Intraflagellar Transport Machinery Research Articles

The IFT20 homolog in Caenorhabditis elegans is required for ciliogenesis and cilia-mediated behavior.

Cilia are microtubule-based organelles that carry out a wide range of critical functions throughout the development of higher animals. Regardless of their type, all cilia rely on a motor-driven, bidirectional transport system known as intraflagellar transport (IFT). Of the many components of the IFT machinery, IFT20 is one of the smallest subunits. Nevertheless, IFT20 has been shown to play critical roles in the assembly of several types of mammalian cilia. Here we show that the IFT20 homolog in Caenorhabditis elegans, IFT-20, is also important for correct cilium assembly in sensory neurons. Strikingly, however, we find that IFT-20-deficient animals are able to assemble short, vestigial cilia. In spite of this, we show that practically all IFT-20-deficient animals fail to respond to environmental cues that are normally detected by cilia to modulate their behavior. Altogether, our results indicate that IFT-20 is critical for both the correct assembly and function of cilia in C. elegans.

A missense mutation in IFT74, encoding for an essential component for intraflagellar transport of Tubulin, causes asthenozoospermia and male infertility without clinical signs of Bardet-Biedl syndrome.

Cilia and flagella are formed around an evolutionary conserved microtubule-based axoneme and are required for fluid and mucus clearance, tissue homeostasis, cell differentiation and movement. The formation and maintenance of cilia and flagella require bidirectional transit of proteins along the axonemal microtubules, a process called intraflagellar transport (IFT). In humans, IFT defects contribute to a large group of systemic diseases, called ciliopathies, which often display overlapping phenotypes. By performing exome sequencing of a cohort of 167 non-syndromic infertile men displaying multiple morphological abnormalities of the sperm flagellum (MMAF) we identified two unrelated patients carrying a homozygous missense variant adjacent to a splice donor consensus site of IFT74 (c.256G > A;p.Gly86Ser). IFT74 encodes for a core component of the IFT machinery that is essential for the anterograde transport of tubulin. We demonstrate that this missense variant affects IFT74 mRNA splicing and induces the production of at least two distinct mutant proteins with abnormal subcellular localization along the sperm flagellum. Importantly, while IFT74 deficiency was previously implicated in two cases of Bardet-Biedl syndrome, a pleiotropic ciliopathy with variable expressivity, our data indicate that this missense mutation only results in primary male infertility due to MMAF, with no other clinical features. Taken together, our data indicate that the nature of the mutation adds a level of complexity to the clinical manifestations of ciliary dysfunction, thus contributing to the expanding phenotypical spectrum of ciliopathies.

Molecular basis of ciliary defects caused by compound heterozygous IFT144/WDR19 mutations found in cranioectodermal dysplasia.

Primary cilia contain specific proteins to achieve their functions as cellular antennae. Ciliary protein trafficking is mediated by the intraflagellar transport (IFT) machinery containing the IFT-A and IFT-B complexes. Mutations in genes encoding the IFT-A subunits (IFT43, IFT121/WDR35, IFT122, IFT139/TTC21B, IFT140 and IFT144/WDR19) often result in skeletal ciliopathies, including cranioectodermal dysplasia (CED). We here characterized the molecular and cellular defects of CED caused by compound heterozygous mutations in IFT144 [the missense variant IFT144(L710S) and the nonsense variant IFT144(R1103*)]. These two variants were distinct with regard to their interactions with other IFT-A subunits and with the IFT-B complex. When exogenously expressed in IFT144-knockout (KO) cells, IFT144(L710S) as well as IFT144(WT) rescued both moderately compromised ciliogenesis and the abnormal localization of ciliary proteins. As the homozygous IFT144(L710S) mutation was found to cause autosomal recessive retinitis pigmentosa, IFT144(L710S) is likely to be hypomorphic at the cellular level. In striking contrast, the exogenous expression of IFT144(R1103*) in IFT144-KO cells exacerbated the ciliogenesis defects. The expression of IFT144(R1103*) together with IFT144(WT) restored the abnormal phenotypes of IFT144-KO cells. However, the coexpression of IFT144(R1103*) with the hypomorphic IFT144(L710S) variant in IFT144-KO cells, which mimics the genotype of compound heterozygous CED patients, resulted in severe ciliogenesis defects. Taken together, these observations demonstrate that compound heterozygous mutations in IFT144 cause severe ciliary defects via a complicated mechanism, where one allele can cause severe ciliary defects when combined with a hypomorphic allele.

