scholarly journals The radial spokes and central apparatus: Mechano-chemical transducers that regulate flagellar motility

2003 ◽  
Vol 57 (1) ◽  
pp. 8-17 ◽  
Author(s):  
Elizabeth F. Smith ◽  
Pinfen Yang
Keyword(s):  
Flagellar Motility ◽  
Radial Spokes ◽  
Central Apparatus ◽  
Chemical Transducers

scholarly journals Regulation of Flagellar Dynein by Calcium and a Role for an Axonemal Calmodulin and Calmodulin-dependent Kinase

2002 ◽  
Vol 13 (9) ◽  
pp. 3303-3313 ◽  
Author(s):  
Elizabeth F. Smith
Keyword(s):  
Calcium Signaling ◽  
Signaling Pathway ◽  
Molecular Mechanism ◽  
Calcium Signal ◽  
Sliding Velocity ◽  
Flagellar Motility ◽  
Wild Type ◽  
Wild Type Level ◽  
Radial Spokes ◽  
Central Apparatus

Ciliary and flagellar motility is regulated by changes in intraflagellar calcium. However, the molecular mechanism by which calcium controls motility is unknown. We tested the hypothesis that calcium regulates motility by controlling dynein-driven microtubule sliding and that the central pair and radial spokes are involved in this regulation. We isolated axonemes from Chlamydomonasmutants and measured microtubule sliding velocity in buffers containing 1 mM ATP and various concentrations of calcium. In buffers with pCa > 8, microtubule sliding velocity in axonemes lacking the central apparatus (pf18 and pf15) was reduced compared with that of wild-type axonemes. In contrast, at pCa4, dynein activity in pf18 and pf15axonemes was restored to wild-type level. The calcium-induced increase in dynein activity in pf18 axonemes was inhibited by antagonists of calmodulin and calmodulin-dependent kinase II. Axonemes lacking the C1 central tubule (pf16) or lacking radial spoke components (pf14 and pf17) do not exhibit calcium-induced increase in dynein activity in pCa4 buffer. We conclude that calcium regulation of flagellar motility involves regulation of dynein-driven microtubule sliding, that calmodulin and calmodulin-dependent kinase II may mediate the calcium signal, and that the central apparatus and radial spokes are key components of the calcium signaling pathway.

scholarly journals Mechanosignaling between central apparatus and radial spokes controls axonemal dynein activity

2014 ◽  
Vol 204 (5) ◽  
pp. 807-819 ◽  
Author(s):  
Toshiyuki Oda ◽  
Haruaki Yanagisawa ◽  
Toshiki Yagi ◽  
Masahide Kikkawa
Keyword(s):  
Regulatory Mechanisms ◽  
Flagellar Motility ◽  
Downstream Effectors ◽  
Concerted Activity ◽  
Radial Spokes ◽  
Axonemal Dynein ◽  
Central Apparatus ◽  
Dynein Arms ◽  
Protein Tags ◽  
Intermolecular Collision

Cilia/flagella are conserved organelles that generate fluid flow in eukaryotes. The bending motion of flagella requires concerted activity of dynein motors. Although it has been reported that the central pair apparatus (CP) and radial spokes (RSs) are important for flagellar motility, the molecular mechanism underlying CP- and RS-mediated dynein regulation has not been identified. In this paper, we identified nonspecific intermolecular collision between CP and RS as one of the regulatory mechanisms for flagellar motility. By combining cryoelectron tomography and motility analyses of Chlamydomonas reinhardtii flagella, we show that binding of streptavidin to RS heads paralyzed flagella. Moreover, the motility defect in a CP projection mutant could be rescued by the addition of exogenous protein tags on RS heads. Genetic experiments demonstrated that outer dynein arms are the major downstream effectors of CP- and RS-mediated regulation of flagellar motility. These results suggest that mechanosignaling between CP and RS regulates dynein activity in eukaryotic flagella.

scholarly journals PF16 encodes a protein with armadillo repeats and localizes to a single microtubule of the central apparatus in Chlamydomonas flagella.

