Ccdc113/Ccdc96 complex, a novel regulator of ciliary beating that connects radial spoke 3 to dynein g and the nexin link

Ciliary beating requires the coordinated activity of numerous axonemal complexes. The protein composition and role of radial spokes (RS), nexin links (N-DRC) and dyneins (ODAs and IDAs) is well established. However, how information is transmitted from the central apparatus to the RS and across other ciliary structures remains unclear. Here, we identify a complex comprising the evolutionarily conserved proteins Ccdc96 and Ccdc113, positioned parallel to N-DRC and forming a connection between RS3, dynein g, and N-DRC. Although Ccdc96 and Ccdc113 can be transported to cilia independently, their stable docking and function requires the presence of both proteins. Deletion of either CCDC113 or CCDC96 alters cilia beating frequency, amplitude and waveform. We propose that the Ccdc113/Ccdc96 complex transmits signals from RS3 and N-DRC to dynein g and thus regulates its activity and the ciliary beat pattern.

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LRRC23 is a conserved component of the radial spoke that is necessary for sperm motility and male fertility in mice

Journal of Cell Science ◽

10.1242/jcs.259381 ◽

2021 ◽

Author(s):

Xin Zhang ◽

Jiang Sun ◽

Yonggang Lu ◽

Jintao Zhang ◽

Keisuke Shimada ◽

...

Keyword(s):

Signal Transduction ◽

Male Infertility ◽

Male Fertility ◽

Gene Encoding ◽

Ciliary Beating ◽

Radial Spoke ◽

Evolutionarily Conserved ◽

Radial Spokes ◽

Cilia And Flagella ◽

Mammalian Spermatozoa

Cilia and flagella are ancient structures that achieve controlled motor functions through the coordinated interaction based on microtubules, and some attached projections. Radial spokes (RSs) facilitate the beating motion of these organelles by mediating signal transduction between dyneins and a central pair (CP) of singlet microtubules. RS complex isolation from Chlamydomonas axonemes enabled the detection of 23 radial spoke proteins (RSP1-23), with the roles of some radial spoke proteins remained unknown. Recently, RSP15 has been reported to be located to the stalk of RS2, but its homolog in mammals has not been explored. Herein, we show that Lrrc23 is an evolutionarily conserved testis-enriched gene encoding an RSP15 homolog in mice. We found that LRRC23 localizes to the RS complex within murine sperm flagella and interacts with RSPH3A/B. The knockout of Lrrc23 resulted in male infertility due to RS disorganization and impaired motility in murine spermatozoa, whereas the ciliary beating was unaffected significantly. These data indicate that LRRC23 is a key regulator underpinning the integrity of RS complex within the flagella of mammalian spermatozoa, whereas it is dispensable in cilia.

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Central Apparatus, the Molecular Kickstarter of Ciliary and Flagellar Nanomachines

International Journal of Molecular Sciences ◽

10.3390/ijms22063013 ◽

2021 ◽

Vol 22 (6) ◽

pp. 3013

Author(s):

Zuzanna Samsel ◽

Justyna Sekretarska ◽

Anna Osinka ◽

Dorota Wloga ◽

Ewa Joachimiak

Keyword(s):

Protein Composition ◽

Eukaryotic Cells ◽

Two Phase ◽

Ciliary Beating ◽

Immotile Cilia ◽

Growing Body ◽

Central Apparatus ◽

Motile Cilia

Motile cilia and homologous organelles, the flagella, are an early evolutionarily invention, enabling primitive eukaryotic cells to survive and reproduce. In animals, cilia have undergone functional and structural speciation giving raise to typical motile cilia, motile nodal cilia, and sensory immotile cilia. In contrast to other cilia types, typical motile cilia are able to beat in complex, two-phase movements. Moreover, they contain many additional structures, including central apparatus, composed of two single microtubules connected by a bridge-like structure and assembling numerous complexes called projections. A growing body of evidence supports the important role of the central apparatus in the generation and regulation of the motile cilia movement. Here we review data concerning the central apparatus structure, protein composition, and the significance of its components in ciliary beating regulation.

