scholarly journals A WDR35-dependent coat protein complex transports ciliary membrane cargo vesicles to cilia

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Tooba Quidwai ◽  
Jiaolong Wang ◽  
Emma A Hall ◽  
Narcis A Petriman ◽  
Weihua Leng ◽  
...  
Keyword(s):  
Coat Protein ◽  
Membrane Proteins ◽  
Sequence Homology ◽  
Protein Complex ◽  
Intraflagellar Transport ◽  
Retrograde Transport ◽  
Ciliary Membrane ◽  
Mouse Mutants ◽  
Core Components

Intraflagellar transport (IFT) is a highly conserved mechanism for motor-driven transport of cargo within cilia, but how this cargo is selectively transported to cilia is unclear. WDR35/IFT121 is a component of the IFT-A complex best known for its role in ciliary retrograde transport. In the absence of WDR35, small mutant cilia form but fail to enrich in diverse classes of ciliary membrane proteins. In Wdr35 mouse mutants, the non-core IFT-A components are degraded and core components accumulate at the ciliary base. We reveal deep sequence homology of WDR35 and other IFT-A subunits to α and ß' COPI coatomer subunits, and demonstrate an accumulation of 'coat-less' vesicles which fail to fuse with Wdr35 mutant cilia. We determine that recombinant non-core IFT-As can bind directly to lipids and provide the first in-situ evidence of a novel coat function for WDR35, likely with other IFT-A proteins, in delivering ciliary membrane cargo necessary for cilia elongation.

scholarly journals A WDR35-dependent coatomer transports ciliary membrane proteins from the Golgi to the cilia

2020 ◽  
Author(s):  
Tooba Quidwai ◽  
Emma A. Hall ◽  
Margaret A. Keighren ◽  
Weihua Leng ◽  
Petra Kiesel ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Diffusion Barrier ◽  
Sequence Homology ◽  
Intraflagellar Transport ◽  
Structural Similarity ◽  
Retrograde Transport ◽  
Ciliary Membrane ◽  
Mouse Mutants ◽  
Core Components

AbstractIntraflagellar transport (IFT) is a highly conserved mechanism for motor-driven transport of cargo within cilia, but how this cargo is selectively transported to cilia and across the diffusion barrier is unclear. WDR35/IFT121 is a component of the IFT-A complex best known for its role in ciliary retrograde transport. In the absence of WDR35, small mutant cilia form but fail to enrich in diverse classes of ciliary membrane proteins. In Wdr35 mouse mutants, the IFT-A peripheral components are degraded and core components accumulate at the transition zone. We reveal deep sequence homology and structural similarity of WDR35 and other IFT-As to the coatomer COPI proteins α and β′, and demonstrate an accumulation of ‘coat-less’ vesicles which fail to fuse with Wdr35 mutant cilia. Our data provides the first in situ evidence of a novel coatomer function for WDR35 likely with other IFT-A proteins in delivering ciliary membrane cargo from the Golgi necessary for cilia elongation.

scholarly journals Flagella Ca2+ elevations regulate pausing of retrograde intraflagellar transport trains in adherent Chlamydomonas flagella

2020 ◽  
Author(s):  
Cecile Fort ◽  
Peter Collingridge ◽  
Colin Brownlee ◽  
Glen Wheeler
Keyword(s):  
Membrane Proteins ◽  
Green Alga ◽  
High Frequency ◽  
Intraflagellar Transport ◽  
Gliding Motility ◽  
Membrane Glycoproteins ◽  
Ciliary Membrane ◽  
Process Uncertainty ◽  
Temporal Properties ◽  
Transient Interactions

AbstractThe movement of ciliary membrane proteins is directed by transient interactions with intraflagellar transport (IFT) trains. The green alga Chlamydomonas has adapted this process for gliding motility, using IFT to move adhesive glycoproteins (FMG-1B) in the flagella membrane. Although Ca2+ signalling contributes directly to the gliding process, uncertainty remains over the mechanisms through which Ca2+ acts to influence the movement of IFT trains. Here we show that flagella Ca2+ elevations regulate IFT primarily by initiating the movement of paused retrograde IFT trains. Flagella Ca2+ elevations exhibit complex spatial and temporal properties, including high frequency repetitive Ca2+ elevations that prevent the accumulation of paused retrograde IFT trains. We show that flagella Ca2+ elevations disrupt the IFT-dependent movement of microspheres along the flagella membrane. The results suggest that flagella Ca2+ elevations directly disrupt the interaction between retrograde IFT particles and flagella membrane glycoproteins to modulate gliding motility and the adhesion of the flagellum to a surface.

scholarly journals Formation of the B9-domain protein complex MKS1–B9D2–B9D1 is essential as a diffusion barrier for ciliary membrane proteins

