scholarly journals Intraflagellar transport (IFT) cargo

2004 ◽  
Vol 164 (2) ◽  
pp. 255-266 ◽  
Author(s):  
Hongmin Qin ◽  
Dennis R. Diener ◽  
Stefan Geimer ◽  
Douglas G. Cole ◽  
Joel L. Rosenbaum
Keyword(s):  
Cell Body ◽  
Soluble Fraction ◽  
Intraflagellar Transport ◽  
Radial Spokes ◽  
Flagellar Assembly ◽  
Protein Particles ◽  
Bidirectional Movement

Intraflagellar transport (IFT) is the bidirectional movement of multisubunit protein particles along axonemal microtubules and is required for assembly and maintenance of eukaryotic flagella and cilia. One posited role of IFT is to transport flagellar precursors to the flagellar tip for assembly. Here, we examine radial spokes, axonemal subunits consisting of 22 polypeptides, as potential cargo for IFT. Radial spokes were found to be partially assembled in the cell body, before being transported to the flagellar tip by anterograde IFT. Fully assembled radial spokes, detached from axonemal microtubules during flagellar breakdown or turnover, are removed from flagella by retrograde IFT. Interactions between IFT particles, motors, radial spokes, and other axonemal proteins were verified by coimmunoprecipitation of these proteins from the soluble fraction of Chlamydomonas flagella. These studies indicate that one of the main roles of IFT in flagellar assembly and maintenance is to transport axonemal proteins in and out of the flagellum.

scholarly journals Chlamydomonas Basal Bodies as Flagella Organizing Centers

Cells ◽  
2018 ◽  
Vol 7 (7) ◽  
pp. 79 ◽  
Author(s):  
Jenna Wingfield ◽  
Karl-Ferdinand Lechtreck
Keyword(s):  
Structure Function ◽  
Chlamydomonas Reinhardtii ◽  
Developmental Disorders ◽  
Intraflagellar Transport ◽  
Specific Protein ◽  
Basal Bodies ◽  
Unicellular Green Alga ◽  
Flagellar Assembly ◽  
Flagellar Axoneme

During ciliogenesis, centrioles convert to membrane-docked basal bodies, which initiate the formation of cilia/flagella and template the nine doublet microtubules of the flagellar axoneme. The discovery that many human diseases and developmental disorders result from defects in flagella has fueled a strong interest in the analysis of flagellar assembly. Here, we will review the structure, function, and development of basal bodies in the unicellular green alga Chlamydomonas reinhardtii, a widely used model for the analysis of basal bodies and flagella. Intraflagellar transport (IFT), a flagella-specific protein shuttle critical for ciliogenesis, was first described in C. reinhardtii. A focus of this review will be on the role of the basal bodies in organizing the IFT machinery.

scholarly journals The role of retrograde intraflagellar transport in flagellar assembly, maintenance, and function

2012 ◽  
Vol 199 (1) ◽  
pp. 151-167 ◽  
Author(s):  
Benjamin D. Engel ◽  
Hiroaki Ishikawa ◽  
Kimberly A. Wemmer ◽  
Stefan Geimer ◽  
Ken-ichi Wakabayashi ◽  
...  
Keyword(s):  
Intraflagellar Transport ◽  
Retrograde Transport ◽  
Full Length ◽  
Temperature Sensitive ◽  
Nonpermissive Temperature ◽  
Flagellar Beat ◽  
Flagellar Assembly ◽  
Ph Shock ◽  
And Function

The maintenance of flagellar length is believed to require both anterograde and retrograde intraflagellar transport (IFT). However, it is difficult to uncouple the functions of retrograde transport from anterograde, as null mutants in dynein heavy chain 1b (DHC1b) have stumpy flagella, demonstrating solely that retrograde IFT is required for flagellar assembly. We isolated a Chlamydomonas reinhardtii mutant (dhc1b-3) with a temperature-sensitive defect in DHC1b, enabling inducible inhibition of retrograde IFT in full-length flagella. Although dhc1b-3 flagella at the nonpermissive temperature (34°C) showed a dramatic reduction of retrograde IFT, they remained nearly full-length for many hours. However, dhc1b-3 cells at 34°C had strong defects in flagellar assembly after cell division or pH shock. Furthermore, dhc1b-3 cells displayed altered phototaxis and flagellar beat. Thus, robust retrograde IFT is required for flagellar assembly and function but is dispensable for the maintenance of flagellar length. Proteomic analysis of dhc1b-3 flagella revealed distinct classes of proteins that change in abundance when retrograde IFT is inhibited.

scholarly journals Chlamydomonas ARMC2/PF27 is an obligate cargo adapter for IFT of radial spokes

