Spatiotemporal manipulation of ciliary glutamylation reveals its roles in intraciliary trafficking and Hedgehog signaling

Hong, Shi‐Rong; Wang, Cuei-Ling; Huang, Yao‐Shen; Chang, Yu‐Chen; Chang, Yaw-Jen; Pusapati, Ganesh V.; Lin, Chih Kuang; Hsu, Nan‐Jung; Cheng, Hsiao-Chi; Chiang, Yueh-Chen; Huang, Wei-En; Shaner, Nathan C.; Rohatgi, Rajat; Inoue, Takanari; Lin, Yu‐Chun

doi:10.1101/282103

Cited by 17 publications

(28 citation statements)

References 71 publications

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“…Polyglutamylation decorates the surface of axonemal microtubules. This reversible modification ranges from 1-17 glutamyl residues in vivo (59), and plays a role in IFT activity and MAP binding (60)(61)(62)(63)(64).…”

Section: Post-translational Modifications Of Ciliary Microtubulesmentioning

confidence: 99%

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ARMC9 and TOGARAM1 define a Joubert syndrome-associated protein module that regulates axonemal post-translational modifications and cilium stability

Latour

Weghe

Rusterholz

et al. 2019

Preprint

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Joubert syndrome (JBTS) is a recessive neurodevelopmental ciliopathy, characterized by a pathognomonic hindbrain malformation. All known JBTS-genes encode proteins involved in the structure or function of primary cilia, ubiquitous antenna-like organelles essential for cellular signal transduction. Here, we use the recently identified JBTS-associated protein ARMC9 in tandem-affinity purification and yeast twohybrid screens to identify a novel ciliary module composed of ARMC9-TOGARAM1-CCDC66-CEP104-CSPP1. TOGARAM1-variants cause JBTS and disrupt its interaction with ARMC9. Using a combination of protein interaction analyses and characterization of patient-derived fibroblasts, CRISPR/Cas9-engineered zebrafish and hTERT-RPE1 cells, we demonstrate that dysfunction of ARMC9 or TOGARAM1 results in short cilia with decreased axonemal acetylation and glutamylation, but relatively intact transition zone function. Aberrant serum-induced ciliary resorption and cold-induced depolymerization in both ARMC9 and TOGARAM1 patient cells lines suggest a role for this new JBTS-associated protein complex in ciliary stability.part by the transition zone (TZ) that connects the axoneme to the membrane and acts as a partition.Approximately half of the known JBTS proteins, including RPGRIP1L (JBTS7) (14) and CC2D2A (JBTS9) (15), assemble into multi-protein complexes at the ciliary TZ where they are thought to organize the molecular gate that regulates ciliary protein entry and exit (16); and dysfunction of TZ organization is thought to play a key role in JBTS. Another subset of JBTS-associated proteins, including ARL13B (JBTS8) and INPP5E (JBTS1) (6), associate with the ciliary membrane beyond the TZ. These proteins are thought to play a role in regulation of signaling pathways such as Hh signaling via maintenance of ciliary lipid composition (17, 18). Different JBTS-associated proteins have been found to function at the basal body or distal segment/tip (4). CSPP1 (JBTS 21) (19) and CEP104 (JBTS25) (20) were mainly detected at the centrosomes and ciliary basal bodies, however their exact molecular function, and how defects in these proteins lead to JBTS, are less well understood. CEP104 localizes to the ciliary tip during ciliogenesis, where it is required for structural integrity in the motile cilia of Chlamydomonas and Tetrahymena (21, 22). Mutations in the gene encoding the ciliary tip kinesin KIF7 (JBTS12) cause JBTS, which were linked to defects in tubulin acetylation and Hedgehog signaling (23).Recently, we identified mutations in the gene encoding armadillo repeat motif containing 9 (ARMC9) in individuals with JBTS (JBTS30). ARMC9 localizes to centrioles (27) and the proximal portion of the cilium (28) in mammalian cilia. ARMC9 transcript levels are upregulated with induction of ciliogenesis and armc9 dysfunction in zebrafish yields typical ciliopathy phenotypes (27). As ARMC9 has not been identified as a component of the ciliary JS-associated protein modules mentioned above, we here use ARMC9 as a bait in protein interaction screen...