IFT54 directly interacts with kinesin-II and IFT dynein to regulate anterograde intraflagellar transport.

The intraflagellar transport (IFT) machinery consists of the anterograde motor kinesin-II, the retrograde motor IFT dynein, and the IFT-A and -B complexes. However, the interaction among IFT motors and IFT complexes during IFT remains elusive. Here, we show that the IFT-B protein IFT54 interacts with both kinesin-II and IFT dynein and regulates anterograde IFT. Deletion of residues 342-356 of Chlamydomonas IFT54 resulted in diminished anterograde traffic of IFT and accumulation of IFT motors and complexes in the proximal region of cilia. IFT54 directly interacted with kinesin-II and this interaction was strengthened for the IFT54Δ342-356 mutant in vitro and in vivo. The deletion of residues 261-275 of IFT54 reduced ciliary entry and anterograde traffic of IFT dynein with accumulation of IFT complexes near the ciliary tip. IFT54 directly interacted with IFT dynein subunit D1bLIC, and deletion of residues 261-275 reduced this interaction. The interactions between IFT54 and the IFT motors were also observed in mammalian cells. Our data indicate a central role for IFT54 in binding the IFT motors during anterograde IFT.

Rabl2 GTP hydrolysis licenses BBSome-mediated export to fine-tune ciliary signaling.

Cilia of higher animals sense various environmental stimuli. Proper ciliary signaling requires appropriate extent of BBSome-mediated export of membrane receptors across ciliary barrier transition zone (TZ) through retrograde intraflagellar transport (IFT) machinery. How the barrier passage is controlled, however, remains unknown. Here, we show that small GTPase Rabl2 functions as a molecular switch for the outward TZ passage. Rabl2-GTP enters cilia by binding to IFT-B complex. Its GTP hydrolysis enables the outward TZ passage of the BBSome and its cargos with retrograde IFT machinery, whereas its persistent association leads to their shedding from IFT-B during the passing process and consequently ciliary retention. Rabl2 deficiency or expression of a GTP-locked mutant impairs the ciliary hedgehog signaling without interfering with ciliation and respectively results in different spectrums of mouse developmental disorders. We propose that the switch role of Rabl2 ensures proper turnover of the BBSome and ciliary membrane receptors to fine-tune cilia-dependent signaling for normal embryonic development and organismic homeostasis.

Cooperation of the IFT-A complex with the IFT-B complex is required for ciliary retrograde protein trafficking and GPCR import.

Cilia sense and transduce extracellular signals via specific receptors. The intraflagellar transport (IFT) machinery mediates not only bidirectional protein trafficking within cilia but also the import/export of ciliary proteins across the ciliary gate. The IFT machinery is known to comprise two multisubunit complexes, namely, IFT-A and IFT-B; however, little is known about how the two complexes cooperate to mediate ciliary protein trafficking. We here show that IFT144–IFT122 from IFT-A and IFT88–IFT52 from IFT-B make major contributions to the interface between the two complexes. Exogenous expression of the IFT88(Δα) mutant, which has decreased binding to IFT-A, partially restores the ciliogenesis defect of IFT88-knockout (KO) cells. However, IFT88(Δα)-expressing IFT88-KO cells demonstrate a defect in IFT-A entry into cilia, aberrant accumulation of IFT-B proteins at the bulged ciliary tips, and impaired import of ciliary G protein–coupled receptors (GPCRs). Furthermore, overaccumulated IFT proteins at the bulged tips appeared to be released as extracellular vesicles. These phenotypes of IFT88(Δα)-expressing IFT88-KO cells resembled those of IFT144-KO cells. These observations together indicate that the IFT-A complex cooperates with the IFT-B complex to mediate the ciliary entry of GPCRs as well as retrograde trafficking of the IFT machinery from the ciliary tip.

Non-ciliary Roles of IFT Proteins in Cell Division and Polycystic Kidney Diseases.

