Mouse Receptor Interacting Protein 3 Does Not Contain a Caspase-Recruiting or a Death Domain but Induces Apoptosis and Activates NF-κB

Abstract

The death domain-containing receptor superfamily and their respective downstream mediators control whether or not cells initiate apoptosis or activate NF-kappaB, events critical for proper immune system function. A screen for upstream activators of NF-kappaB identified a novel serine-threonine kinase capable of activating NF-kappaB and inducing apoptosis. Based upon domain organization and sequence similarity, this novel kinase, named mRIP3 (mouse receptor interacting protein 3), appears to be a new RIP family member. RIP, RIP2, and mRIP3 contain an N-terminal kinase domain that share 30 to 40% homology. In contrast to the C-terminal death domain found in RIP or the C-terminal caspase-recruiting domain found in RIP2, the C-terminal tail of mRIP3 contains neither motif and is unique. Despite this feature, overexpression of the mRIP3 C terminus is sufficient to induce apoptosis, suggesting that mRIP3 uses a novel mechanism to induce death. mRIP3 also induced NF-kappaB activity which was inhibited by overexpression of either dominant-negative NIK or dominant-negative TRAF2. In vitro kinase assays demonstrate that mRIP3 is catalytically active and has autophosphorylation site(s) in the C-terminal domain, but the mRIP3 catalytic activity is not required for mRIP3 induced apoptosis and NF-kappaB activation. Unlike RIP and RIP2, mRIP3 mRNA is expressed in a subset of adult tissues and is thus likely to be a tissue-specific regulator of apoptosis and NF-kappaB activity. While the lack of a dominant-negative mutant precludes linking mRIP3 to a known upstream regulator, characterizing the expression pattern and the in vitro functions of mRIP3 provides insight into the mechanism(s) by which cells modulate the balance between survival and death in a cell-type-specific manner.

Receptor Interacting Protein 3 Suppresses Vascular Smooth Muscle Cell Growth by Inhibition of the Phosphoinositide 3-Kinase-Akt Axis

“…5A) and cell death ELISA (Fig. 5B), consistent with previous studies on other cell types (33)(34)(35)(36). Next, we envisioned that normalizing Akt activation by adenoviral gene transfer of the constitutively active PI3K mutant might protect cells from rRIP3-induced cell death.…”

“…Previous studies have shown that the kinase activity of RIP3 contributes to its necrotic effect (40), whereas other studies suggest that the kinase activity is not necessary for its apoptotic effect (34). To determine whether the kinase activity is involved in rRIP3-induced growth arrest and apoptosis, we made a kinaseinactive mutant of RIP3 (rRIP3-C, which lacks the kinase domain).…”

“…Previous studies have demonstrated that RIP1 is obligated to tumor necrosis factor receptor-1-mediated NF-B activation and induces apoptosis and necrosis when overexpressed (30,31), whereas RIP1 deficiency prevents tumor necrosis factor-mediated activation of NF-B and enhances cell death (32). A large body of evidence suggests that RIP3 is also essentially involved in the tumor necrosis factor receptor-1 signaling pathway (33)(34)(35)(36) and that RIP3 binds to RIP1 and then exerts a potent apoptotic effect by interacting with some procaspases or by attenuating RIP1-and tumor necrosis factor receptor-1-mediated NF-B activation through phosphorylation of RIP1 (37). Moreover, in response to death receptor ligands, RIP3 can mediate both caspase-dependent and -independent apoptosis as well as NF-B activation (38).…”

RIP3 overexpression sensitizes human breast cancer cells to parthenolide in vitro via intracellular ROS accumulation

“…This experiment also proved that RIP3 sensitized MCF-7 and MDA-MB-231 cells to PTL-induced apoptosis. Contrary to the report that RIP3 overexpression could induce apoptosis [19][20][21] , we did not observe any spontaneous apoptosis in our RIP3-MCF-7 or RIP3-MDA-MB-231 cells. This may be because previous work studied apoptosis as soon as 24 h after transient transfection of the RIP3 constructs, while we tested the cells after one-week antibiotic selection.…”

RIPK1- and RIPK3-induced cell death mode is determined by target availability

“…6,7 RIPK3, like RIPK1, bears a kinase domain and RIP homology interaction motif (RHIM), but unlike RIPK1 does not have a DD. [8][9][10][11] RIPK3 is required for necroptosis. 12,13 Furthermore, RIPK1 appears to activate RIPK3 in this pathway, as cell death could be blocked by nec-1.…”