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Role of the Cilia Membrane in Control of Microtubule Sliding

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s1431927600002683 ◽

1980 ◽

Vol 38 ◽

pp. 612-615

Author(s):

Edna S. Kaneshiro

Keyword(s):

Control Mechanism ◽

Beat Frequency ◽

Surface Membrane ◽

Ciliary Membrane ◽

Ciliary Beating ◽

Ciliary Reversal ◽

Voltage Dependent ◽

Reversal Mechanism ◽

And Function

It is currently believed that ciliary beating results from microtubule sliding which is restricted in regions to cause bending. Cilia beat can be modified to bring about changes in beat frequency, cessation of beat and reversal in beat direction. In ciliated protozoans these modifications which determine swimming behavior have been shown to be related to intracellular (intraciliary) Ca2+ concentrations. The Ca2+ levels are in turn governed by the surface ciliary membrane which exhibits increased Ca2+ conductance (permeability) in response to depolarization. Mutants with altered behaviors have been isolated. Pawn mutants fail to exhibit reversal of the effective stroke of ciliary beat and therefore cannot swim backward. They lack the increased inward Ca2+ current in response to depolarizing stimuli. Both normal and pawn Paramecium made leaky to Ca2+ by Triton extrac¬tion of the surface membrane exhibit backward swimming only in reactivating solutions containing greater than IO-6 M Ca2+ Thus in pawns the ciliary reversal mechanism itself is left operational and only the control mechanism at the membrane is affected. The topographic location of voltage-dependent Ca2+ channels has been identified as a component of the ciliary mem¬brane since the inward Ca2+ conductance response is eliminated by deciliation and the return of the response occurs during cilia regeneration. Since the ciliary membrane has been impli¬cated in the control of Ca2+ levels in the cilium and therefore is the site of at least one kind of control of microtubule sliding, we have focused our attention on understanding the structure and function of the membrane.

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Caenorhabditis elegans DYF-2, an Orthologue of Human WDR19, Is a Component of the Intraflagellar Transport Machinery in Sensory Cilia

Molecular Biology of the Cell ◽

10.1091/mbc.e06-04-0260 ◽

2006 ◽

Vol 17 (11) ◽

pp. 4801-4811 ◽

Cited By ~ 46

Author(s):

Evgeni Efimenko ◽

Oliver E. Blacque ◽

Guangshuo Ou ◽

Courtney J. Haycraft ◽

Bradley K. Yoder ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Aspartic Acid ◽

Critical Role ◽

Intraflagellar Transport ◽

Bardet Biedl Syndrome ◽

Evolutionarily Conserved ◽

Sensory Cilia ◽

Mutant Phenotypes ◽

And Function

The intraflagellar transport (IFT) machinery required to build functional cilia consists of a multisubunit complex whose molecular composition, organization, and function are poorly understood. Here, we describe a novel tryptophan-aspartic acid (WD) repeat (WDR) containing IFT protein from Caenorhabditis elegans, DYF-2, that plays a critical role in maintaining the structural and functional integrity of the IFT machinery. We determined the identity of the dyf-2 gene by transgenic rescue of mutant phenotypes and by sequencing of mutant alleles. Loss of DYF-2 function selectively affects the assembly and motility of different IFT components and leads to defects in cilia structure and chemosensation in the nematode. Based on these observations, and the analysis of DYF-2 movement in a Bardet–Biedl syndrome mutant with partially disrupted IFT particles, we conclude that DYF-2 can associate with IFT particle complex B. At the same time, mutations in dyf-2 can interfere with the function of complex A components, suggesting an important role of this protein in the assembly of the IFT particle as a whole. Importantly, the mouse orthologue of DYF-2, WDR19, also localizes to cilia, pointing to an important evolutionarily conserved role for this WDR protein in cilia development and function.

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HMGN dynamics and chromatin function

Biochemistry and Cell Biology ◽

10.1139/o03-040 ◽

2003 ◽

Vol 81 (3) ◽

pp. 113-122 ◽

Cited By ~ 21

Author(s):

Frédéric Catez ◽

Jae-Hwan Lim ◽

Robert Hock ◽

Yuri V Postnikov ◽

Michael Bustin

Keyword(s):

Binding Sites ◽

Protein Composition ◽

Nuclear Proteins ◽

Chromatin Interactions ◽

Binding Partners ◽

Nucleosomal Dna ◽

Transient Interactions ◽

And Function ◽

Nucleosome Binding

Recent studies indicate that most nuclear proteins, including histone H1 and HMG are highly mobile and their interaction with chromatin is transient. These findings suggest that the structure of chromatin is dynamic and the protein composition at any particular chromatin site is not fixed. Here we discuss how the dynamic behavior of the nucleosome binding HMGN proteins affects the structure and function of chromatin. The high intranuclear mobility of HMGN insures adequate supply of protein throughout the nucleus and serves to target these proteins to their binding sites. Transient interactions of the proteins with nucleosomes destabilize the higher order chromatin, enhance the access to nucleosomal DNA, and impart flexibility to the chromatin fiber. While roaming the nucleus, the HMGN proteins encounter binding partners and form metastable multiprotein complexes, which modulate their chromatin interactions. Studies with HMGN proteins underscore the important role of protein dynamics in chromatin function.Key words: HMG, nuclear proteins, chromatin, HMGN.

