The adaptor protein p62/SQSTM1 in osteoclast signaling pathways

Abstract

Paget's disease of bone (PDB) is a skeletal disorder characterized by focal and disorganized increases in bone turnover and overactive osteoclasts. The discovery of mutations in the SQSTM1/p62 gene in numerous patients has identified protein p62 as an important modulator of bone turnover. In both precursors and mature osteoclasts, the interaction between receptor activator of NF-κB ligand (RANKL) and its receptor RANK results in signaling cascades that ultimately activate transcription factors, particularly NF-κB and NFATc1, promoting and regulating the osteoclast differentiation, activity, and survival. As a scaffold with multiple protein-protein interaction motifs, p62 is involved in virtually all the RANKL-activated osteoclast signaling pathways, along with being implicated in numerous other cellular processes. The p62 adaptor protein is one of the functional links reported between RANKL and TRAF6-mediated NF-κB activation, and also plays a major role as a shuttling factor that targets polyubiquitinated proteins for degradation by either the autophagy or proteasome pathways. The dysregulated expression and/or activity of p62 in bone disease up-regulates osteoclast functions. This review aims to outline and summarize the role of p62 in RANKL-induced signaling pathways and in ubiquitin-mediated signaling in osteoclasts, and the impact of PDB-associated p62 mutations on these processes.

Figure 1

Interaction motifs and domains of RANKL signal intermediaries. A: p62 and an aPKC. The cytosolic p62 protein, encoded by the SQSTM1 gene, is a scaffolding protein that interacts with the RANK signaling complex, and is one of the functional links reported between RANKL and TRAF-6-mediated NF-κB activation. Multiple interaction motifs located within p62 enable recruitment of specific proteins and regulation of downstream signaling pathways. RIP binds to the ZZ domain, whereas TRAF6 interacts with the TF6-b sequence, and the aPKC isoforms, ERK, and others interact with the PB1 domain. This PB1 interaction directs the aPKCs in the NF-κB pathway. The UBA domain binds to polyubiquitin chains, and is important for the ubiquitination of TRAF6. PB1, PB1 dimerisation domain; ZZ ZNF, ZZ-type zinc finger; TF6-b, TRAF6 binding sequence; PEST, (P, Proline; E, Glutamate; S, Serine; T, Threonine) rich sequence; UBA, ubiquitin associated; PS, pseudosubstrate region; MATH, meprin and TRAF homology. B: The RANK receptor. RANK has six known putative TRAF-binding motifs (PTM 1-6) of which three have been involved in osteoclast signaling. TRAF6 binds to membrane-proximal motifs such as PTM3 (PFQEP^369-373), whereas PTM5 (PVQEET^559-564) and PTM6 (PVQEQ^604-609) most likely interact with TRAF2 and TRAF5 [22,23]. In addition, three other TRAF6- binding sites have been identified, BS I (PTEDEY^340-345), BS II (PLEVGE^373-378) and BS III (PGEDHE^447-452 [25,26].

Figure 2

p62 in osteoclast signaling and protein trafficking. The binding of RANKL to the receptor protein RANK at the plasma membrane induces the formation of a trimer, triggering the recruitment of a series of adaptor proteins. TRAF6 catalyses Lys63-linked autoubiquitination via intrinsic E3 ubiquitin ligase activity, which is regulated by the UBA domain of p62, and eventually deubiquitinated post-recruitment of CYLD to p62. In the interim, this ubiquitination permits activation of the TAB1-TAB2-TAK1 complex, in turn activating the MAP kinases, as well as NF-κB-inducing kinase (NIK), which leads to phosphorylation and activation of IKKβ. Activation of TRAF6 and p62 also leads to activation of the Akt/PKB pathway. Simultaneously, p62 binds aPKC through its N-terminal PB1, allowing for the phosphorylation of IKKβ by the aPKC. Once activated, IKKβ phosphorylates IκB, which is subsequently ubiquitinated, and degraded through the proteasome system, liberating NF-κB to translocate to the nucleus and interact with transcription promoters.