| First Author | Liu Y | Year | 2023 |
| Journal | Mol Cell Proteomics | Volume | 22 |
| Issue | 2 | Pages | 100495 |
| PubMed ID | 36634736 | Mgi Jnum | J:333847 |
| Mgi Id | MGI:7442337 | Doi | 10.1016/j.mcpro.2023.100495 |
| Citation | Liu Y, et al. (2023) In-Cell Chemical Crosslinking Identifies Hotspots for SQSTM-1/p62-IkappaBalpha Interaction That Underscore a Critical Role of p62 in Limiting NF-kappaB Activation Through IkappaBalpha Stabilization. Mol Cell Proteomics 22(2):100495 |
| abstractText | We have previously documented that in liver cells, the multifunctional protein scaffold p62/SQSTM1 is closely associated with IkappaBalpha, an inhibitor of the transcriptional activator NF-kappaB. Such an intimate p62-IkappaBalpha association we now document leads to a marked 18-fold proteolytic IkappaBalpha-stabilization, enabling its nuclear entry and termination of the NF-kappaB-activation cycle. In p62(-/-)-cells, such termination is abrogated resulting in the nuclear persistence and prolonged activation of NF-kappaB following inflammatory stimuli. Utilizing various approaches both classic (structural deletion, site-directed mutagenesis) as well as novel (in-cell chemical crosslinking), coupled with proteomic analyses, we have defined the precise structural hotspots of p62-IkappaBalpha association. Accordingly, we have identified such IkappaBalpha hotspots to reside around N-terminal (K38, K47, and K67) and C-terminal (K238/C239) residues in its fifth ankyrin repeat domain. These sites interact with two hotspots in p62: One in its PB-1 subdomain around K13, and the other comprised of a positively charged patch (R(183)/R(186)/K(187)/K(189)) between its ZZ- and TB-subdomains. APEX proximity analyses upon IkappaBalpha-cotransfection of cells with and without p62 have enabled the characterization of the p62 influence on IkappaBalpha-protein-protein interactions. Interestingly, consistent with p62's capacity to proteolytically stabilize IkappaBalpha, its presence greatly impaired IkappaBalpha's interactions with various 20S/26S proteasomal subunits. Furthermore, consistent with p62 interaction with IkappaBalpha on an interface opposite to that of its NF-kappaB-interacting interface, p62 failed to significantly affect IkappaBalpha-NF-kappaB interactions. These collective findings together with the known dynamic p62 nucleocytoplasmic shuttling leads us to speculate that it may be involved in "piggy-back" nuclear transport of IkappaBalpha following its NF-kappaB-elicited transcriptional activation and de novo synthesis, required for termination of the NF-kappaB-activation cycle. Consequently, mice carrying a liver-specific deletion of p62-residues 68 to 252 reveal age-dependent-enhanced liver inflammation. Our findings reveal yet another mode of p62-mediated pathophysiologically relevant regulation of NF-kappaB. |
