Antagonistic regulation of trafficking to
            <i>Caenorhabditis elegans</i>
            sensory cilia by a
            <i>Retinal Degeneration 3</i>
            homolog and retromer

Martínez-Velázquez, Luis A.; Ringstad, Niels

doi:10.1073/pnas.1712302115

Cited by 10 publications

(5 citation statements)

References 75 publications

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“…In addition, GCY-22::GFP was more diffuse in the cell body and dendrite (Supplementary Fig. 1,3), in agreement with RDL-1 regulating an early trafficking pathway for GCYs 43 .…”

Section: Gfpsupporting

confidence: 75%

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Ciliary Tip Signaling Compartment Is Formed and Maintained by Intraflagellar Transport

et al. 2020

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Primary 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. signaling domains, remains limited, however.Cilia use two mechanisms, a trafficking system and a diffusion barrier, that function together to regulate the trafficking of proteins into, within, and out of cilia. The main ciliary trafficking machinery, intraflagellar transport (IFT), facilitates bidirectional transport of cargo, including signaling proteins, from the base/foundation (basal body) to the tip of the axoneme 2 . Anterograde IFT to the tip relies on kinesins, and cytoplasmic dynein enables retrograde transport back 3,4 . Two IFT modules, subcomplexes-A and -B 5,6 , together with another module containing BBS proteins (BBSome) that is thought to bridge the subcomplexes, play essential roles in cargo transport 7,8 . The best-known IFT cargos are axoneme structure components, including tubulin 9,10 , but signaling proteins, like the TRPV channel subunits OSM-9 and OCR-2 in the nematode C. elegans, are also transported 11 . Additionally, several mammalian ciliary signaling proteins, namely the GPCR SSTR3 and Hedgehog signaling component SMO, traverse the cilium by both IFT and diffusion 12 .To help confine proteins to cilia, a subdomain immediately distal to the basal body, called the transition zone (TZ), acts as a diffusion barrier for both membrane and soluble proteins [13][14][15] . How the TZ acts with IFT or other trafficking systems to regulate the composition of the sensory organelle is not well understood 16,17 .

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“…In addition, GCY-22::GFP was more diffuse in the cell body and dendrite (Supplementary Fig. 1,3), in agreement with RDL-1 regulating an early trafficking pathway for GCYs 43 .…”

Section: Gfpsupporting

confidence: 75%

“…The C. elegans RD3 orthologue, RDL-1, influences the trafficking of GCYs to the PCMC and cilium 43 .…”

Section: Gfpmentioning

confidence: 99%

Ciliary Tip Signaling Compartment Is Formed and Maintained by Intraflagellar Transport

et al. 2020

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“…Low brood numbers, such as we observed in our vps-26 knockout worms, can also be correlated with defects in ciliogenesis, albeit in a 3D culture environment (Lee et al, 2016). Moreover, a recent study indicates that ciliogenesis in C. elegans BAG sensory neurons is coordinated by retromer-mediated vesicular trafficking (Martinez-Velazquez and Ringstad, 2018). Two recent studies in mammalian hint at the potential involvement of the retromer complex in ciliogenesis.…”

Section: Discussionmentioning

confidence: 99%

“…Whereas the role of the retromer complex has been studied extensively in yeast and in mammalian systems, the function of retromer and its role in development in invertebrates such as C. elegans is incompletely understood. It has been observed that worms with mutant vps-29 or vps-35 genes display impaired trafficking of the receptor-type guanylate cyclase GCY-9, which accumulates in BAG chemosensory neurons (Coudreuse et al, 2006; Martinez-Velazquez and Ringstad, 2018). Recent studies have shown a role for worm retromer subunits in anteroposterior polarity, Wnt signaling and Q cell migration (Prasad and Clark, 2006).…”

