scholarly journals Dynein-2 intermediate chains play crucial but distinct roles in primary cilia formation and function

2018 ◽  
Author(s):  
Laura Vuolo ◽  
Nicola L. Stevenson ◽  
Kate J. Heesom ◽  
David J. Stephens
Keyword(s):  
Transition Zone ◽  
Cell Lines ◽  
Primary Cilia ◽  
Point Mutations ◽  
Intraflagellar Transport ◽  
Jeune Syndrome ◽  
Cilia Formation ◽  
Microtubule Motor ◽  
And Function ◽  
Intermediate Chains

AbstractThe dynein-2 microtubule motor is the retrograde motor for intraflagellar transport. Mutations in dynein-2 components cause skeletal ciliopathies, notably Jeune syndrome. Dynein-2 comprises a heterodimer of two non-identical intermediate chains, WDR34 and WDR60. Here, we use knockout cell lines to demonstrate that each intermediate chain has a distinct role in cilia function. Both proteins are required to maintain a functional transition zone and for efficient bidirectional intraflagellar transport, only WDR34 is essential for axoneme extension. In contrast, only WDR60 is essential for co-assembly of the other subunits. Furthermore, WDR60 cannot compensate for loss of WDR34 or vice versa. This work defines a functional asymmetry to match the subunit asymmetry within the dynein-2 motor. Analysis of causative point mutations in WDR34 and WDR60 can partially restore function to knockout cells. Our data show that Jeune syndrome is caused by defects in transition zone architecture as well as intraflagellar transport.SUMMARYHere, Vuolo and colleagues use engineered knockout human cell lines to define roles for dynein-2 intermediate chains. WDR34 is required for axoneme extension, while WDR60 is not. Both subunits are required for cilia transition zone organization and bidirectional intraflagellar transport.

scholarly journals Dynein-2 intermediate chains play crucial but distinct roles in primary cilia formation and function

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Laura Vuolo ◽  
Nicola L Stevenson ◽  
Kate J Heesom ◽  
David J Stephens
Keyword(s):  
Transition Zone ◽  
Primary Cilia ◽  
Complex Function ◽  
Intraflagellar Transport ◽  
Loss Of Function ◽  
Jeune Syndrome ◽  
Cilia Formation ◽  
Microtubule Motor ◽  
And Function ◽  
Intermediate Chains

The dynein-2 microtubule motor is the retrograde motor for intraflagellar transport. Mutations in dynein-2 components cause skeletal ciliopathies, notably Jeune syndrome. Dynein-2 contains a heterodimer of two non-identical intermediate chains, WDR34 and WDR60. Here, we use knockout cell lines to demonstrate that each intermediate chain has a distinct role in cilium function. Using quantitative proteomics, we show that WDR34 KO cells can assemble a dynein-2 motor complex that binds IFT proteins yet fails to extend an axoneme, indicating complex function is stalled. In contrast, WDR60 KO cells do extend axonemes but show reduced assembly of dynein-2 and binding to IFT proteins. Both proteins are required to maintain a functional transition zone and for efficient bidirectional intraflagellar transport. Our results indicate that the subunit asymmetry within the dynein-2 complex is matched with a functional asymmetry between the dynein-2 intermediate chains. Furthermore, this work reveals that loss of function of dynein-2 leads to defects in transition zone architecture, as well as intraflagellar transport.

scholarly journals STORM imaging reveals the spatial arrangement of transition zone components and IFT particles at the ciliary base in Tetrahymena

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Khodor S. Hazime ◽  
Zhu Zhou ◽  
Ewa Joachimiak ◽  
Natalia A. Bulgakova ◽  
Dorota Wloga ◽  
...  
Keyword(s):  
Transition Zone ◽  
Spatial Organization ◽  
Intraflagellar Transport ◽  
Spatial Arrangement ◽  
Interaction Sites ◽  
Narrow Region ◽  
Cilia Formation ◽  
Axial Dimension ◽  
Optical Reconstruction ◽  
And Function

