Docking-complex-independent alignment of <i>Chlamydomonas</i> outer dynein arms with 24-nm periodicity <i>in vitro</i>

Oda, Toshiyuki; Abe, Tatsuki; Yanagisawa, Haruaki; Kikkawa, Masahide

doi:10.1242/jcs.184598

Cited by 27 publications

(28 citation statements)

References 33 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…It was recently shown that the ODA-DC is more likely to be a flexible stabilizer of ODAs where it strengthens the electrostatic interactions between ODAs and microtubules, rather than a molecular ruler on which ODAs attach [ 34 ]. In contrast to the prior assumption that ODA-DC is an intermediate between ODAs and microtubules, it seems possible that ODAs at this point also interact directly with microtubules.…”

Section: Discussionmentioning

confidence: 99%

Homozygous loss-of-function mutations in MNS1 cause laterality defects and likely male infertility

et al. 2018

View full text Add to dashboard Cite

The clinical spectrum of ciliopathies affecting motile cilia spans impaired mucociliary clearance in the respiratory system, laterality defects including heart malformations, infertility and hydrocephalus. Using linkage analysis and whole exome sequencing, we identified two recessive loss-of-function MNS1 mutations in five individuals from four consanguineous families: 1) a homozygous nonsense mutation p.Arg242* in four males with laterality defects and infertility and 2) a homozygous nonsense mutation p.Gln203* in one female with laterality defects and recurrent respiratory infections additionally carrying homozygous mutations in DNAH5. Consistent with the laterality defects observed in these individuals, we found Mns1 to be expressed in mouse embryonic ventral node. Immunofluorescence analysis further revealed that MNS1 localizes to the axonemes of respiratory cilia as well as sperm flagella in human. In-depth ultrastructural analyses confirmed a subtle outer dynein arm (ODA) defect in the axonemes of respiratory epithelial cells resembling findings reported in Mns1-deficient mice. Ultrastructural analyses in the female carrying combined mutations in MNS1 and DNAH5 indicated a role for MNS1 in the process of ODA docking (ODA-DC) in the distal respiratory axonemes. Furthermore, co-immunoprecipitation and yeast two hybrid analyses demonstrated that MNS1 dimerizes and interacts with the ODA docking complex component CCDC114. Overall, we demonstrate that MNS1 deficiency in humans causes laterality defects (situs inversus) and likely male infertility and that MNS1 plays a role in the ODA-DC assembly.

show abstract

Section: Discussionmentioning

confidence: 99%

Homozygous loss-of-function mutations in MNS1 cause laterality defects and likely male infertility

et al. 2018

View full text Add to dashboard Cite

show abstract

“…Previously, the 24-nm-long ODA docking complex (ODA-DC) was proposed to be responsible for the assembly and 24-nm periodicity of ODAs, but the localization of the ODA-DC remained undetermined (Owa et al 2014). However, it was recently shown that Chlamydomonas ODAs can assemble onto DMTs lacking ODA-DC from the oda3 mutant in vitro, with proper periodicity (Oda et al 2016). In addition, the cryo-ET structure revealed that the part of ODA-DC contacts with DMT near to the microtubule binding site of ODA, but the rest of the complex binds to the outer surface of ODA (Oda et al 2016).…”

Section: Molecular Rulers Facilitate Docking Of Microtubule Associatementioning

confidence: 99%

“…However, it was recently shown that Chlamydomonas ODAs can assemble onto DMTs lacking ODA-DC from the oda3 mutant in vitro, with proper periodicity (Oda et al 2016). In addition, the cryo-ET structure revealed that the part of ODA-DC contacts with DMT near to the microtubule binding site of ODA, but the rest of the complex binds to the outer surface of ODA (Oda et al 2016). Together, this suggested that ODA-DC may not be able to serve as a molecular 11 ruler.…”

Section: Molecular Rulers Facilitate Docking Of Microtubule Associatementioning

confidence: 99%

Structure of the ciliary axoneme at nanometer resolution reconstructed by TYGRESS