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Section: Post-translational Modifications Of Ciliary Microtubulesmentioning

confidence: 99%

“…Decreased ciliary polyglutamylation interferes with kinesin2-mediated anterograde transport, also on the B-tubule, and subsequently negatively impacts Hedgehog signaling (61,63). Some MAPs are sensitive to the amount of glutamylation.…”

Section: Post-translational Modifications Of Ciliary Microtubulesmentioning

confidence: 99%

ARMC9 and TOGARAM1 define a Joubert syndrome-associated protein module that regulates axonemal post-translational modifications and cilium stability

Latour

Weghe

Rusterholz

et al. 2019

Preprint

View full text Add to dashboard Cite

show abstract

“…Similar skeletal malformations have been noted in humans and rodent models of FASD (69–71). The decrease in Cep41 was particularly interesting, as polyglutamylation of tubulin is important for protein trafficking within the cilia, including Shh pathway proteins (72). In addition, Evc is known to play a role in Gli protein trafficking (67).…”

Section: Discussionmentioning

confidence: 99%

Prenatal alcohol exposure disrupts Shh pathway and primary cilia genes in the mouse neural tube

Boschen¹,

Fish²,

Parnell³

2019

Preprint

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Neurulation-stage alcohol exposure (NAE; embryonic day [E] 8-10) is associated with midline craniofacial and CNS defects that likely arise from disruption of morphogen pathways, such as Sonic hedgehog (Shh). Notably, midline anomalies are also a hallmark of genetic ciliopathies such as Joubert syndrome. We tested whether NAE alters Shh pathway signaling and the number and function of primary cilia, organelles critical for Shh pathway transduction. Female C57BL/6J mice were administered two doses of alcohol (2.9 g/kg/dose) or vehicle on E9. Embryos were collected 6, 12, or 24 hr later, and changes to Shh, cell cycle genes, and primary cilia were measured in the rostroventral neural tube (RVNT). Within the first 24 hours post-NAE, reductions in Shh pathway and cell cycle gene expression and the ratio of Gli3 forms in the full-length activator state were observed. RVNT volume and cell layer width were reduced at 12 hr. In addition, expression of multiple cilia-related genes were observed at 6 hr post-NAE. As a further test of cilia gene-ethanol interaction, mice heterozygous for Kif3a exhibited perturbed behavior during adolescence following NAE compared to vehicle-treated mice, and Kif3a heterozygosity exacerbated the hyperactive effects of NAE on exploratory activity. These data demonstrate that NAE downregulates the Shh pathway in a region of the neural tube that gives rise to alcohol-sensitive brain structures and identifies disruption of primary cilia function, or a “transient ciliopathy”, as a possible cellular mechanism of prenatal alcohol pathogenesis.

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“…The B-tubule of the axonemal dMT was previously shown as a physical substrate of anterograde intraflagellar transport and post-translational modifications in cilia and flagella (Lechtreck and Geimer, 2000;O'Hagan et al, 2011;Redeker et al, 2005;Stepanek and Pigino, 2016). Through its ultrastructure, tubulin composition and post-translational modifications, the B-tubule of the dMT regulates cell-specific coordination of intraflagellar transport motors and, as such, enabling flagellar motility and mediating specialization of ciliary function (Hong et al, 2018;Silva et al, 2017;Suryavanshi et al, 2010).…”

Section: Tz Maturation and Remodeling Occur Via B-tubule Assembly Andmentioning

confidence: 99%

Cell type-specific structural plasticity of the ciliary transition zone inC. elegans

Akella

Silva

Morsci³

et al. 2018

Preprint

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Running Title: Transition zone structure can change on cilia type. Both cases result in structural TZ intermediates different from TZ in adult cilia. In CEM cilia, axonemal extension and maturation occurs concurrently with TZ structural maturation. Conclusions and SignificanceOur work extends the current model to include the structural plasticity of metazoan transition zone, which can be structurally delayed, maintained or remodeled in cell typespecific manner.

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Spatiotemporal manipulation of ciliary glutamylation reveals its roles in intraciliary trafficking and Hedgehog signaling

Cited by 17 publications

References 71 publications

ARMC9 and TOGARAM1 define a Joubert syndrome-associated protein module that regulates axonemal post-translational modifications and cilium stability

ARMC9 and TOGARAM1 define a Joubert syndrome-associated protein module that regulates axonemal post-translational modifications and cilium stability

Prenatal alcohol exposure disrupts Shh pathway and primary cilia genes in the mouse neural tube

Cell type-specific structural plasticity of the ciliary transition zone inC. elegans

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