1996 ◽  
Vol 132 (3) ◽  
pp. 359-370 ◽  
Author(s):  
E F Smith ◽  
P A Lefebvre
Keyword(s):  
Protein Interactions ◽  
Insertional Mutagenesis ◽  
Flagellar Motility ◽  
Wild Type ◽  
Protein Protein Interactions ◽  
Cellular Functions ◽  
Central Pair ◽  
Central Apparatus ◽  
Immunofluorescence Labeling ◽  
Armadillo Repeats

Several studies have indicated that the central pair of microtubules and their associated structures play a significant role in regulating flagellar motility. To begin a molecular analysis of these components we have generated central apparatus-defective mutants in Chlamydomonas reinhardtii using insertional mutagenesis. One paralyzed mutant recovered in our screen, D2, is an allele of a previously identified mutant, pf16. Mutant cells have paralyzed flagella, and the C1 microtubule of the central apparatus is missing in isolated axonemes. We have cloned the wild-type PF16 gene and confirmed its identity by rescuing pf16 mutants upon transformation. The rescued pf16 cells were wild-type in motility and in axonemal ultrastructure. A full-length cDNA clone for PF16 was obtained and sequenced. Database searches using the predicted 566 amino acid sequence of PF16 indicate that the protein contains eight contiguous armadillo repeats. A number of proteins with diverse cellular functions also contain armadillo repeats including pendulin, Rch1, importin, SRP-1, and armadillo. An antibody was raised against a fusion protein expressed from the cloned cDNA. Immunofluorescence labeling of wild-type flagella indicates that the PF16 protein is localized along the length of the flagella while immunogold labeling further localizes the PF16 protein to a single microtubule of the central pair. Based on the localization results and the presence of the armadillo repeats in this protein, we suggest that the PF16 gene product is involved in protein-protein interactions important for C1 central microtubule stability and flagellar motility.

scholarly journals Regulation of flagellar motility by the conserved flagellar protein CG34110/Ccdc135/FAP50

2011 ◽  
Vol 22 (7) ◽  
pp. 976-987 ◽  
Author(s):  
Yong Yang ◽  
Deborah A. Cochran ◽  
Mary D. Gargano ◽  
Iryna King ◽  
Nayef K. Samhat ◽  
...  
Keyword(s):  
Flagellar Motility ◽  
Genetic Screens ◽  
Sperm Flagellum ◽  
Respiratory Epithelia ◽  
Downstream Effectors ◽  
Radial Spokes ◽  
Flagellar Proteins ◽  
Flagellar Protein ◽  
Cilia And Flagella ◽  
Lost Boys

Eukaryotic cilia and flagella are vital sensory and motile organelles. The calcium channel PKD2 mediates sensory perception on cilia and flagella, and defects in this can contribute to ciliopathic diseases. Signaling from Pkd2-dependent Ca2+ rise in the cilium to downstream effectors may require intermediary proteins that are largely unknown. To identify these proteins, we carried out genetic screens for mutations affecting Drosophila melanogaster sperm storage, a process mediated by Drosophila Pkd2. Here we show that a new mutation lost boys (lobo) encodes a conserved flagellar protein CG34110, which corresponds to vertebrate Ccdc135 (E = 6e-78) highly expressed in ciliated respiratory epithelia and sperm, and to FAP50 (E = 1e-28) in the Chlamydomonas reinhardtii flagellar proteome. CG34110 localizes along the fly sperm flagellum. FAP50 is tightly associated with the outer doublet microtubules of the axoneme and appears not to be a component of the central pair, radial spokes, dynein arms, or structures defined by the mbo waveform mutants. Phenotypic analyses indicate that both Pkd2 and lobo specifically affect sperm movement into the female storage receptacle. We hypothesize that the CG34110/Ccdc135/FAP50 family of conserved flagellar proteins functions within the axoneme to mediate Pkd2-dependent processes in the sperm flagellum and other motile cilia.

scholarly journals PF15p Is the Chlamydomonas Homologue of the Katanin p80 Subunit and Is Required for Assembly of Flagellar Central Microtubules

2004 ◽  
Vol 3 (4) ◽  
pp. 870-879 ◽  
Author(s):  
Erin E. Dymek ◽  
Paul A. Lefebvre ◽  
Elizabeth F. Smith
Keyword(s):  
Amino Acid ◽  
Significant Role ◽  
Insertional Mutagenesis ◽  
Flagellar Motility ◽  
Wild Type ◽  
Central Pair ◽  
Coding Sequence ◽  
Microtubule Severing ◽  
Central Apparatus

