HCα and HCβ) and three ICs (IC140, IC138, and IC97), and among the three ICs of IDA f/I1, the exact molecular function(s) of IC97 in flagellar motility is not well understood. In this study, we isolated a Chlamydomonas mutant lacking IC97 and analyzed the phenotypes. The ic97 mutant phenocopied several aspects of the previously isolated IDA-f/I1-related mutants in Chlamydomonas and showed slow swimming compared to the wild type but retained the ability to phototaxis. Further analysis revealed that the mutant had low flagellar beat frequency and miscoordination between the two (cis- and trans-) flagella. In addition, the mutant cells swam in a comparatively straight path compared to the wild-type cells. Taken together, our results highlight the importance of proper assembly of IC97 in the IDA-f/I1 complex for the regulation of flagellar and cellular motility in Chlamydomonas and provide valuable insights into both the molecular functions of IC97 orthologs in higher eukaryotes and the pathogenetic mechanisms of human ciliopathies caused by IDA-f/I1 defects.ImportanceIDA f/I1 is a hetero-dimeric flagellar dynein that is particularly important for the regulation of flagellar waveform and whose defects are associated with human ciliopathies. IC97 is an evolutionarily conserved intermediate chain of IDA f/I1, but the detailed molecular functions of IC97 in flagellar motility have not been elucidated. In this study, mutational and biochemical analyses of the previously uncharacterized Chlamydomonas ic97 mutant revealed that IC97 is required for both the normal flagellar and cellular motility. In particular, IC97 appears to play an important role in both the control of flagellar beat frequency and the coordination between the two (cis- and trans-) flagella in Chlamydomonas. Our results provide important insights into the regulation of IDA-f/I1 activity by IC97 and the pathogenetic mechanisms of human ciliopathies caused by IDA-f/I1 defects.

Dataset Information

Structural organization of the intermediate and light chain complex of Chlamydomonas ciliary I1 dynein.

Publications

Structural organization of the intermediate and light chain complex of Chlamydomonas ciliary I1 dynein.

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