AbstractThe base of the cilium comprising the transition zone (TZ) and transition fibers (TF) acts as a selecting gate to regulate the intraflagellar transport (IFT)-dependent trafficking of proteins to and from cilia. Before entering the ciliary compartment, IFT complexes and transported cargoes accumulate at or near the base of the cilium. The spatial organization of IFT proteins at the cilia base is key for understanding cilia formation and function. Using stochastic optical reconstruction microscopy (STORM) and computational averaging, we show that seven TZ, nine IFT, three Bardet–Biedl syndrome (BBS), and one centrosomal protein, form 9-clustered rings at the cilium base of a ciliate Tetrahymena thermophila. In the axial dimension, analyzed TZ proteins localize to a narrow region of about 30 nm while IFT proteins dock approximately 80 nm proximal to TZ. Moreover, the IFT-A subcomplex is positioned peripheral to the IFT-B subcomplex and the investigated BBS proteins localize near the ciliary membrane. The positioning of the HA-tagged N- and C-termini of the selected proteins enabled the prediction of the spatial orientation of protein particles and likely cargo interaction sites. Based on the obtained data, we built a comprehensive 3D-model showing the arrangement of the investigated ciliary proteins.

scholarly journals Centrioles initiate cilia assembly but are dispensable for maturation and maintenance in C. elegans

2017 ◽  
Vol 216 (6) ◽  
pp. 1659-1671 ◽  
Author(s):  
Daniel Serwas ◽  
Tiffany Y. Su ◽  
Max Roessler ◽  
Shaohe Wang ◽  
Alexander Dammermann
Keyword(s):  
Fluorescence Microscopy ◽  
Transition Zone ◽  
Electron Tomography ◽  
Intraflagellar Transport ◽  
Basal Bodies ◽  
C Elegans ◽  
Cilia Formation ◽  
Syndrome Protein

Cilia are cellular projections that assemble on centriole-derived basal bodies. While cilia assembly is absolutely dependent on centrioles, it is not known to what extent they contribute to downstream events. The nematode C. elegans provides a unique opportunity to address this question, as centrioles do not persist at the base of mature cilia. Using fluorescence microscopy and electron tomography, we find that centrioles degenerate early during ciliogenesis. The transition zone and axoneme are not completely formed at this time, indicating that cilia maturation does not depend on intact centrioles. The hydrolethalus syndrome protein HYLS-1 is the only centriolar protein known to remain at the base of mature cilia and is required for intraflagellar transport trafficking. Surprisingly, targeted degradation of HYLS-1 after initiation of ciliogenesis does not affect ciliary structures. Taken together, our results indicate that while centrioles are essential to initiate cilia formation, they are dispensable for cilia maturation and maintenance.

scholarly journals NRF2-dependent gene expression promotes ciliogenesis and Hedgehog signaling

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Ana Martin-Hurtado ◽  
Raquel Martin-Morales ◽  
Natalia Robledinos-Antón ◽  
Ruth Blanco ◽  
Ines Palacios-Blanco ◽  
...  
Keyword(s):  
Hedgehog Signaling ◽  
Primary Cilia ◽  
Embryonic Stem ◽  
Mtor Inhibitors ◽  
Gene Promoters ◽  
Protein Levels ◽  
Functional Connections ◽  
Differentiated Cells ◽  
Cilia Formation ◽  
And Function