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Chlamydomonas flagellar mutants lacking radial spokes and central tubules. Structure, composition, and function of specific axonemal components.

The Journal of Cell Biology ◽

10.1083/jcb.76.3.729 ◽

1978 ◽

Vol 76 (3) ◽

pp. 729-747 ◽

Cited By ~ 256

Author(s):

G B Witman ◽

J Plummer ◽

G Sander

Keyword(s):

Lattice Spacing ◽

Sodium Dodecyl ◽

Internal Standard ◽

Protein Composition ◽

Wild Type ◽

Microscope Examination ◽

Radial Spokes ◽

Dynein Arms ◽

High Molecular Weight Proteins ◽

And Function

The fine structure, protein composition, and roles in flagellar movement of specific axonemal components were studied in wild-type Chlamydomonas and paralyzed mutants pf-14, pf-15A, and pf-19. Electron microscope examination of the isolated axoneme of pf-14 showed that it lacks the radial spokes but is otherwise structurally normal. Comparison of isolated axonemes of wild type and pf-14 by sodium dodecyl sulfate-acrylamide gel electrophoresis indicated that the mutant is missing a protein of 118,000 mol wt; this protein is apparently a major component of the spokes. Pf-15A and pf-19 lack the central tubules and sheath; axonemes of these mutants are missing three high molecular weight proteins which are probably components of the central tubule-central sheath complex. Under conditions where wild-type axonemes reactivated, axonemes of the three mutants remained intact but did not form bends. However, mutant and wild-type axonemes underwent identical adenosine triphosphate-induced disintegration after treatment with trypsin; the dynein arms of the mutants are therefore capable of generating interdoublet shearing forces. These findings indicated that both the radial spokes and the central tubule-central sheath complex are essential for conversion of interdoublet sliding into axonemal bending. Moreover, because axonemes of pf-14 remained intact under reactivating conditions, the nexin links alone are sufficient to limit the amount of interdoublet sliding that occurs. The axial periodicities of the central sheath, dynein arms, radial spokes, and nexin links of Chlamydomonas were determined by electron microscopy using the lattice-spacing of crystalline catalase as an internal standard. Some new ultrastructural details of the components are described.

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TMEM231, mutated in orofaciodigital and Meckel syndromes, organizes the ciliary transition zone

The Journal of Cell Biology ◽

10.1083/jcb.201411087 ◽

2015 ◽

Vol 209 (1) ◽

pp. 129-142 ◽

Cited By ~ 59

Author(s):

Elle C. Roberson ◽

William E. Dowdle ◽

Aysegul Ozanturk ◽

Francesc R. Garcia-Gonzalo ◽

Chunmei Li ◽

...

Keyword(s):

Transition Zone ◽

Transmembrane Protein ◽

Syndrome Type ◽

Zone Formation ◽

Meckel Syndrome ◽

Evolutionarily Conserved ◽

Complex Functions ◽

And Function ◽

Type 3

The Meckel syndrome (MKS) complex functions at the transition zone, located between the basal body and axoneme, to regulate the localization of ciliary membrane proteins. We investigated the role of Tmem231, a two-pass transmembrane protein, in MKS complex formation and function. Consistent with a role in transition zone function, mutation of mouse Tmem231 disrupts the localization of proteins including Arl13b and Inpp5e to cilia, resulting in phenotypes characteristic of MKS such as polydactyly and kidney cysts. Tmem231 and B9d1 are essential for each other and other complex components such as Mks1 to localize to the transition zone. As in mouse, the Caenorhabditis elegans orthologue of Tmem231 localizes to and controls transition zone formation and function, suggesting an evolutionarily conserved role for Tmem231. We identified TMEM231 mutations in orofaciodigital syndrome type 3 (OFD3) and MKS patients that compromise transition zone function. Thus, Tmem231 is critical for organizing the MKS complex and controlling ciliary composition, defects in which cause OFD3 and MKS.

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Recent progress in research on molecular mechanisms of autophagy in the heart

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00711.2014 ◽

2015 ◽

Vol 308 (4) ◽

pp. H259-H268 ◽

Cited By ~ 21

Author(s):

Yasuhiro Maejima ◽

Yun Chen ◽

Mitsuaki Isobe ◽

Åsa B. Gustafsson ◽

Richard N. Kitsis ◽

...