2021 ◽  
Author(s):  
Karl F Lechtreck ◽  
Yi Liu ◽  
Jin Dai ◽  
Rama Alkhofash ◽  
Jack Butler ◽  
...  
Keyword(s):  
Lung Function ◽  
Male Infertility ◽  
Cell Body ◽  
Intraflagellar Transport ◽  
Proximal Region ◽  
Repeat Protein ◽  
Radial Spokes ◽  
Armadillo Repeat ◽  
Dynein Arms

Intraflagellar transport (IFT) carries proteins into flagella but how IFT trains interact with the large number of diverse proteins required to assemble flagella remains largely unknown. Here, we show that IFT of radial spokes in Chlamydomonas requires ARMC2/PF27, a conserved armadillo repeat protein associated with male infertility and reduced lung function. Chlamydomonas ARMC2 was highly enriched in growing flagella and tagged ARMC2 and the spoke protein RSP3 comigrated on anterograde trains. In contrast, a cargo and an adapter of inner and outer dynein arms moved independently of ARMC2, indicating that unrelated cargoes distribute stochastically onto the IFT trains. After concomitant unloading at the flagellar tip, RSP3 attached to the axoneme whereas ARMC2 diffused back to the cell body. In armc2/pf27 mutants, IFT of radial spokes was abolished and the presence of radial spokes was limited to the proximal region of flagella. We conclude that ARMC2 is a cargo adapter required for IFT of radial spokes to ensure their assembly along flagella. ARMC2 belongs to a growing class of cargo-specific adapters that enable flagellar transport of preassembled axonemal substructures by IFT.

scholarly journals The GTPase IFT27 is involved in both anterograde and retrograde intraflagellar transport

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Diego Huet ◽  
Thierry Blisnick ◽  
Sylvie Perrot ◽  
Philippe Bastin
Keyword(s):  
Trypanosoma Brucei ◽  
Protein Complexes ◽  
Intraflagellar Transport ◽  
Retrograde Transport ◽  
Small Gtpase ◽  
Anterograde Transport ◽  
Cargo Transport ◽  
Cilia And Flagella ◽  
Bidirectional Movement

The construction of cilia and flagella depends on intraflagellar transport (IFT), the bidirectional movement of two protein complexes (IFT-A and IFT-B) driven by specific kinesin and dynein motors. IFT-B and kinesin are associated to anterograde transport whereas IFT-A and dynein participate to retrograde transport. Surprisingly, the small GTPase IFT27, a member of the IFT-B complex, turns out to be essential for retrograde cargo transport in Trypanosoma brucei. We reveal that this is due to failure to import both the IFT-A complex and the IFT dynein into the flagellar compartment. To get further molecular insight about the role of IFT27, GDP- or GTP-locked versions were expressed in presence or absence of endogenous IFT27. The GDP-locked version is unable to enter the flagellum and to interact with other IFT-B proteins and its sole expression prevents flagellum formation. These findings demonstrate that a GTPase-competent IFT27 is required for association to the IFT complex and that IFT27 plays a role in the cargo loading of the retrograde transport machinery.

scholarly journals The role of the dynein light intermediate chain in retrograde IFT and flagellar function in Chlamydomonas

2016 ◽  
Vol 27 (15) ◽  
pp. 2404-2422 ◽  
Author(s):  
Jaimee Reck ◽  
Alexandria M. Schauer ◽  
Kristyn VanderWaal Mills ◽  
Raqual Bower ◽  
Douglas Tritschler ◽  
...  
Keyword(s):  
Intraflagellar Transport ◽  
Small Subset ◽  
Dependent Manner ◽  
Microtubule Motor ◽  
Flagellar Assembly ◽  
Cilia And Flagella ◽  
The Stability ◽  
Mutant Phenotypes ◽  
Intermediate Chain

The assembly of cilia and flagella depends on the activity of two microtubule motor complexes, kinesin-2 and dynein-2/1b, but the specific functions of the different subunits are poorly defined. Here we analyze Chlamydomonas strains expressing different amounts of the dynein 1b light intermediate chain (D1bLIC). Disruption of D1bLIC alters the stability of the dynein 1b complex and reduces both the frequency and velocity of retrograde intraflagellar transport (IFT), but it does not eliminate retrograde IFT. Flagellar assembly, motility, gliding, and mating are altered in a dose-dependent manner. iTRAQ-based proteomics identifies a small subset of proteins that are significantly reduced or elevated in d1blic flagella. Transformation with D1bLIC-GFP rescues the mutant phenotypes, and D1bLIC-GFP assembles into the dynein 1b complex at wild-type levels. D1bLIC-GFP is transported with anterograde IFT particles to the flagellar tip, dissociates into smaller particles, and begins processive retrograde IFT in <2 s. These studies demonstrate the role of D1bLIC in facilitating the recycling of IFT subunits and other proteins, identify new components potentially involved in the regulation of IFT, flagellar assembly, and flagellar signaling, and provide insight into the role of D1bLIC and retrograde IFT in other organisms.