Cilia are small organelles present at the surface of most differentiated cells where they act as sensors for mechanical or biochemical stimuli. Cilia assembly and function require the Intraflagellar Transport (IFT) machinery, an intracellular transport system that functions in association with microtubules and motors. If IFT proteins have long been studied for their ciliary roles, recent evidences indicate that their functions are not restricted to the cilium. Indeed, IFT proteins are found outside the ciliary compartment where they are involved in a variety of cellular processes in association with non-ciliary motors. Recent works also provide evidence that non-ciliary roles of IFT proteins could be responsible for the development of ciliopathies related phenotypes including polycystic kidney diseases. In this review, we will discuss the interactions of IFT proteins with microtubules and motors as well as newly identified non-ciliary functions of IFT proteins, focusing on their roles in cell division. We will also discuss the potential contribution of non-ciliary IFT proteins functions to the etiology of kidney diseases.

Anterograde trafficking of ciliary MAP kinase–like ICK/CILK1 by the intraflagellar transport machinery is required for intraciliary retrograde protein trafficking

ICK (also known as CILK1) is a mitogen-activated protein kinase-like kinase localized at the ciliary tip. Its deficiency is known to result in the elongation of cilia and causes ciliopathies in humans. However, little is known about how ICK is transported to the ciliary tip. We here show that the C-terminal noncatalytic region of ICK interacts with the intraflagellar transport (IFT)-B complex of the IFT machinery and participates in its transport to the ciliary tip. Furthermore, total internal reflection fluorescence microscopy demonstrated that ICK undergoes bidirectional movement within cilia, similarly to IFT particles. Analysis of ICK knockout cells demonstrated that ICK deficiency severely impairs the retrograde trafficking of IFT particles and ciliary G protein-coupled receptors. In addition, we found that in ICK knockout cells, ciliary proteins are accumulated at the bulged ciliary tip, which appeared to be torn off and released into the environment as an extracellular vesicle. The exogenous expression of various ICK constructs in ICK knockout cells indicated that the IFT-dependent transport of ICK, as well as its kinase activity and phosphorylation at the canonical TDY motif, is essential for ICK function. Thus, we unequivocally show that ICK transported to the ciliary tip is required for retrograde ciliary protein trafficking and consequently for normal ciliary function.

Purification and crystal structure of human ODA16: Implications for ciliary import of outer dynein arms by the intraflagellar transport machinery

Motile cilia protrude from cell surfaces and are necessary to create movement of cells and fluids in the body. At the molecular level, cilia contain several dynein molecular motor complexes including outer dynein arms (ODAs) that are attached periodically to the ciliary axoneme, where they hydrolyse ATP to create the force required for bending and motility of the cilium. ODAs are preassembled in the cytoplasm and subsequently trafficked into the cilium by the intraflagellar transport (IFT) system. In the case of the green alga Chlamydomonas reinhardtii, the adaptor protein ODA16 binds to ODAs and directly to the IFT complex component IFT46 to facilitate the ciliary import of ODAs. Here, we purified recombinant human IFT46 and ODA16, determined the high-resolution crystal structure of the ODA16 protein, and carried out direct interaction studies of IFT46 and ODA16. The human ODA16 C-terminal 320 residues adopt the fold of an eight-bladed β-propeller with high overall structural similarity to the Chlamydomonas ODA16. However, the small 80 residue N-terminal domain, which in Chlamydomonas ODA16 is located on top of the β-propeller and is required to form the binding cleft for IFT46, has no visible electron density in case of the human ODA16 structure. Furthermore, size exclusion chromatography and pull-down experiments failed to detect a direct interaction between human ODA16 and IFT46. These data suggest that additional factors may be required for the ciliary import of ODAs in human cells with motile cilia.

Cardiovascular Regulation by the Neuronal BBSome.

The BBSome, a complex of 8 BBS (Bardet-Biedl syndrome) proteins known for its role in the control of cilia function and other cellular processes, has been implicated in blood pressure control, but the underlying mechanisms are not fully understood. Here, we show that neuronal BBSome plays an important role in blood pressure regulation. Targeted inactivation of the BBSome in the nervous system through Bbs1 gene deletion causes sympathetically mediated increase in blood pressure in mice. This phenotype is reproduced by selective ablation of the Bbs1 gene from the LRb (leptin receptor)-expressing neurons. Strikingly, the well-known role of the BBSome in the regulation of cilia formation and function is unlikely to account for the prohypertensive effect of BBSome inactivation as disruption of the IFT (intraflagellar transport) machinery required for ciliogenesis by deleting the Ift88 gene in LRb neurons had no effect on arterial pressure and sympathetic nerve activity. Furthermore, we found that Bbs1 gene deletion from AgRP (agouti-related protein) neurons or POMC (proopiomelanocortin) neurons increased renal and splanchnic sympathetic nerve activity without altering blood pressure. This lack of blood pressure increase despite the sympathetic overdrive may be explained by vascular adrenergic desensitization as indicated by the reduced vascular contractile response evoked by phenylephrine and the decreased expression of adrenergic receptors. Our results identify the neuronal BBSome as a new player in hemodynamic, sympathetic, and vascular regulation, in a manner independent of cilia.

