Research Project 7:
Adaptive lysosomal signaling mechanisms in CLN3-defective cells
Recent findings discover that lysosomal biogenesis, autophagy, exocytosis, and lysosome-associated processes such as phagocytosis, are subject to global transcriptional regulation in which the transcription factor EB (TFEB) is thought to play a key role (1). The transcriptional activity of TFEB is regulated by phosphorylation/dephosphorylation processes catalyzed by various kinases (e.g. mTORC1, ERK1/2 or protein kinase C beta; 2) and the phosphatase calcineurin (3) which determine the translocation of TFEB into the nucleus. TFEB is posttranslationally modified at multiple amino acid residues that leads to numerous cellular isoforms of different isoelectric points. The functional significance of these TFEB isoforms and their modification stoichiometry for transcriptional activity, subcellular localization and stability of TFEB remains unclear. Furthermore, regulatory mechanisms in response to environmental cues and the molecular components affecting the modification of TFEB at different sites and subsequently lysosome functions are unsolved.
In this project we will isolate distinct TFEB isoforms and identify their posttranslationally modified residues. In the second step we will investigate whether these modifications correlate with the expression of defined lysosomal genes (expression studies followed by MS-based quantitative lysosome proteomics; 4,5).
- Settembre C, Fraldi A, Medina DL, Ballabio A (2013) Signals from the lysosome: a control centre for cellular clearance and energy metabolism. Nat Rev Mol cell Biol 14: 283-96
- Settembre C, diMalta C, Polito VA, Garcia Arencibia M, Vetrini F, Erdin S, Erdin SU, Huynh T, Medina D, Colella P, Sardiello M, Rubinsztein DC, Ballabio A (2011) TFEB links autophagy to lysosomal biogenesis. Science 332: 1429-33
- Medina DL, Di Paola S, Peluso I, Armani A, De Stefani D, Venditti R, Montefusco S, Scotto-Rosato A, Prezioso C, Forrester A, Settembre C, Wang W, Gao Q, Xu H, Sandri M, Rizzuto R, De Matteis MA, Ballabio A (2015) Lysosomal calcium signalling regulates autophagy through calcineurin and TFEB. Nat Cell Biol 17: 288-99
- Markmann S, Thelen M, Willnow T, Gieselmann V, Braulke T, Kollmann K (2015) Lrp1/LDL receptor play critical roles in mannose 6-phosphate-independent lysosomal enzyme targeting. Traffic 16: 743-59
- Markmann S, Krambeck S, Hughes CJ, Mirzaian M, Aerts JMFG, Saftig P, Schweizer M, Vissers JPC, Braulke T, Damme M (2017) Quantitative proteome analysis of mouse liver lysosomes provides evidence for mannose 6-phosphate-independent targeting mechanisms of acid hydrolases in mucolipidosis II. Mol Cell Proteomics 16: 438-50