2020 ◽  
Vol 31 (20) ◽  
pp. 2259-2268
Author(s):  
Misato Okazaki ◽  
Takuya Kobayashi ◽  
Shuhei Chiba ◽  
Ryota Takei ◽  
Luxiaoxue Liang ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Transition Zone ◽  
Diffusion Barrier ◽  
Protein Complex ◽  
Ciliary Membrane ◽  
Meckel Syndrome ◽  
Interaction Mode ◽  
Domain Protein

Meckel syndrome (MKS)1, B9 domain (B9D)1, and B9D2 are soluble transition zone (TZ) proteins and share a B9D. We demonstrate the interaction mode of these B9D proteins to be MKS1-B9D2-B9D1 and their interdependent localization to the TZ. We also show that formation of the B9D protein complex is crucial for creating a diffusion barrier for ciliary membrane proteins.

scholarly journals Single molecule imaging reveals a major role for diffusion in the exploration of ciliary space by signaling receptors

eLife ◽  
2013 ◽  
Vol 2 ◽  
Author(s):  
Fan Ye ◽  
David K Breslow ◽  
Elena F Koslover ◽  
Andrew J Spakowitz ◽  
W James Nelson ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Active Transport ◽  
Single Molecule ◽  
Primary Cilia ◽  
Somatostatin Receptor ◽  
Intraflagellar Transport ◽  
Signal Propagation ◽  
Protein Diffusion ◽  
Ciliary Membrane ◽  
Signaling Receptors

The dynamic organization of signaling cascades inside primary cilia is key to signal propagation. Yet little is known about the dynamics of ciliary membrane proteins besides a possible role for motor-driven Intraflagellar Transport (IFT). To characterize these dynamics, we imaged single molecules of Somatostatin Receptor 3 (SSTR3, a GPCR) and Smoothened (Smo, a Hedgehog signal transducer) in the ciliary membrane. While IFT trains moved processively from one end of the cilium to the other, single SSTR3 and Smo underwent mostly diffusive behavior interspersed with short periods of directional movements. Statistical subtraction of instant velocities revealed that SSTR3 and Smo spent less than a third of their time undergoing active transport. Finally, SSTR3 and IFT movements could be uncoupled by perturbing either membrane protein diffusion or active transport. Thus ciliary membrane proteins move predominantly by diffusion, and attachment to IFT trains is transient and stochastic rather than processive or spatially determined.

scholarly journals The Bardet–Biedl syndrome protein complex is an adapter expanding the cargo range of intraflagellar transport trains for ciliary export

2018 ◽  
Vol 115 (5) ◽  
pp. E934-E943 ◽  
Author(s):  
Peiwei Liu ◽  
Karl F. Lechtreck
Keyword(s):  
Protein Complex ◽  
Intraflagellar Transport ◽  
Negative Regulator ◽  
Membrane Composition ◽  
Bardet Biedl Syndrome ◽  
Ciliary Membrane ◽  
Phototactic Behavior ◽  
Dependent Transport ◽  
Particle Imaging ◽  
Single Particle Imaging

Bardet–Biedl syndrome (BBS) is a ciliopathy resulting from defects in the BBSome, a conserved protein complex. BBSome mutations affect ciliary membrane composition, impairing cilia-based signaling. The mechanism by which the BBSome regulates ciliary membrane content remains unknown. Chlamydomonas bbs mutants lack phototaxis and accumulate phospholipase D (PLD) in the ciliary membrane. Single particle imaging revealed that PLD comigrates with BBS4 by intraflagellar transport (IFT) while IFT of PLD is abolished in bbs mutants. BBSome deficiency did not alter the rate of PLD entry into cilia. Membrane association and the N-terminal 58 residues of PLD are sufficient and necessary for BBSome-dependent transport and ciliary export. The replacement of PLD’s ciliary export sequence (CES) caused PLD to accumulate in cilia of cells with intact BBSomes and IFT. The buildup of PLD inside cilia impaired phototaxis, revealing that PLD is a negative regulator of phototactic behavior. We conclude that the BBSome is a cargo adapter ensuring ciliary export of PLD on IFT trains to regulate phototaxis.

scholarly journals Trafficking of ciliary membrane proteins by the intraflagellar transport/BBSome machinery

2018 ◽  
Vol 62 (6) ◽  
pp. 753-763 ◽  
Author(s):  
Jenna L. Wingfield ◽  
Karl-Ferdinand Lechtreck ◽  
Esben Lorentzen
Keyword(s):  
Membrane Proteins ◽  
Molecular Motors ◽  
Intraflagellar Transport ◽  
Inherited Disease ◽  
Cell Organelles ◽  
Bardet Biedl Syndrome ◽  
Ciliary Membrane ◽  
Peripheral Membrane Proteins ◽  
Abnormal Accumulation ◽  
And Function

