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Author Notes:

Address correspondence to: Tamara Caspary (tcaspar@emory.edu).

We thank Kate Cameron for technical assistance with FACS sorting at the Academic Medical Center, Sarah Bay for comments on the manuscript and heroic figure revisions, and Cheryl Timms Strauss for editing.

Subjects:

Research Funding:

This work was supported by National Institutes of Health Grants NS056380, GM110663, and NS090029 (T.C.), KWF Project Grant UVA 2012–5607, and continuous support from the AMC Foundation (M.F.B.).

L.E.M. was supported by National Institutes of Health training grants (GM08605 and EY007092) and an American Heart Association predoctoral fellowship (11PRE7200011).

S.S. received support from National Institutes of Health Grant T32 GM008490.

Essential services were provided by the Emory Custom Cloning Core Facility, the Emory·Children’s Pediatric Research Center Flow Cytometry Core, the Emory University School of Medicine Core Facility for Flow Cytometry, and the Emory Viral Vector Core (National Institute of Neurological Disorders and Stroke Core Facilities Grant P30NS055077).

Keywords:

  • Science & Technology
  • Life Sciences & Biomedicine
  • Cell Biology
  • ADP-RIBOSYLATION FACTOR
  • GTPASE-ACTIVATING PROTEIN
  • JOUBERT-SYNDROME
  • SONIC HEDGEHOG
  • ARF-FAMILY
  • NEURAL-TUBE
  • IN-VIVO
  • INTRAFLAGELLAR TRANSPORT
  • MUTATIONS
  • BINDING

Arl13b regulates Shh signaling from both inside and outside the cilium

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Journal Title:

Molecular Biology of the Cell

Volume:

Volume 27, Number 23

Publisher:

, Pages 3780-3790

Type of Work:

Article | Final Publisher PDF

Abstract:

The regulatory GTPase Arl13b localizes to primary cilia, where it regulates Sonic hedgehog (Shh) signaling. Missense mutations in ARL13B can cause the ciliopathy Joubert syndrome (JS), and the mouse null allele is embryonic lethal. We used mouse embryonic fibroblasts as a system to determine the effects of Arl13b mutations on Shh signaling. We tested seven different mutants-Three JS-causing variants, two point mutants predicted to alter guanine nucleotide handling, one that disrupts cilia localization, and one that prevents palmitoylation and thus membrane binding-in assays of transcriptional and nontranscriptional Shh signaling. We found that mutations disrupting Arl13b's palmitoylation site, cilia localization signal, or GTPase handling altered the Shh response in distinct assays of transcriptional or nontranscriptional signaling. In contrast, JS-causing mutations in Arl13b did not affect Shh signaling in these same assays, suggesting that these mutations result in more subtle defects, likely affecting only a subset of signaling outputs. Finally, we show that restricting Arl13b from cilia interferes with its ability to regulate Shh-stimulated chemotaxis, despite previous evidence that cilia themselves are not required for this nontranscriptional Shh response. This points to a more complex relationship between the ciliary and nonciliary roles of this regulatory GTPase than previously envisioned.

Copyright information:

© 2016 Mariani et al.

This is an Open Access work distributed under the terms of the Creative Commons Attribution-NonCommercial-ShareAlike 3.0 Unported License (http://creativecommons.org/licenses/by-nc-sa/3.0/).

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