scholarly journals A Dynein Light Intermediate Chain, D1bLIC, Is Required for Retrograde Intraflagellar Transport

2004 ◽  
Vol 15 (10) ◽  
pp. 4382-4394 ◽  
Author(s):  
Yuqing Hou ◽  
Gregory J. Pazour ◽  
George B. Witman
Keyword(s):  
Intraflagellar Transport ◽  
Cytoplasmic Dynein ◽  
Wild Type ◽  
Phosphate Binding ◽  
Conserved Domain ◽  
Wild Type Phenotype ◽  
Flagellar Assembly ◽  
Bidirectional Movement ◽  
Mutant Cells ◽  
Intermediate Chain

Intraflagellar transport (IFT), the bidirectional movement of particles along flagella, is essential for flagellar assembly. The motor for retrograde IFT in Chlamydomonas is cytoplasmic dynein 1b, which contains the dynein heavy chain DHC1b and the light intermediate chain (LIC) D1bLIC. To investigate a possible role for the LIC in IFT, we identified a d1blic mutant. DHC1b is reduced in the mutant, indicating that D1bLIC is important for stabilizing dynein 1b. The mutant has variable length flagella that accumulate IFT-particle proteins, indicative of a defect in retrograde IFT. Interestingly, the remaining DHC1b is normally distributed in the mutant flagella, strongly suggesting that the defect is in binding of cargo to the retrograde motor rather than in motor activity per se. Cell growth and Golgi apparatus localization and morphology are normal in the mutant, indicating that D1bLIC is involved mainly in retrograde IFT. Like mammalian LICs, D1bLIC has a phosphate-binding domain (P-loop) at its N-terminus. To investigate the function of this conserved domain, d1blic mutant cells were transformed with constructs designed to express D1bLIC proteins with mutated P-loops. The constructs rescued the mutant cells to a wild-type phenotype, indicating that the function of D1bLIC in IFT is independent of its P-loop.

scholarly journals Chlamydomonas ARMC2/PF27 is an obligate cargo adapter for IFT of radial spokes

eLife ◽  
2022 ◽  
Vol 11 ◽  
Author(s):  
Karl F Lechtreck ◽  
Yi Liu ◽  
Jin Dai ◽  
Rama A Alkhofash ◽  
jack Butler ◽  
...  
Keyword(s):  
Lung Function ◽  
Male Infertility ◽  
Cell Body ◽  
Intraflagellar Transport ◽  
Proximal Region ◽  
Repeat Protein ◽  
Radial Spokes ◽  
Armadillo Repeat ◽  
Dynein Arms

Intraflagellar transport (IFT) carries proteins into flagella but how IFT trains interact with the large number of diverse proteins required to assemble flagella remains largely unknown. Here, we show that IFT of radial spokes in Chlamydomonas requires ARMC2/PF27, a conserved armadillo repeat protein associated with male infertility and reduced lung function. Chlamydomonas ARMC2 was highly enriched in growing flagella and tagged ARMC2 and the spoke protein RSP3 comigrated on anterograde trains. In contrast, a cargo and an adapter of inner and outer dynein arms moved independently of ARMC2, indicating that unrelated cargoes distribute stochastically onto the IFT trains. After concomitant unloading at the flagellar tip, RSP3 attached to the axoneme whereas ARMC2 diffused back to the cell body. In armc2/pf27 mutants, IFT of radial spokes was abolished and the presence of radial spokes was limited to the proximal region of flagella. We conclude that ARMC2 is a cargo adapter required for IFT of radial spokes to ensure their assembly along flagella. ARMC2 belongs to a growing class of cargo-specific adapters that enable flagellar transport of preassembled axonemal substructures by IFT.

scholarly journals RABL2 interacts with the intraflagellar transport-B complex and CEP19 and participates in ciliary assembly

2017 ◽  
Vol 28 (12) ◽  
pp. 1652-1666 ◽  
Author(s):  
Yuya Nishijima ◽  
Yohei Hagiya ◽  
Tomohiro Kubo ◽  
Ryota Takei ◽  
Yohei Katoh ◽  
...  
Keyword(s):  
Basal Body ◽  
Intraflagellar Transport ◽  
Bound State ◽  
Dependent Manner ◽  
Centrosomal Protein ◽  
Mother Centriole ◽  
Flagellar Assembly ◽  
Exogenous Expression ◽  
And Function