Abstract The transcription factor NRF2 is a master regulator of cellular antioxidant and detoxification responses, but it also regulates other processes such as autophagy and pluripotency. In human embryonic stem cells (hESCs), NRF2 antagonizes neuroectoderm differentiation, which only occurs after NRF2 is repressed via a Primary Cilia-Autophagy-NRF2 (PAN) axis. However, the functional connections between NRF2 and primary cilia, microtubule-based plasma membrane protrusions that function as cellular antennae, remain poorly understood. For instance, nothing is known about whether NRF2 affects cilia, or whether cilia regulation of NRF2 extends beyond hESCs. Here, we show that NRF2 and primary cilia reciprocally regulate each other. First, we demonstrate that fibroblasts lacking primary cilia have higher NRF2 activity, which is rescued by autophagy-activating mTOR inhibitors, indicating that the PAN axis also operates in differentiated cells. Furthermore, NRF2 controls cilia formation and function. NRF2-null cells grow fewer and shorter cilia and display impaired Hedgehog signaling, a cilia-dependent pathway. These defects are not due to increased oxidative stress or ciliophagy, but rather to NRF2 promoting expression of multiple ciliogenic and Hedgehog pathway genes. Among these, we focused on GLI2 and GLI3, the transcription factors controlling Hh pathway output. Both their mRNA and protein levels are reduced in NRF2-null cells, consistent with their gene promoters containing consensus ARE sequences predicted to bind NRF2. Moreover, GLI2 and GLI3 fail to accumulate at the ciliary tip of NRF2-null cells upon Hh pathway activation. Given the importance of NRF2 and ciliary signaling in human disease, our data may have important biomedical implications.

scholarly journals Signaling compartment at the ciliary tip is formed and maintained by intraflagellar transport and functions as sensitive salt detector

2019 ◽  
Author(s):  
Servaas N. van der Burght ◽  
Suzanne Rademakers ◽  
Jacque-Lynne Johnson ◽  
Chunmei Li ◽  
Gert-Jan Kremers ◽  
...  
Keyword(s):  
Primary Cilia ◽  
Physiological Function ◽  
Intraflagellar Transport ◽  
Distal Region ◽  
Central Question ◽  
Membrane Diffusion ◽  
High Concentration ◽  
C Elegans ◽  
Cgmp Signaling ◽  
And Function

AbstractPrimary cilia are ubiquitous antenna-like organelles that mediate cellular signaling and represent hotspots for human diseases termed ciliopathies. How signaling subcompartments are established within the microtubule-based organelle, and for example support Hedgehog or cGMP signal transduction pathways, remains a central question. Here we show that a C. elegans salt-sensing receptor type guanylate cyclase, GCY-22, accumulates at a high concentration within the distal region of the cilium. This receptor uses DAF-25 (Ankmy2 in mammals) to cross the transition zone (TZ) membrane diffusion barrier in the proximal-most region of the ciliary axoneme. Targeting of GCY-22 to the ciliary tip is dynamic, requiring the cargo-mobilizing intraflagellar transport (IFT) system. Disruption of transit across the TZ barrier or IFT trafficking causes GCY-22 protein mislocalization and defects in the formation, maintenance, and function of the ciliary tip compartment required for chemotaxis to low NaCl concentrations. Together, our findings reveal how a previously undescribed cilium tip cGMP signaling compartment is established and contributes to the physiological function of a primary cilium.

scholarly journals Phosphoinositide lipids in primary cilia biology

2020 ◽  
Vol 477 (18) ◽  
pp. 3541-3565
Author(s):  
Sarah E. Conduit ◽  
Bart Vanhaesebroeck
Keyword(s):  
Primary Cilia ◽  
Critical Role ◽  
Phospholipid Composition ◽  
Cell Types ◽  
Phosphoinositide Metabolism ◽  
Lipid Phosphatases ◽  
Regulatory Enzymes ◽  
Lipid Species ◽  
Cilia Formation ◽  
And Function

