Background
Our study reveals that the lysosome plays a critical role in Parkinson’s disease pathogenesis. We identified a lysosomal integral membrane protein, TMEM55B, which interacts with a complex of RILPL1, a scaffolding protein, bound with LRRK2-mediated phosphorylated Rabs. Based on this interaction, our current hypothesis is that the function of TMEM55B becomes compromised due to its engagement with RILPL1 in the presence of hyperphosphorylated Rabs. The previously reported and sometimes conflicting functions of TMEM55B provided the rationale for our study, aimed at better defining its molecular role.
Methods and Results
We resolved the crystal structure of TMEM55B residues 80-166 bound with RILPL1-peptide. From this structure, we identified a novel motif, termed as TMEM55B-binding motif (TBM), which mediates the interaction of RILPL1 with TMEM55B. Co-immunoprecipitation of wild-type and mutant TMEM55B, followed by mass spectrometry analysis, revealed a set of new TBM-containing proteins, many of which are implicated in autophagy, lysosomal trafficking, or ciliogenesis. Structural comparisons using FoldSEEK and DALI showed significant similarity between TMEM55B and the RING domains of E3 ubiquitin ligases. Based on these findings, we purified full-length TMEM55B and designed an in-vitro autoubiquitylation assay which demonstrated that thirteen UBE2 enzymes can generate polyubiquitin chains in the presence of full-length TMEM55B albeit to a low stoichiometric level. Ongoing studies aim to clarify whether TMEM55B acts as a non-canonical RING-type E3 ubiquitin ligase and/or modulates the activity of other E3 ligases.
Conclusions
Our study identifies a novel motif namely TBM in multiple proteins involved in autophagy, lysosomal biology, and ciliogenesis. This motif mediates direct physical interaction with TMEM55B. Preliminary biochemical data hint that TMEM55B may function as an E3 ubiquitin ligase, but further analysis is required to validate this conclusion. Given the growing evidence for crosstalk between lysosomal dysfunction, autophagy, and ciliogenesis in neurodegenerative diseases, further defining TMEM55B substrates and downstream signalling pathways could enable the identification of new biomarkers and therapeutic targets for Parkinson’s disease.