Single-Molecule Tracking Reveals Complex Motility of Transmembrane Proteins in the Chemosensory Cilia of C. elegans

Cilia are vital for the cell's ability to sense its environment and rely on a process called intra flagellar transport (IFT) for their development, maintenance and function in signal-transduction. In IFT, motor proteins transport ciliary components along the polarized microtubule axoneme of the cilium. Among the cargoes of IFT are transmembrane proteins involved in signal transduction. As a model system, we study C. elegans chemosensory cilia, which we study using fluorescence microscopy with single-molecule sensitivity. First, we demonstrate that IFT machinery and ciliary components, including TRPV transmembrane channel protein OCR-2 are redistributed away from the ciliary tip upon external chemical stimulation, in a robust, extensive and reversible way. To elucidate the dynamics underlying this dramatic protein redistribution, we performed single-molecule imaging of OCR-2 in live C. elegans. Advanced analysis of the single-molecule trajectories shows that, in dendrite and transition zone, active transport is the prevailing motility mode of OCR-2. In the proximal and distal segments, however, motility is a much more complex, location-specific interplay between active transport, normal diffusion and sub diffusion. At the tip, confinement of the membrane proteins plays an important role. Together, our data and analysis demonstrate an intricate interplay between modes of transportation that ensure the proper ciliary distribution of OCR-2. These insights in the dynamics of cellular signal-transduction contributes to a wider understanding of IFT dynamics and to cilia as chemosensory organelles.

Structure of the human BBSome core complex.

The BBSome is a heterooctameric protein complex that plays a central role in primary cilia homeostasis. Its malfunction causes the severe ciliopathy Bardet-Biedl syndrome (BBS). The complex acts as a cargo adapter that recognizes signaling proteins such as GPCRs and links them to the intraflagellar transport machinery. The underlying mechanism is poorly understood. Here we present a high-resolution cryo-EM structure of a human heterohexameric core subcomplex of the BBSome. The structure reveals the architecture of the complex in atomic detail. It explains how the subunits interact with each other and how disease-causing mutations hamper this interaction. The complex adopts a conformation that is open for binding to membrane-associated GTPase Arl6 and a large positively charged patch likely strengthens the interaction with the membrane. A prominent negatively charged cleft at the center of the complex is likely involved in binding of positively charged signaling sequences of cargo proteins.

Trypanosomes have divergent kinesin-2 proteins that function differentially in flagellum biosynthesis and cell viability.

Trypanosoma brucei, the causative agent of African sleeping sickness, has a flagellum that is crucial for motility, pathogenicity, and viability. In most eukaryotes, the intraflagellar transport (IFT) machinery drives flagellum biogenesis, and anterograde IFT requires kinesin-2 motor proteins. In this study, we investigated the function of the two T. brucei kinesin-2 proteins, TbKin2a and TbKin2b, in bloodstream form trypanosomes. We found that, compared to kinesin-2 proteins across other phyla, TbKin2a and TbKin2b show greater variation in neck, stalk and tail domain sequences. Both kinesins contributed additively to flagellar lengthening. Silencing TbKin2a inhibited cell proliferation, cytokinesis and motility, whereas silencing TbKin2b did not. TbKin2a was localized on the flagellum and colocalized with IFT components near the basal body, consistent with it performing a role in IFT. TbKin2a was also detected on the flagellar attachment zone, a specialized structure that connects the flagellum to the cell body. Our results indicate that kinesin-2 proteins in trypanosomes play conserved roles in flagellar biosynthesis and exhibit a specialized localization, emphasizing the evolutionary flexibility of motor protein function in an organism with a large complement of kinesins.

