Research Unit Proteolysis
in the Golgi Apparatus
Dr. Matthias Voss
+49 431 880-1676
Research in the Voss lab aims to elucidate how proteolytic processing of glycosylation enzymes in the Golgi apparatus contributes to or is interconnected with the overall organisation of the Golgi. Combining tailored engineered cell culture models, protein biochemistry and cell biology tools as well as state-of-the-art proteomics, the lab studies in particular the intramembrane protease SPPL3, which has emerged as a potent physiological regulator of glycosylation.
Research Vosslab
Proteolysis in the Golgi apparatus
The Golgi apparatus is a central organelle of the secretory pathway in eukaryotic cells. It ensures proper maturation of newly synthesized secretory proteins but also membrane proteins and lipids, most prominently through glycosylation. The precise mode and extent of glycosylation is a critical determinant of secretory and membrane protein function, can globally affect cell-cell interactions and hence is of fundamental importance for the organisation of organs and tissues. Inborn defects in Golgi glycosylation are associated with severe disease in humans and alterations in cellular glycosylation are known to occur in the context of and drive human, e.g. malignancies, underscoring the importance of a properly tuned cellular glycosylation machinery.
Close to 200 membrane-anchored enzymes catalyse glycosylation reactions in the Golgi apparatus. For proper Golgi glycosylation to occur, the overall organization of the Golgi ensures that these important glycosylation enzymes remain in the Golgi apparatus, distributed asymmetrically across the cis/medial/trans axis of the Golgi stack reflecting their precise role in the concerted sequence of Golgi glycosylation reactions, while Golgi cargo (i.e. newly synthesized proteins subject to Golgi glycosylation yet destined for e.g. the cell surface) is rapidly trafficked forward. This particular organisation of the Golgi glycosylation machinery is explained by the fact that glycosylation enzymes are actively retained in the Golgi apparatus through molecular interactions of their membrane-anchors and their cytosolic N-termini.
Interestingly, it is well established that the vast majority of Golgi enzymes is subject to membrane-proximal endoproteolysis. This irreversibly separates a Golgi enzyme’s active ectodomain in the Golgi lumen from its membrane anchor, effectively uncoupling the enzyme from retrieval mechanisms that ensure its correct distribution and residence in the Golgi apparatus.
The intramembrane protease signal peptide peptidase-like 3 (SPPL3) was found to cleave a large number of Golgi enzymes, leading to their secretion and thus depletion from the Golgi apparatus (Voss et al. (2014); Kuhn*, Voss* et al. (2015)). As it can fine-tune abundance and thus intra-Golgi activity of numerous Golgi enzymes, SPPL3 in fact constitutes a potent regulator of cellular glycosylation. Numerous independent genetic screens have since confirmed that hyperglycosylation due to Golgi enzyme accumulation in SPPL3-deficient cells has profound physiological consequences (e.g. in respect to tumour immune evasion) (reviewed in Voss et al. (2024)). This clearly warrants further investigation, in particular as the mechanisms regulating SPPL3 activity and its spatiotemporal access to substrates in the Golgi stack have remained enigmatic.
Recently, through the use of tailored cellular models and state-of-the-art N-terminome analyses, our lab further expanded the spectrum of SPPL3 substrates in the Golgi apparatus and the identification of numerous intramembrane cleavage sites suggested the presence of SPPL3 cleavage products in e.g. human blood (Hobohm et al. (2022)). In parallel, we resolved the intra-Golgi localization of cell-endogenous SPPL3 using high-resolution imaging (Truberg et al. (2022)). To reveal novel aspects of the regulation of SPPL3 activity and localization, the lab’s present interest is centred around the interconnections of SPPL3 activity and the organisation of the Golgi apparatus and we assess changes in SPPL3 activity upon e.g. genetic disruption of Golgi organization. Most recently, this has led us to establish lysosomal sorting as a novel post-cleavage fate of select SPPL3 substrates (Brauer et al. (2024)).
Key papers
Voss M, Künzel U, Higel F, Kuhn PH, Colombo A, Fukumori A, Haug-Kröper M, Klier B, Grammer G, Seidl, A, Schröder B, Obst R, Steiner H, Lichtenthaler SF, Haass C, Fluhrer R (2014). Shedding of glycan- modifying enzymes by signal peptide peptidase-like 3 (SPPL3) regulates cellular N-glycosylation. EMBO J 33, 2890-2905.
Kuhn PH*, Voss M*, Haug-Kröper M, Schröder B, Scheppers U, Bräse S, Haass C, Lichtenthaler SF, Fluhrer R (2015). Secretome analysis identifies novel signal Peptide peptidase-like 3 (Sppl3) substrates and reveals a role of Sppl3 in multiple Golgi glycosylation pathways. Mol Cell Proteomics 14, 1584-1598.
Hobohm L, Koudelka T, Bahr FH, Truberg J, Kapell S, Schacht SS, Meisinger D, Mengel M, Jochimsen A, Hofmann A, Heintz L, Tholey A, Voss M (2022). N-terminome analyses underscore the prevalence of SPPL3-mediated intramembrane proteolysis among Golgi-resident enzymes and its role in Golgi enzyme secretion. Cell Mol Life Sci 79, 185.
Truberg J, Hobohm L, Jochimsen A, Desel C, Schweizer M, Voss M (2022). Endogenous tagging reveals a mid-Golgi localization of the glycosyltransferase-cleaving intramembrane protease SPPL3. Biochim Biophys Acta Mol Cell Res 1869, 119345.
Voss M (2024). Proteolytic cleavage of Golgi glycosyltransferases by SPPL3 and other proteases and its implications for cellular glycosylation. Biochim Biophys Acta Gen Subj 1868, 130668.
Brauer BK*, Chen Z*, Beirow F, Li J, Meisinger D, Capriotti E, Schweizer M, Wagner L, Wienberg J, Hobohm L, Blume L, Qiao W, Narimatsu Y, Carette JE, Clausen H, Winter D, Braulke T, Jabs S‡, Voss M‡ (2024). GOLPH3/3L-dependent Golgi retention of LYSET ensures M6P-tagging of lysosomal enzymes and B4GALT5. EMBO J 43, 6264-6290.