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Methods to Investigate the β-Arrestin-Mediated Control of ARF6 Activation to Regulate Trafficking and Actin Cytoskeleton Remodeling

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Beta-Arrestins

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1957))

Abstract

ADP-ribosylation factors (ARF) are GTPases that act to control the activation of numerous signaling events and cellular responses. The ARF6 isoform, present at the plasma membrane, can be activated by the angiotensin II type 1 receptor (AT1R), a process dependent upon β-arrestin recruitment to the activated receptor. Here, we describe classical methods used to assess β-arrestin-dependent activation of ARF6 following agonist stimulation of cells. In addition, because ARF6 and β-arrestin can form a complex, we describe the procedures used to detect the interaction of β-arrestin with this GTPase.

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References

  1. Boulay PL, Cotton M, Melancon P, Claing A (2008) ADP-ribosylation factor 1 controls the activation of the phosphatidylinositol 3-kinase pathway to regulate epidermal growth factor-dependent growth and migration of breast cancer cells. J Biol Chem 283(52):36425–36434

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Hashimoto S, Hirose M, Hashimoto A, Morishige M, Yamada A, Hosaka H, Akagi K, Ogawa E, Oneyama C, Agatsuma T, Okada M, Kobayashi H, Wada H, Nakano H, Ikegami T, Nakagawa A, Sabe H (2006) Targeting AMAP1 and cortactin binding bearing an atypical src homology 3/proline interface for prevention of breast cancer invasion and metastasis. Proc Natl Acad Sci U S A 103(18):7036–7041

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Galas MC, Helms JB, Vitale N, Thierse D, Aunis D, Bader MF (1997) Regulated exocytosis in chromaffin cells. A potential role for a secretory granule-associated ARF6 protein. J Biol Chem 272(5):2788–2793

    Article  CAS  PubMed  Google Scholar 

  4. Mitchell R, Robertson DN, Holland PJ, Collins D, Lutz EM, Johnson MS (2003) ADP-ribosylation factor-dependent phospholipase D activation by the M3 muscarinic receptor. J Biol Chem 278(36):33818–33830

    Article  CAS  PubMed  Google Scholar 

  5. Cohen LA, Honda A, Varnai P, Brown FD, Balla T, Donaldson JG (2007) Active Arf6 recruits ARNO/cytohesin GEFs to the PM by binding their PH domains. Mol Biol Cell 18(6):2244–2253

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Poupart ME, Fessart D, Cotton M, Laporte SA, Claing A (2007) ARF6 regulates angiotensin II type 1 receptor endocytosis by controlling the recruitment of AP-2 and clathrin. Cell Signal 19(11):2370–2378

    Article  CAS  PubMed  Google Scholar 

  7. Charles R, Namkung Y, Cotton M, Laporte SA, Claing A (2015) Betaarrestin-mediated angiotensin II signaling controls the activation of ARF6 and endocytosis in migration of vascular smooth muscle cells. J Biol Chem 291(8):3967–3981

    Article  PubMed  PubMed Central  Google Scholar 

  8. Claing A, Chen W, Miller WE, Vitale N, Moss J, Premont RT, Lefkowitz RJ (2001) Beta arrestin-mediated ARF6 activation and beta2-adrenergic receptor endocytosis. J Biol Chem 276(45):42509–42513

    Article  CAS  PubMed  Google Scholar 

  9. Benard V, Bohl BP, Bokoch GM (1999) Characterization of rac and cdc42 activation in chemoattractant-stimulated human neutrophils using a novel assay for active GTPases. J Biol Chem 274(19):13198–13204

    Article  CAS  PubMed  Google Scholar 

  10. Boman AL, Zhang C, Zhu X, Kahn RA (2000) A family of ADP-ribosylation factor effectors that can alter membrane transport through the trans-Golgi. Mol Biol Cell 11(4):1241–1255

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Dell’Angelica EC, Puertollano R, Mullins C, Aguilar RC, Vargas JD, Hartnell LM, Bonifacino JS (2000) GGAs: a family of ADP ribosylation factor-binding proteins related to adaptors and associated with the Golgi complex. J Cell Biol 149(1):81–94

    Article  PubMed  PubMed Central  Google Scholar 

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Acknowledgments

This work was supported by the Canadian Institutes of Health Research (MOP-79470 and MOP-106596 to AC).

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Correspondence to Audrey Claing .

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Charles, R., Bourmoum, M., Campbell, S., Claing, A. (2019). Methods to Investigate the β-Arrestin-Mediated Control of ARF6 Activation to Regulate Trafficking and Actin Cytoskeleton Remodeling. In: Scott, M., Laporte, S. (eds) Beta-Arrestins. Methods in Molecular Biology, vol 1957. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-9158-7_10

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  • DOI: https://doi.org/10.1007/978-1-4939-9158-7_10

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-9157-0

  • Online ISBN: 978-1-4939-9158-7

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