Primary cilia are solitary signalling organelles projecting from the surface of most cell types. Although the ciliary membrane is continuous with the plasma membrane it exhibits a unique phospholipid composition, a feature essential for normal cilia formation and function. Recent studies have illustrated that distinct phosphoinositide lipid species localise to specific cilia subdomains, and have begun to build a ‘phosphoinositide map’ of the cilium. The abundance and localisation of phosphoinositides are tightly regulated by the opposing actions of lipid kinases and lipid phosphatases that have also been recently discovered at cilia. The critical role of phosphoinositides in cilia biology is highlighted by the devastating consequences of genetic defects in cilia-associated phosphoinositide regulatory enzymes leading to ciliopathy phenotypes in humans and experimental mouse and zebrafish models. Here we provide a general introduction to primary cilia and the roles phosphoinositides play in cilia biology. In addition to increasing our understanding of fundamental cilia biology, this rapidly expanding field may inform novel approaches to treat ciliopathy syndromes caused by deregulated phosphoinositide metabolism.

scholarly journals Aurora A and AKT Kinase Signaling Associated with Primary Cilia

Cells ◽  
2021 ◽  
Vol 10 (12) ◽  
pp. 3602
Author(s):  
Yuhei Nishimura ◽  
Daishi Yamakawa ◽  
Takashi Shiromizu ◽  
Masaki Inagaki
Keyword(s):  
Primary Cilia ◽  
Tissue Type ◽  
Aurora A ◽  
Kinase Signaling ◽  
Cellular Processes ◽  
Pathological Conditions ◽  
Cell Tissue ◽  
Selective Targeting ◽  
Cilia Formation ◽  
And Function

Dysregulation of kinase signaling is associated with various pathological conditions, including cancer, inflammation, and autoimmunity; consequently, the kinases involved have become major therapeutic targets. While kinase signaling pathways play crucial roles in multiple cellular processes, the precise manner in which their dysregulation contributes to disease is dependent on the context; for example, the cell/tissue type or subcellular localization of the kinase or substrate. Thus, context-selective targeting of dysregulated kinases may serve to increase the therapeutic specificity while reducing off-target adverse effects. Primary cilia are antenna-like structures that extend from the plasma membrane and function by detecting extracellular cues and transducing signals into the cell. Cilia formation and signaling are dynamically regulated through context-dependent mechanisms; as such, dysregulation of primary cilia contributes to disease in a variety of ways. Here, we review the involvement of primary cilia-associated signaling through aurora A and AKT kinases with respect to cancer, obesity, and other ciliopathies.

Role of Primary Cilia in Bone and Cartilage

2021 ◽  
pp. 002203452110466
Author(s):  
Z. Chinipardaz ◽  
M. Liu ◽  
D.T. Graves ◽  
S. Yang
Keyword(s):  
Growth Factor ◽  
Bone Formation ◽  
Primary Cilium ◽  
Primary Cilia ◽  
Transforming Growth Factor ◽  
Tooth Development ◽  
Critical Role ◽  
Intraflagellar Transport ◽  
Growth Factor Signaling ◽  
Cilia Formation

The primary cilium is a nonmotile microtubule-based organelle in most vertebrate cell types. The primary cilium plays a critical role in tissue development and homeostasis by sensing and transducing various signaling pathways. Ciliary proteins such as intraflagellar transport (IFT) proteins as well as ciliary motor proteins, kinesin and dynein, comprise a bidirectional intraflagellar transport system needed for cilia formation and function. Mutations in ciliary proteins that lead to loss or dysfunction of primary cilia cause ciliopathies such as Jeune syndrome and Ellis–van Creveld syndrome and cause abnormalities in tooth development. These diseases exhibit severe skeletal and craniofacial dysplasia, highlighting the significance of primary cilia in skeletal development. Cilia are necessary for the propagation of hedgehog, transforming growth factor β, platelet-derived growth factor, and fibroblast growth factor signaling during osteogenesis and chondrogenesis. Ablation of ciliary proteins such as IFT80 or IFT20 blocks cilia formation, which inhibits osteoblast differentiation, osteoblast polarity, and alignment and reduces bone formation. Similarly, cilia facilitate chondrocyte differentiation and production of a cartilage matrix. Cilia also play a key role in mechanosensing and are needed for increased bone formation in response to mechanical forces.

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