Keyword(s):

Heart Disease ◽

Molecular Mechanisms ◽

Therapeutic Potential ◽

Degradation Process ◽

Structure And Function ◽

Cellular Functions ◽

Evolutionarily Conserved ◽

And Function ◽

Cellular Dysfunction

Dysregulation of autophagy, an evolutionarily conserved process for degradation of long-lived proteins and organelles, has been implicated in the pathogenesis of human disease. Recent research has uncovered pathways that control autophagy in the heart and molecular mechanisms by which alterations in this process affect cardiac structure and function. Although initially thought to be a nonselective degradation process, autophagy, as it has become increasingly clear, can exhibit specificity in the degradation of molecules and organelles, such as mitochondria. Furthermore, it has been shown that autophagy is involved in a wide variety of previously unrecognized cellular functions, such as cell death and metabolism. A growing body of evidence suggests that deviation from appropriate levels of autophagy causes cellular dysfunction and death, which in turn leads to heart disease. Here, we review recent advances in understanding the role of autophagy in heart disease, highlight unsolved issues, and discuss the therapeutic potential of modulating autophagy in heart disease.

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Radial spokes of Chlamydomonas flagella: genetic analysis of assembly and function.

The Journal of Cell Biology ◽

10.1083/jcb.88.1.80 ◽

1981 ◽

Vol 88 (1) ◽

pp. 80-88 ◽

Cited By ~ 111

Author(s):

B Huang ◽

G Piperno ◽

Z Ramanis ◽

D J Luck

Keyword(s):

Gene Product ◽

Biochemical Analysis ◽

Indirect Evidence ◽

Mutant Gene ◽

Molecular Defect ◽

Molecular Weight Range ◽

Radial Spoke ◽

Radial Spokes ◽

And Function

In addition to the previously studied pf-14 and pf-1 loci in Chlamydomonas reinhardtii, mutations for another five genes (pf-17, pf-24, pf-25, pf-26, and pf-27) have been identified and characterized as specifically affecting the assembly and function of the flagellar radial spokes. Mutants for each of the newly identified loci show selective alterations for one or more of the 17 polypeptides in the molecular weight range of 20,000-130,000 which form the radial spoke structure. In specific instances the molecular defect has been correlated with altered radial spoke morphology. Biochemical analysis of in vivo complementation in mutant X wild-type dikaryons has provided indirect evidence that mutations for four of the five new loci (pf-17, pf-24, pf-25, and pf-26) reside in structural genes for spoke components. In the case of pf-24, the identity of the mutant gene product was supported by analysis of induced intragenic revertants. In contrast to the other radial spoke mutants thus far investigated, evidence suggests that the gene product in pf-27 is extrinsic to the radial spokes and is required for the specific in vivo phosphorylation of spoke polypeptides.

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The CSC proteins FAP61 and FAP251 build the basal substructures of radial spoke 3 in cilia

Molecular Biology of the Cell ◽

10.1091/mbc.e14-11-1545 ◽

2015 ◽

Vol 26 (8) ◽

pp. 1463-1475 ◽

Cited By ~ 30

Author(s):

Paulina Urbanska ◽

Kangkang Song ◽

Ewa Joachimiak ◽

Lucja Krzemien-Ojak ◽

Piotr Koprowski ◽

...

Keyword(s):

Radial Spoke ◽

Ciliary Motility ◽

Distinct Structure ◽

Radial Spokes ◽

Dynein Arms ◽

Motile Cilia ◽

The Individual ◽

And Function

Dynein motors and regulatory complexes repeat every 96 nm along the length of motile cilia. Each repeat contains three radial spokes, RS1, RS2, and RS3, which transduct signals between the central microtubules and dynein arms. Each radial spoke has a distinct structure, but little is known about the mechanisms of assembly and function of the individual radial spokes. In Chlamydomonas, calmodulin and spoke-associated complex (CSC) is composed of FAP61, FAP91, and FAP251 and has been linked to the base of RS2 and RS3. We show that in Tetrahymena, loss of either FAP61 or FAP251 reduces cell swimming and affects the ciliary waveform and that RS3 is either missing or incomplete, whereas RS1 and RS2 are unaffected. Specifically, FAP251-null cilia lack an arch-like density at the RS3 base, whereas FAP61-null cilia lack an adjacent portion of the RS3 stem region. This suggests that the CSC proteins are crucial for stable and functional assembly of RS3 and that RS3 and the CSC are important for ciliary motility.

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