The TBK1-SCF^FBXO3-TMEM192-TAX1BP1 axis: a novel regulatory mechanism for lysophagy

ABSTRACT

Lysophagy, the selective macroautophagic/autophagic clearance of damaged lysosomes, is a critical mechanism for maintaining cellular homeostasis. Our recent study identified a novel regulatory axis involving TBK1, SCF^FBXO3, TMEM192, and TAX1BP1 that orchestrates lysophagic flux following lysosomal damage. We demonstrated that TBK1-dependent phosphorylation of FBXO3 facilitates its interaction with TMEM192, promoting its ubiquitination and subsequent recognition by the autophagy receptor TAX1BP1. Perturbing this pathway significantly reduces lysophagic flux and results in accumulation of damaged lysosomes. These findings establish a previously unrecognized mechanistic link between ubiquitination, receptor recruitment, and lysophagic degradation, broadening our understanding of lysosomal quality control.

Lysosomes house the primary catabolic activities of the cell, breaking down macromolecules and recycling their components. Maintaining lysosomal homeostasis is therefore essential to prevent detrimental effects of protease leakage and the buildup of damaged organelles. While several selective autophagy pathways – such as mitophagy – are increasingly well characterized, the specific regulatory events that govern lysophagy have remained elusive. Our recent findings shed light on this question by identifying a coordinated mechanism that senses lysosomal damage, tags compromised lysosomes with ubiquitin and recruits downstream effectors to ensure timely degradation [1].

Using a targeted ubiquitination-focused compound library screen, we discovered that the SCF^FBXO3 E3 ubiquitin ligase complex, is an essential regulator of lysophagy. Inhibition of FBXO3 markedly reduces lysophagic flux, as evidenced by impaired clearance of LGALS3 (galectin 3) puncta and accumulation of damaged lysosomes by LLOMe. Mechanistically, FBXO3 selectively interacts with the lysosomal membrane protein TMEM192, leading to its ubiquitination. This modification serves as a crucial signal for the recruitment of TAX1BP1, an autophagic receptor protein that facilitates the subsequent sequestration of damaged lysosomes into phagophores and ultimately autophagosomes. Notably, TBK1-mediated phosphorylation of FBXO3 enhances its function, linking stress signaling to lysophagic machinery activation. Consistently, loss of FBXO3 or TMEM192 significantly impairs the recruitment of TAX1BP1 to damaged lysosomes, leading to inefficient lysosomal clearance. Furthermore, we observed that pharmacological inhibition of TBK1 reduces the ability of FBXO3 to ubiquitinate TMEM192, emphasizing the critical role of phosphorylation in orchestrating lysophagy (Figure 1). Notably, our data indicates a process similar to mitophagy, where PINK1-PRKN/Parkin coordinates the ubiquitination of mitochondrial outer membrane proteins to recruit autophagy receptors. By analogy, TBK1-FBXO3 serves as the damage sensor-ubiquitin ligase module for lysosomes, while TMEM192 emerges as a membrane substrate for ubiquitination. Thus, understanding the interplay among these factors will be critical to obtaining a more complete pathway of lysophagic regulation.

Figure 1.

Proposed model of ubiquitin-dependent selective autophagy for lysosomes. Mitochondrial damage triggers mitophagy, regulated by the PINK1-PRKN-VDAC1-SQSTM1 axis. Upon mitochondrial stress, PINK1 accumulates on the mitochondrial membrane and phosphorylates PRKN, facilitating the ubiquitination of membrane proteins such as VDAC1, which, in turn, recruit autophagy receptors. Similarly, lysophagy is initiated in response to lysosomal damage induced by lysosomotropic agents such as LLOMe. Upon lysosomal rupture, TBK1 is activated through phosphorylation at S172 and subsequently phosphorylates FBXO3. FBXO3 then facilitates the ubiquitination of the lysosomal membrane protein TMEM192, serving as a signal for the recruitment of autophagy receptors, including SQSTM1 and TAX1BP1. This TBK1-FBXO3-TMEM192-TAX1BP1 cascade orchestrates the selective degradation of damaged lysosomes.

Our work prompts several new lines of inquiry. First, it will be important to determine how TBK1 senses lysosomal damage signals, especially under conditions of partial injury versus catastrophic rupture. Second, the exact nature of the membrane-based cues that attract TBK1 to damaged lysosomes remains to be clarified; one possibility involves local changes in lipid composition or exposure of glycosylated proteins. Finally, while FBXO3 modulation influences lysophagic efficiency, further studies are needed to assess its specificity and potential side effects.

In conclusion, our discovery of the TBK1-FBXO3-TMEM192-TAX1BP1 axis reveals a major regulatory component of lysophagy. We demonstrate that lysophagy is a highly orchestrated process rather than a mere byproduct of generalized autophagy. These insights highlight the power of selective autophagy in safeguarding cellular integrity and underscore its potential for therapeutic intervention.

Funding Statement

This research was supported by the National Research Foundation of Korea, funded by the Ministry of Science & ICT [RS-2024-00338475], and by Basic Science Research Program through the National Research Foundation of Korea (NRF), funded by the Ministry of Education [RS-2024-00453488 and RS-2024-00463344].

Disclosure statement

No potential conflict of interest was reported by the author(s).