Song

Shang

et al. 2018

Preprint

View full text Add to dashboard Cite

The resolution of subtomogram averages calculated from cryo-electron tomograms (cryo-ET) of crowded cellular environments is often limited due to signal loss in, and misalignment of the subtomograms. In contrast, single-particle cryo-electron microcopy (SP-cryo-EM) routinely reaches near-atomic resolution of isolated complexes. We developed a novel hybrid-method called "TomographY-Guided 3D REconstruction of Subcellular Structures" (TYGRESS) that combines cryo-ET with SP-cryo-EM to achieve close-to-nanometer resolution of complexes inside crowded environments. Using TYGRESS, we determined the native 3D structures of the intact ciliary axoneme with up to 12 Å resolution. These results reveal many structures and details that were not visible by cryo-ET. TYGRESS is generally applicable to cellular complexes that are amenable to subtomogram averaging, bringing us a step closer to (pseudo-)atomic models of cells. One Sentence Summary:A hybrid cryo-electron microscopy method reveals subcellular structures at unprecedented resolution. 4 Main Text: Due to recent hardware and image processing advances, single-particle (SP-) cryo-EM can produce 3D reconstructions of purified, native proteins and macromolecular complexes (from ~50 kDa to several MDa size) with near-atomic detail, i.e. with a resolution of 3 Å or better (Bai et al. 2015, Cheng 2015, Danev and Baumeister 2017). Even single-particle cryo-ET of relatively thin (<100 nm) samples containing isolated complexes (Bartesaghi et al. 2012), and of viruses with a high abundance of capsomers (Schur et al. 2016, Wan et al. 2017) has reached sub-nanometer resolution. However, a few hundred nanometer thick and complex cellular samples are notpresently amenable to structural study by SP-cryo-EM. The reason is that for unambiguous particle picking and accurate particle alignment, SP-cryo-EM usually requires the particles to be purified, structurally relatively homogeneous and distributed in a thin monolayer that avoids superposition of particles in the cryo-EM projection images .Cryo-ET is unique in that it can 3D-reconstruct and visualize pleomorphic structures, such as intact cells and organelles in situ. In addition, repeating identical components in the reconstructed tomograms, such as axonemal repeats in cilia or chemoreceptors in bacterial membranes, can be resolved with molecular detail using subtomogram averaging that increased the signal-to-noise ratio and thus resolution of the reconstruction , Briggs 2013). However, the resolution of cellular cryo-ET and subtomogram averaging is ultimately limited by the need to balance several irreconcilable factors (table S1). For example, a higher electron dose improves the signal-to-noise ratio of the tilt images, increasing accuracy of image alignment and correction of the contrast transfer function (CTF), with positive effects on resolution. On the other hand, a higher electron dose also leads to more structural degradation by radiation damage, limiting useful high-resolution signal to the early exposures in a tomogram. In

show abstract

“…Recent studies have revealed that the ODA-DC is not required for the periodicity of the ODA’s [20]; rather, the 24-nm periodicity is an intrinsic feature of the ODAs. Structural studies indicate that the ODA-DC is a flexible structure that serves to enhance the binding of ODAs to the doublet microtubules.…”

Section: Discussionmentioning

confidence: 99%

“…(Note that the nomenclature for the ODA-DC genes and subunits is DCC1 , DCC2 and DLE3 for gene names; oda3 , oda1 and oda14 for mutant alleles; DC1, DC2 and DC3 for proteins [19]). It is required to anchor the ODA to the ciliary outer doublet microtubules [13, 15] but is not required for the 24-nm periodicity of the ODA [20]. …”

Section: Introductionmentioning

confidence: 99%

Characterization of a new oda3 allele, oda3-6, defective in assembly of the outer dynein arm-docking complex in Chlamydomonas reinhardtii

et al. 2017

View full text Add to dashboard Cite

We have used an insertional mutagenesis approach to generate new C. reinhardtii motility mutants. Of 56 mutants isolated, one is a new allele at the ODA3 locus, called oda3-6. Similar to the previously characterized oda3 alleles, oda3-6 has a slow-jerky swimming phenotype and reduced swimming speed. The oda3-6 mutant fails to assemble the outer dynein arm motor and outer dynein arm—docking complex (ODA-DC) in the ciliary axoneme due to an insertion in the 5’ end of the DCC1 gene, which encodes the DC1 subunit of the ODA-DC. Transformation of oda3-6 with the wild-type DCC1 gene rescues the mutant swimming phenotype and restores assembly of the ODA-DC and the outer dynein arm in the cilium. This is the first oda3 mutant to be characterized at the molecular level and is likely to be very useful for further analysis of DC1 function.

show abstract

Docking-complex-independent alignment of Chlamydomonas outer dynein arms with 24-nm periodicity in vitro

Cited by 27 publications

References 33 publications

Homozygous loss-of-function mutations in MNS1 cause laterality defects and likely male infertility

Homozygous loss-of-function mutations in MNS1 cause laterality defects and likely male infertility

Structure of the ciliary axoneme at nanometer resolution reconstructed by TYGRESS

Characterization of a new oda3 allele, oda3-6, defective in assembly of the outer dynein arm-docking complex in Chlamydomonas reinhardtii

Contact Info

Product

Resources

About