ORCID
https://orcid.org/0000-0002-9268-3907
Date of Award
12-20-2023
Degree Name
Doctor of Philosophy (PhD)
Degree Type
Dissertation
Abstract
Transcription factor EB (TFEB) is a master regulator of lysosomal biogenesis and autophagy that plays a key role in the regulation of cellular clearance pathways. TFEB is regulated via a complex array of post-translational modifications (PTMs), but the exact molecular mechanism that regulates TFEB stability has remained elusive. Here, we show that TFEB levels are critically regulated by a defined phosphorylation-ubiquitination cascade. A human kinome screen identifies IKK (inhibitor of κB kinase) as a TFEB modifier, and a combination of phosphorylation assays, mass spectrometry analyses, and site-specific mutagenesis unveils a previously unrecognized TFEB phospho-degron (423SPFPSLS429) as the target of IKK. We show that the phosphorylation event triggers ubiquitination of adjacent lysine residues (K430 and K431) by the E3 ligase β-TrCP2 (β-Transducin repeat-containing protein 2), thereby tagging TFEB for degradation. Modified TFEB constructs that abolish these PTMs show much increased stability and expression levels but remain equally sensitive to autophagy- or stress-related stimuli while maintaining the capability to promote the expression of TFEB target genes and the clearance of Alzheimer’s associated tau in a cellular model of disease. Our results therefore uncover an IKK/β-TrCP2 phosphorylation-ubiquitination cascade as a major mechanism that governs TFEB stability independently of other TFEB regulators.
Language
English (en)
Chair and Committee
Marco Sardiello
Recommended Citation
Xiong, Yan, "TFEB degradation is regulated by an IKK/β-TrCP2 phosphorylation-ubiquitination cascade" (2023). Arts & Sciences Electronic Theses and Dissertations. 3211.
https://openscholarship.wustl.edu/art_sci_etds/3211