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.
Keywords:
ADP-ribosylation factor; Co-immunoprecipitation; GGA3; GST-pulldown; β-Arrestin interacting partner.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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ADP-Ribosylation Factor 6
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ADP-Ribosylation Factors / metabolism*
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Actin Cytoskeleton / metabolism*
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DNA, Complementary / metabolism
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Enzyme Activation
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GTP Phosphohydrolases / metabolism
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Guanosine Triphosphate / metabolism
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HEK293 Cells
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Humans
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Molecular Biology / methods*
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Muscle, Smooth, Vascular / cytology
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Myocytes, Smooth Muscle / metabolism
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Protein Transport
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RNA, Small Interfering / metabolism
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Recombinant Fusion Proteins / metabolism
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beta-Arrestins / metabolism*
Substances
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ADP-Ribosylation Factor 6
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DNA, Complementary
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RNA, Small Interfering
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Recombinant Fusion Proteins
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beta-Arrestins
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Guanosine Triphosphate
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GTP Phosphohydrolases
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ADP-Ribosylation Factors
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ARF6 protein, human