Abstract C07: Ciliogenesis and Hh signaling are suppressed downstream of KRas during ADM

Abstract Acinar-to-ductal metaplasia (ADM) is a homeostatic mechanism where in response to inflammation for example during pancreatitis, acini dedifferentiate, adopting a progenitor-like cell fate before proliferating and redifferentiating to regenerate the pancreas. In pancreatic ductal adenocarcinoma (PDAC) constitutive activation of KRas promotes ADM; however, redifferentiation is blocked and instead progenitor-like cells are locked in an irreversible duct-like state leading to neoplastic transformation and PanIN formation. Hedgehog (Hh) signaling drives regeneration of the pancreas following ADM. Hh signaling is transduced via the primary cilium, a non-motile microtubule-based organelle that projects from the cell surface. Most vertebrate cells have a single cilium; however, acini lack cilia. Upon ADM progenitor-like cells assemble a cilium, consistent with a requirement of Hh signaling for redifferentiation. Cilia are lost in many tumors, including PDAC, where cilia are absent from low-grade PanIN, but the consequence of this is not fully understood. It has recently been reported that cilia loss potentiates KRas signaling in PDAC. Here we have assessed the requirement for cilia during ADM. We show that upon ADM progenitor-like cells expressing activated KRas (from KC mice; LSL-KRasG12D/+;Pdx1Cre) assemble fewer cilia compared to WT cells (from control litter mates). The ciliary axoneme is assembled via intraflagellar transport (IFT). We demonstrate that components of the IFT machinery are present and that the basal body, a structure derived from the mother centriole that nucleates the axoneme, is competent to form cilia in KC cells. Indicating that KC cells are able to assemble cilia, however, we show that ciliogenesis is blocked due in part to ectopic activation of the cilium disassembly pathway. This pathway is mediated via AurkA, known to be upregulated in PDAC. We show that AurkA is also upregulated in KC ADM cells and that inhibition of AurkA restores cilia to WT levels. Cells that lack cilia are unable to respond to Hh signals. We show that decreased cilia in KC ADM cells diminishes their response to Hh signaling. Rescuing cilia through inhibition of AurkA reduces the capacity of KC cells to undergo ADM and restores their response to Hh signals. We conclude that fewer cilia in KC cells caused by upregulation of AurkA prevents a full response to Hh signaling and as such redifferentiation of progenitor-like cells formed during ADM fails to occur. Instead these cells are locked in a metaplastic state predisposed to neoplastic transformation, resulting in the formation of PanIN. This study suggests that restoration of cilia could be used as a mechanism to restore responsiveness to Hh signaling during early disease stages such as chronic pancreatitis, to prevent formation of precursor lesions. Citation Format: Fiona Bangs, Eric O'Neill. Ciliogenesis and Hh signaling are suppressed downstream of KRas during ADM [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Advances in Science and Clinical Care; 2019 Sept 6-9; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2019;79(24 Suppl):Abstract nr C07.

Interactions of the dynein-2 intermediate chain WDR34 with the light chains are required for ciliary retrograde protein trafficking.

The dynein-2 complex drives retrograde ciliary protein trafficking by associating with the intraflagellar transport (IFT) machinery, containing IFT-A and IFT-B complexes. We recently showed that the dynein-2 complex, which comprises 11 subunits, can be divided into three subcomplexes: DYNC2H1–DYNC2LI1, WDR34–DYNLL1/DYNLL2–DYNLRB1/DYNLRB2, and WDR60–TCTEX1D2–DYNLT1/DYNLT3. In this study, we demonstrated that the WDR34 intermediate chain interacts with the two light chains, DYNLL1/DYNLL2 and DYNLRB1/DYNLRB2, via its distinct sites. Phenotypic analyses of WDR34-knockout cells exogenously expressing various WDR34 constructs showed that the interactions of the WDR34 intermediate chain with the light chains are crucial for ciliary retrograde protein trafficking. Furthermore, we found that expression of the WDR34 N-terminal construct encompassing the light chain–binding sites but lacking the WD40 repeat domain inhibits ciliary biogenesis and retrograde trafficking in a dominant-negative manner, probably by sequestering WDR60 or the light chains. Taken together with phenotypic differences of several WDR34-knockout cell lines, these results indicate that incorporation of DYNLL1/DYNLL2 and DYNLRB1/DYNLRB2 into the dynein-2 complex via interactions with the WDR34 intermediate chain is crucial for dynein-2 function in retrograde ciliary protein trafficking.

Requirement of IFT-B-BBSome complex interaction in export of GPR161 from cilia.