ABSTRACT Numerous studies have indicated that the central apparatus plays a significant role in regulating flagellar motility, yet little is known about how the central pair of microtubules or their associated projections assemble. Several Chlamydomonas mutants are defective in central apparatus assembly. For example, mutant pf15 cells have paralyzed flagella that completely lack the central pair of microtubules. We have cloned the wild-type PF15 gene and confirmed its identity by rescuing the motility and ultrastructural defects in two pf15 alleles, the original pf15a mutant and a mutant generated by insertional mutagenesis. Database searches using the 798-amino-acid polypeptide predicted from the complete coding sequence indicate that the PF15 gene encodes the Chlamydomonas homologue of the katanin p80 subunit. Katanin was originally identified as a heterodimeric protein with a microtubule-severing activity. These results reveal a novel role for the katanin p80 subunit in the assembly and/or stability of the central pair of flagellar microtubules.

scholarly journals The unity and diversity of the ciliary central apparatus

2019 ◽  
Vol 375 (1792) ◽  
pp. 20190164 ◽  
Author(s):  
Lei Zhao ◽  
Yuqing Hou ◽  
Nathan A. McNeill ◽  
George B. Witman
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Recent Work ◽  
Protein Composition ◽  
Model System ◽  
Flagellar Motility ◽  
Phylogenetic Groups ◽  
Wide Range ◽  
Central Apparatus ◽  
Cilia And Flagella ◽  
Eukaryotic Organisms

Nearly all motile cilia and flagella (terms here used interchangeably) have a ‘9+2’ axoneme containing nine outer doublet microtubules and two central microtubules. The central pair of microtubules plus associated projections, termed the central apparatus (CA), is involved in the control of flagellar motility and is essential for the normal movement of ‘9+2’ cilia. Research using the green alga Chlamydomonas reinhardtii , an important model system for studying cilia, has provided most of our knowledge of the protein composition of the CA, and recent work using this organism has expanded the number of known and candidate CA proteins nearly threefold. Here we take advantage of this enhanced proteome to examine the genomes of a wide range of eukaryotic organisms, representing all of the major phylogenetic groups, to identify predicted orthologues of the C. reinhardtii CA proteins and explore how widely the proteins are conserved and whether there are patterns to this conservation. We also discuss in detail two contrasting groups of CA proteins—the ASH-domain proteins, which are broadly conserved, and the PAS proteins, which are restricted primarily to the volvocalean algae. This article is part of the Theo Murphy meeting issue ‘Unity and diversity of cilia in locomotion and transport’.

scholarly journals The CSC connects three major axonemal complexes involved in dynein regulation

2012 ◽  
Vol 23 (16) ◽  
pp. 3143-3155 ◽  
Author(s):  
Thomas Heuser ◽  
Erin E. Dymek ◽  
Jianfeng Lin ◽  
Elizabeth F. Smith ◽  
Daniela Nicastro
Keyword(s):  
Electron Tomography ◽  
Structural Heterogeneity ◽  
Flagellar Motility ◽  
Wild Type ◽  
Localization Model ◽  
Cryo Electron Tomography ◽  
Radial Spokes ◽  
Regulatory Complex ◽  
Health And Development ◽  
Cilia And Flagella

Motile cilia and flagella are highly conserved organelles that play important roles in human health and development. We recently discovered a calmodulin- and spoke-associ­ated complex (CSC) that is required for wild-type motility and for the stable assembly of a subset of radial spokes. Using cryo–electron tomography, we present the first structure-based localization model of the CSC. Chlamydomonas flagella have two full-length radial spokes, RS1 and RS2, and a shorter RS3 homologue, the RS3 stand-in (RS3S). Using newly developed techniques for analyzing samples with structural heterogeneity, we demonstrate that the CSC connects three major axonemal complexes involved in dynein regulation: RS2, the nexin–dynein regulatory complex (N-DRC), and RS3S. These results provide insights into how signals from the radial spokes may be transmitted to the N-DRC and ultimately to the dynein motors. Our results also indicate that although structurally very similar, RS1 and RS2 likely serve different functions in regulating flagellar motility.

The role of central apparatus components in flagellar motility and microtubule assembly

1997 ◽  
Vol 38 (1) ◽  
pp. 1-8 ◽  
Author(s):  
Elizabeth F. Smith ◽  
Paul A. Lefebvre
Keyword(s):  
Microtubule Assembly ◽  
Flagellar Motility ◽  
Central Apparatus
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