Background: The homeostasis of palmitoylation and depalmitoylation is
involved in various cellular processes, the disruption of which induces severe
physiological consequences. Acyl-protein thioesterase (APT) and palmitoyl-protein
thioesterases (PPT) catalyze the depalmitoylation process. The natural mutation in
human PPT1 caused neurodegenerative disease, yet the understanding of APT1
remains to be elucidated. While the deletion of APT1 in mice turned out to be potentially
embryonically lethal, the decoding of its function strictly relied on the identification of its
Objective: To determine the potential substrates of APT1 by using the generated
human APT1 knockout cell line.
Methods : The combined techniques of palmitoyl-protein enrichment and massspectrometry
were used to analyze the different proteins. Palmitoyl-proteins both in
HEK293T and APT1-KO cells were extracted by resin-assisted capture (RAC) and data
independent acquisition (DIA) quantitative method of proteomics for data collection.
Results: In total, 382 proteins were identified. The gene ontology classification
segregated these proteins into diverse biological pathways e.g. endoplasmic reticulum
process and ubiquitin-mediated proteolysis. A few potential substrates were selected for
verification; indeed, major proteins were palmitoylated. Importantly, their levels of
palmitoylation were clearly changed in APT1-KO cells. Interestingly, the proliferation of
APT1-KO cells escalated dramatically as compared to that of the WT cells, which could
be rescued by APT1 overexpression.
Conclusion: Our study provides a large scale of potential substrates of APT1, thus
facilitating the understanding of its intervened molecular functions.