Bardet–Biedl syndrome (BBS) is a rare inherited disease caused by defects in the BBSome, an octameric complex of BBS proteins. The BBSome is conserved in most organisms with cilia, which are microtubule (MT)-based cell organelles that protrude from the cell surface and function in motility and sensing. Cilia assembly, maintenance, and function require intraflagellar transport (IFT), a bidirectional motility of multi-megadalton IFT trains propelled by molecular motors along the ciliary MTs. IFT has been shown to transport structural proteins, including tubulin, into growing cilia. The BBSome is an adapter for the transport of ciliary membrane proteins and cycles through cilia via IFT. While both the loss and the abnormal accumulation of ciliary membrane proteins have been observed in bbs mutants, recent data converge on a model where the BBSome mainly functions as a cargo adapter for the removal of certain transmembrane and peripheral membrane proteins from cilia. Here, we review recent data on the ultrastructure of the BBSome and how the BBSome recognizes its cargoes and mediates their removal from cilia.

scholarly journals Ca2+ elevations disrupt interactions between intraflagellar transport and the flagella membrane in Chlamydomonas

2021 ◽  
Vol 134 (3) ◽  
pp. jcs253492
Author(s):  
Cecile Fort ◽  
Peter Collingridge ◽  
Colin Brownlee ◽  
Glen Wheeler
Keyword(s):  
Membrane Proteins ◽  
Green Alga ◽  
High Frequency ◽  
Traction Force ◽  
Intraflagellar Transport ◽  
Gliding Motility ◽  
Membrane Glycoproteins ◽  
Ciliary Membrane ◽  
Precise Control ◽  
Transient Interactions

ABSTRACTThe movement of ciliary membrane proteins is directed by transient interactions with intraflagellar transport (IFT) trains. The green alga Chlamydomonas has adapted this process for gliding motility, using retrograde IFT motors to move adhesive glycoproteins in the flagella membrane. Ca2+ signalling contributes directly to the gliding process, although uncertainty remains over the mechanism through which it acts. Here, we show that flagella Ca2+ elevations initiate the movement of paused retrograde IFT trains, which accumulate at the distal end of adherent flagella, but do not influence other IFT processes. On highly adherent surfaces, flagella exhibit high-frequency Ca2+ elevations that prevent the accumulation of paused retrograde IFT trains. Flagella Ca2+ elevations disrupt the IFT-dependent movement of microspheres along the flagella membrane, suggesting that Ca2+ acts by directly disrupting an interaction between retrograde IFT trains and flagella membrane glycoproteins. By regulating the extent to which glycoproteins on the flagella surface interact with IFT motor proteins on the axoneme, this signalling mechanism allows precise control of traction force and gliding motility in adherent flagella.

scholarly journals Tyrosine Phosphorylation of Selected Secretory Carrier Membrane Proteins, SCAMP1 and SCAMP3, and Association with the EGF Receptor

1998 ◽  
Vol 9 (7) ◽  
pp. 1661-1674 ◽  
Author(s):  
Theodore T. Wu ◽  
J. David Castle
Keyword(s):  
Membrane Proteins ◽  
Egf Receptor ◽  
Tyrosine Phosphatase ◽  
Chinese Hamster ◽  
Tyrosine Residue ◽  
Cell Interior ◽  
Murine Fibroblasts ◽  
Secretory Carrier Membrane Proteins

Secretory carrier membrane proteins (SCAMPs) are ubiquitously expressed proteins of post-Golgi vesicles. In the presence of the tyrosine phosphatase inhibitor vanadate, or after overexpression in Chinese hamster ovary (CHO) cells, SCAMP1 and SCAMP3 are phosphorylated selectively on tyrosine residue(s). Phosphorylation is reversible after vanadate washout in situ or when isolated SCAMP3 is incubated with the recombinant tyrosine phosphatase PTP1B. Vanadate also causes the partial accumulation of SCAMP3, but not SCAMP1, in “patches” at or near the cell surface. A search for SCAMP kinase activities has shown that SCAMPs 1 and 3, but not SCAMP2, are tyrosine phosphorylated in EGF-stimulated murine fibroblasts overexpressing the EGF receptor (EGFR). EGF catalyzes the progressive phosphorylation of the SCAMPs up to 1 h poststimulation and may enhance colocalization of the EGFR and SCAMP3 within the cell interior. EGF also induces SCAMP–EGFR association, as detected by coimmunoprecipitation, and phosphorylation of SCAMP3 is stimulated by the EGFR in vitro. These results suggest that phosphorylation of SCAMPs, either directly or indirectly, may be functionally linked to the internalization/down-regulation of the EGFR.

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