Section: Introductionmentioning

confidence: 99%

The retromer complex regulates C. elegans development and mammalian ciliogenesis

Xie

Smith

Dierlam

et al. 2021

Preprint

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The mammalian retromer is comprised of subunits VPS26, VPS29 and VPS35, and a more loosely-associated sorting nexin (SNX) heterodimer. Despite known roles for the retromer in multiple trafficking events in yeast and mammalian cells, its role in development is poorly understood, and its potential function in primary ciliogenesis remains unknown. Using CRISPR-Cas9 editing, we demonstrated that vps-26 homozygous knockout C. elegans have reduced brood sizes and impaired vulval development, as well as decreased body length which has been linked to defects in primary ciliogenesis. Since many endocytic proteins are implicated in the generation of primary cilia, we addressed whether the retromer regulates ciliogenesis in mammalian cells. We observed VPS35 localized to the primary cilium, and depletion of VPS26, VPS35 or SNX1/SNX5 led to decreased ciliogenesis. Retromer also coimmunoprecipitated with the capping protein, CP110, and was required for its removal from the mother centriole. Herein, we characterize new roles for the retromer in C. elegans development and in the regulation of ciliogenesis in mammalian cells, and suggest a novel role for the retromer in CP110 removal from the mother centriole.

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“…Therefore, it is rather tempting to speculate that some of the conserved N- and/or C-proximal portions of the RD3 molecule are specifically required for the cyclase trafficking, such as by playing the role of a recognition signal or participating in the interactions with the trafficking machinery, after the high-affinity binding of RD3 to the cyclase has occurred via the interface identified in our present study. Experimental evaluation of such a possibility may also help shed light on additional biological processes potentially involving RD3, because some recent reports suggest that RD3 (or its homologs) can also interact with other trafficked or nontrafficked proteins ( 45 , 46 ), and not only in the retina but also perhaps in different organs of vertebrate and invertebrate species.…”

Section: Discussionmentioning

confidence: 99%

Two clusters of surface-exposed amino acid residues enable high-affinity binding of retinal degeneration-3 (RD3) protein to retinal guanylyl cyclase

Peshenko

Dizhoor

2020

Journal of Biological Chemistry

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Retinal degeneration-3 (RD3) protein protects photoreceptors from degeneration by preventing retinal guanylyl cyclase (RetGC) activation via calcium-sensing guanylyl cyclase–activating proteins (GCAP), and RD3 truncation causes severe congenital blindness in humans and other animals. The three-dimensional structure of RD3 has recently been established, but the molecular mechanisms of its inhibitory binding to RetGC remain unclear. Here, we report the results of probing 133 surface-exposed residues in RD3 by single substitutions and deletions to identify side chains that are critical for the inhibitory binding of RD3 to RetGC. We tested the effects of these substitutions and deletions in vitro by reconstituting purified RD3 variants with GCAP1-activated human RetGC1. Whereas the vast majority of the surface-exposed residues tolerated substitutions without loss of RD3’s inhibitory activity, substitutions in two distinct narrow clusters located on the opposite sides of the molecule effectively suppressed RD3 binding to the cyclase. The first surface-exposed cluster included residues adjacent to Leu-63 in the loop connecting helices 1 and 2. The second cluster surrounded Arg-101 on a surface of helix 3. Single substitutions in those two clusters drastically, i.e. up to 245-fold, reduced the IC50 for the cyclase inhibition. Inactivation of the two binding sites completely disabled binding of RD3 to RetGC1 in living HEK293 cells. In contrast, deletion of 49 C-terminal residues did not affect the apparent affinity of RD3 for RetGC. Our findings identify the functional interface on RD3 required for its inhibitory binding to RetGC, a process essential for protecting photoreceptors from degeneration.

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Antagonistic regulation of trafficking to Caenorhabditis elegans sensory cilia by a Retinal Degeneration 3 homolog and retromer

Cited by 10 publications

References 75 publications

Ciliary Tip Signaling Compartment Is Formed and Maintained by Intraflagellar Transport

Ciliary Tip Signaling Compartment Is Formed and Maintained by Intraflagellar Transport

The retromer complex regulates C. elegans development and mammalian ciliogenesis

Two clusters of surface-exposed amino acid residues enable high-affinity binding of retinal degeneration-3 (RD3) protein to retinal guanylyl cyclase

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