ABSTRACTThe intraflagellar transport (IFT) machinery, which includes the IFT-A and IFT-B complexes, mediates bidirectional trafficking of ciliary proteins. In addition to these complexes, the BBSome, which is composed of eight subunits that are encoded by the causative genes of Bardet-Biedl syndrome (BBS), has been proposed to connect the IFT machinery to ciliary membrane proteins, such as G protein-coupled receptors, to mediate their export from cilia. However, little is known about the connection between the IFT machinery and the BBSome. Using the visible immunoprecipitation assay, we here identified the interaction between IFT38 from the IFT-B complex and BBS1, BBS2 and BBS9 from the BBSome. Furthermore, by analyzing phenotypes of IFT38-knockout cells exogenously expressing wild-type IFT38 or its mutant lacking the ability to interact with BBS1+BBS2+BBS9, we showed that knockout cells expressing the IFT38 mutant have restored ciliogenesis; however, similar to BBS1-knockout cells, they demonstrated significant accumulation of GPR161 within cilia upon stimulation of Hedgehog signaling. These results indicate that the IFT-B–BBSome interaction is required for the export of GPR161 across the ciliary gate.

Architectures and functions of motor proteins underlying the intraflagellar transport machinery

Primary cilia, which protrude from the surfaces of most human cells, function as cellular antennae that receive extracellular signals. To serve as antennae, primary cilia contain unique proteins, such as G-protein-coupled receptors and ion channels. Defects in the assembly and functions of primary cilia cause hereditary disorders with a wide range of symptoms, including cystic kidney and retinal degeneration. The assembly and maintenance of cilia depend on protein trafficking mediated by the intraflagellar transport (IFT) machinery, which contains three protein complexes (IFT-A, IFT-B, and BBSome) and two motor proteins (kinesin-2 and dynein-2 complex) and is composed of more than 40 subunits in total. We recently revealed the interaction between the kinesin-2 and IFT-B complexes and overall architecture of the dynein-2 complex by taking advantage of the visible immunoprecipitation (VIP) assay. In addition, we clarified the roles of dynein-2 subunits using gene knockout cell lines established using the CRISPR/Cas9 system. This review focuses on recent advances in the architectures and functions of two motor proteins underlying the IFT machinery.

In situ cryo-electron tomography and subtomogram averaging of intraflagellar transport trains.

In situ cryo-electron tomography (cryo-ET) and subtomogram averaging are powerful tools, able to provide 3D structures of biological samples at sub-nanometer resolution, while preserving information about cellular context and higher-order assembly. Best results are typically achieved, when applied to highly repetitive structures, such as viruses. Other typical examples are protein complexes that decorate long stretches along ciliary microtubules at stereotypical and precise repeats, such as axonemal dyneins. For such cases, a plethora of subtomogram averaging protocols exist. In this chapter, we show how we use cryo-ET and subtomogram averaging to study the architecture of the intraflagellar transport (IFT) machinery, a more challenging target that appears only in low copy numbers per tomogram. In the IFT trains, repeating units of IFT adaptor proteins engage two oppositely directed molecular motors to quickly shuttle ciliary building blocks and other proteins to the tip of the cilium and/or back to the base. This dynamic and sporadic nature of IFT trains poses challenges for determining the localization or precise orientation of the particles to be averaged. Solutions to these problems are described in this chapter.

Chlamydomonas Basal Bodies as Flagella Organizing Centers.

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.

Loss-of-function of IFT88 determines metabolic phenotypes in thyroid cancer.

Primary cilia are microtubule-based, dynamic organelles characterized by continuous assembly and disassembly. The intraflagellar transport (IFT) machinery, including IFT88 in cilia, is involved in the maintenance of bidirectional motility along the axonemes, which is required for ciliogenesis and functional competence. Cancer cells are frequently associated with loss of primary cilia and IFT functions. However, there is little information on the role of IFT88 or primary cilia in the metabolic remodeling of cancer cells. Therefore, we investigated the cellular and metabolic effects of the loss-of-function (LOF) mutations of IFT88/primary cilia in thyroid cancer cells. IFT88-deficient 8505C thyroid cancer cells were generated using the CRISPR/Cas9 system, and RNA-sequencing analysis was performed. LOF of the IFT88 gene resulted in a marked defect in ciliogenesis and mitochondrial oxidative function. Gene expression patterns in IFT88-deficient thyroid cancer cells favored glycolysis and lipid biosynthesis. However, LOF of IFT88/primary cilia did not promote thyroid cancer cell proliferation, migration, and invasion. The results suggest that IFT88/primary cilia play a role in metabolic reprogramming in thyroid cancer cells.