Abstract
Prolonged stimulation of the beta2-adrenergic receptor (beta2AR) leads to receptor ubiquitination and downregulation. Using a genome-wide RNA interference screen, we identified arrestin domain-containing 3 (ARRDC3) as a gene required for beta2AR regulation. The ARRDC3 protein interacts with ubiquitin ligase neural precursor development downregulated protein 4 (NEDD4) through two conserved PPXY motifs and recruits NEDD4 to the activated receptor. The ARRDC3 protein also interacts and co-localizes with activated beta2AR. Knockdown of ARRDC3 expression abolishes the association between NEDD4 and beta2AR. Furthermore, functional inactivation of ARRDC3, either through small interfering RNA (siRNA)-mediated knockdown or overexpression of a mutant that does not interact with NEDD4, blocks receptor ubiquitination and degradation. Our results establish ARRDC3 as an essential adaptor for beta2AR ubiquitination.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Sequence
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Animals
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Arrestins / genetics
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Arrestins / metabolism*
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Cell Line
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Endosomal Sorting Complexes Required for Transport / genetics
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Endosomal Sorting Complexes Required for Transport / metabolism*
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Gene Knockdown Techniques
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Humans
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Molecular Sequence Data
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Nedd4 Ubiquitin Protein Ligases
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RNA Interference
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RNA, Small Interfering / genetics
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RNA, Small Interfering / metabolism
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Receptors, Adrenergic, beta-2 / genetics
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Receptors, Adrenergic, beta-2 / metabolism*
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
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Sequence Alignment
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Ubiquitin-Protein Ligases / genetics
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Ubiquitin-Protein Ligases / metabolism*
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Ubiquitination
Substances
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ARRDC3 protein, human
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Arrestins
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Endosomal Sorting Complexes Required for Transport
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RNA, Small Interfering
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Receptors, Adrenergic, beta-2
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Recombinant Fusion Proteins
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Nedd4 Ubiquitin Protein Ligases
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Nedd4 protein, human
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Ubiquitin-Protein Ligases