RNF170 Protein, an Endoplasmic Reticulum Membrane Ubiquitin Ligase, Mediates Inositol 1,4,5-Trisphosphate Receptor Ubiquitination and Degradation

Abstract

Inositol 1,4,5-trisphosphate (IP 3 ) receptors are endoplasmic reticulum membrane calcium channels that, upon activation, are degraded via the ubiquitin-proteasome pathway. While searching for novel mediators of IP 3 receptor processing, we discovered that RNF170, an uncharacterized RING domaincontaining protein, associates rapidly with activated IP 3 receptors. RNF170 is predicted to have three membrane-spanning helices, is localized to the ER membrane, and possesses ubiquitin ligase activity. Depletion of endogenous RNF170 by RNA interference inhibited stimulus-induced IP 3 receptor ubiquitination, and degradation and overexpression of a catalytically inactive RNF170 mutant suppressed stimulus-induced IP 3 receptor processing. A substantial proportion of RNF170 is constitutively associated with the erlin1/2 (SPFH1/2) complex, which has been shown previously to bind to IP 3 receptors immediately after their activation. Depletion of RNF170 did not affect the binding of the erlin1/2 complex to stimulated IP 3 receptors, whereas erlin1/2 complex depletion inhibited RNF170 binding. These results suggest a model in which the erlin1/2 complex recruits RNF170 to activated IP 3 receptors where it mediates IP 3 receptor ubiquitination. Thus, RNF170 plays an essential role in IP 3 receptor processing via the ubiquitin-proteasome pathway.Inositol 1,4,5-trisphosphate (IP 3 ) 3 receptors form tetrameric, IP 3 -and Ca 2ϩ -gated Ca 2ϩ channels in endoplasmic reticulum (ER) membranes of mammalian cells and play a key role in cell signaling (1). Stimulation of certain cell surface receptors triggers IP 3 formation at the plasma membrane, which then diffuses through the cytosol and binds to IP 3 receptors (1). This, in concert with Ca 2ϩ binding, induces yet to be defined conformational changes in the tetrameric channel that permit Ca 2ϩ to flow from stores within the ER lumen into the cytosol (1). There are three IP 3 receptor types in mammals, IP 3 R1, IP 3 R2, and IP 3 R3, and although they differ considerably in their tissue distribution, they have broadly similar properties and are often coexpressed (1).In recent years it has become clear that G-protein-coupled receptor-induced activation of endogenous IP 3 receptors can lead to their rapid degradation via the ubiquitin-proteasome pathway (UPP) (2). This IP 3 receptor "down-regulation" has been demonstrated in many mammalian cell types, including gonadotropin-releasing hormone (GnRH)-stimulated ␣T3-1 mouse pituitary gonadotropes (3), endothelin 1 (ET1)-stimulated Rat1 fibroblasts (4), and muscarinic agonist-stimulated SH-SY5Y human neuroblastoma cells (5) and mHeLa cells (6).The generally accepted summary of the UPP is that substrates are first polyubiquitinated and then processed by the proteasome, a multi-subunit protease that can recognize and degrade polyubiquitinated proteins (7,8). Ubiquitination, the key step in targeting a protein for proteasomal degradation, is achieved through the hierarchical action of three enzymes, termed ubiquitin-activating enzyme (E1), ubiqu...

The Stability and Expression Level of Bok Are Governed by Binding to Inositol 1,4,5-Trisphosphate Receptors

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Tumor suppressive Ca2+ signaling is driven by IP3 receptor fitness

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ERAD‐dependent control of the Wnt secretory factor Evi

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