Optineurin Negatively Regulates TNFα- Induced NF-κB Activation by Competing with NEMO for Ubiquitinated RIP

Abstract

NF-kappaB essential modulator (NEMO), the regulatory subunit of the IkappaB kinase (IKK) that activates NF-kappaB, is essential for NF-kappaB activation. NEMO was recently found to contain a region that preferentially binds Lys (K)63-linked but not K48-linked polyubiquitin (polyUb) chains, and the ability of NEMO to bind to K63-linked polyUb RIP (receptor-interacting protein) is necessary for efficient tumor necrosis factor alpha (TNFalpha)-induced NF-kappaB activation. Optineurin is a homolog of NEMO, and mutations in the optineurin gene are found in a subset of patients with glaucoma, a neurodegenerative disease involving the loss of retinal ganglion cells. Although optineurin shares considerable homology with NEMO, in resting cells, it is not present in the high-molecular-weight complex containing IKKalpha and IKKbeta, and optineurin cannot substitute for NEMO in lipopolysaccharide (LPS)-induced NF-kappaB activation. On the other hand, the overexpression of optineurin blocks the protective effect of E3-14.7K on cell death caused by the overexpression of TNFalpha receptor 1 (TNFR1). Here we show that optineurin has a K63-linked polyUb-binding region similar to that of NEMO, and like NEMO, it bound K63- but not K48-linked polyUb. Optineurin competitively antagonized NEMO's binding to polyUb RIP, and its overexpression inhibited TNFalpha-induced NF-kappaB activation. This competition occurs at physiologic protein levels because microRNA silencing of optineurin resulted in markedly enhanced TNFalpha-induced NF-kappaB activity. These results reveal a physiologic role for optineurin in dampening TNFalpha signaling, and this role might provide an explanation for its association with glaucoma.

Ubiquitin binding mediates the NF-κB inhibitory potential of ABIN proteins

“…The identified motif is also present in ABIN-1 and ABIN-2. Therefore, we speculated that this motif might be responsible for binding to ubiquitin, not only in NEMO and optineurin as reported recently (Ea et al, 2006;Wu et al, 2006;Zhu et al, 2007), but also in ABIN-1, ABIN-2, ABIN-3, and called it UBAN (UBD in ABIN proteins and NEMO) domain (Figure 1a). To verify this assumption, we used GST-monoUb and GST-tetraUb to pull down ABIN-1, ABIN-2 and ABIN-3 proteins that were expressed in HEK293T cells.…”

Toward an integrative view of Optineurin functions

“…5,8 Zhu and colleagues have proposed that, following TNFα stimulation, Optn would compete with NEMO for binding to polyubiquitinated substrates (such as RIP1, Table 1), therefore downregulating NFκB activation. 9 Accordingly, a point mutation Asp 474 →Asn (D474N) in the UBD of Optn abolished its binding activity to Lys63-linked polyUb and prevents its interaction with RIP1. The UBD of Optn has also been recently demonstrated to interact with the onco-protein Tax of HTLV1 to modulate Tax ubiquitination and NFκB activation ( Fig.…”

TNF-α signaling in glaucomatous neurodegeneration

“…In a recent study, a glaucoma-associated mutant of optineurin has been shown to selectively induce oxidative stress-mediated RGC death, and optineurin has been suggested to be a likely component of the TNF-α signaling pathway leading to RGC death (Chalasani et al, 2007). More recently, microRNA silencing of optineurin has resulted in markedly enhanced TNF-α-induced nuclear factor kappaB (NF-KB) activity, thereby supporting a physiologic role of optineurin in dampening TNF-α signaling in association with glaucoma (Zhu et al, 2007).…”