Proteins localized to the basal body and the centrosome play crucial roles in ciliary assembly and function. Although RABL2 and CEP19 are conserved in ciliated organisms and have been implicated in ciliary/flagellar functions, their roles are poorly understood. Here we show that RABL2 interacts with CEP19 and is recruited to the mother centriole and basal body in a CEP19-dependent manner and that CEP19 is recruited to the centriole probably via its binding to the centrosomal protein FGFR1OP. Disruption of the RABL2 gene in Chlamydomonas reinhardtii results in the nonflagellated phenotype, suggesting a crucial role of RABL2 in ciliary/flagellar assembly. We also show that RABL2 interacts, in its GTP-bound state, with the intraflagellar transport (IFT)-B complex via the IFT74–IFT81 heterodimer and that the interaction is disrupted by a mutation found in male infertile mice (Mot mice) with a sperm flagella motility defect. Intriguingly, RABL2 binds to CEP19 and the IFT74–IFT81 heterodimer in a mutually exclusive manner. Furthermore, exogenous expression of the GDP-locked or Mot-type RABL2 mutant in human cells results in mild defects in ciliary assembly. These results indicate that RABL2 localized to the basal body plays crucial roles in ciliary/flagellar assembly via its interaction with the IFT-B complex.

scholarly journals Protein Particles in Chlamydomonas Flagella Undergo a Transport Cycle Consisting of Four Phases

2001 ◽  
Vol 153 (1) ◽  
pp. 13-24 ◽  
Author(s):  
Carlo Iomini ◽  
Veronica Babaev-Khaimov ◽  
Massimo Sassaroli ◽  
Gianni Piperno
Keyword(s):  
Phase I ◽  
Intraflagellar Transport ◽  
Retrograde Transport ◽  
Cytoplasmic Dynein ◽  
Phase Iii ◽  
Cycle Phase ◽  
Flagellar Assembly ◽  
Protein Particles ◽  
Velocity Changes ◽  
Phase I And Ii

We used an improved procedure to analyze the intraflagellar transport (IFT) of protein particles in Chlamydomonas and found that the frequency of the particles, not only the velocity, changes at each end of the flagella. Thus, particles undergo structural remodeling at both flagellar locations. Therefore, we propose that the IFT consists of a cycle composed of at least four phases: phases II and IV, in which particles undergo anterograde and retrograde transport, respectively, and phases I and III, in which particles are remodeled/exchanged at the proximal and distal end of the flagellum, respectively. In support of our model, we also identified 13 distinct mutants of flagellar assembly (fla), each defective in one or two consecutive phases of the IFT cycle. The phase I-II mutant fla10-1 revealed that cytoplasmic dynein requires the function of kinesin II to participate in the cycle. Phase I and II mutants accumulate complex A, a particle component, near the basal bodies. In contrast, phase III and IV mutants accumulate complex B, a second particle component, in flagellar bulges. Thus, fla mutations affect the function of each complex at different phases of the cycle.

scholarly journals In vivo imaging of radial spoke proteins reveals independent assembly and turnover of the spoke head and stalk

2017 ◽  
Author(s):  
Karl F. Lechtreck ◽  
Ilaria Mengoni ◽  
Batare Okivie ◽  
Kiersten B. Hilderhoff
Keyword(s):  
Exchange Rate ◽  
Cell Body ◽  
In Vivo Imaging ◽  
Fluorescent Protein ◽  
De Novo ◽  
Intraflagellar Transport ◽  
Wild Type ◽  
Radial Spokes ◽  
Head Protein

AbstractRadial spokes (RSs) are multiprotein complexes regulating dynein activity. In the cell body and ciliary matrix, RS proteins are present in a 12S precursor, which is converted into axonemal 20S spokes consisting of a head and stalk. To study RS assembly in vivo, we expressed fluorescent protein (FP)-tagged versions of the head protein RSP4 and the stalk protein RSP3 to rescue the corresponding Chlamydomonas mutants pfl, lacking spoke heads, and pf14, lacking RSs entirely. RSP3 and RSP4 mostly co-migrated by intraflagellar transport (IFT). Transport was elevated during ciliary assembly. IFT of RSP4-FP depended on RSP3. To study RS assembly independently of ciliogenesis, strains expressing FP-tagged RS proteins were mated to untagged cells with, without, or with partial RSs. RSP4-FP is added a tip-to-base fashion to preexisting pf1 spoke stalks while de novo RS assembly occurred lengthwise. In wild-type cilia, the exchange rate of head protein RSP4 exceeded that of the stalk protein RSP3 suggesting increased turnover of spoke heads. The data indicate that RSP3 and RSP4 while transported together separate inside cilia during RS repair and maintenance. The 12S RS precursor encompassing both proteins could represent transport form of the RS ensuring stoichiometric delivery by IFT. (196 of 200)

Sign in / Sign up

Export Citation Format

Share Document