CellPalmSeq: A curated RNAseq database of palmitoylating and de-palmitoylating enzyme expression in human cell types and laboratory cell lines

Wild, Angela R.; Hogg, Peter W; Flibotte, Stéphane; Kochhar, Shruti; Hollman, Rocio; Haas, Kurt; Bamji, Shernaz X.

doi:10.3389/fphys.2023.1110550

Cited by 11 publications

(7 citation statements)

References 60 publications

(76 reference statements)

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“…To identify the palmitoylated cysteines in caveolin‐3 we first generated single cysteine to alanine mutations and expressed them in HEK‐293 cells, which possess a full repertoire of zDHHC‐PATs 30,31 . In HEK‐293 cells, low levels of expression of endogenous caveolin‐1 have previously been observed 32 but caveolin‐3 is not expressed at all 33 .…”

Section: Resultsmentioning

confidence: 99%

Cysteine post‐translational modifications regulate protein interactions of caveolin‐3

Ashford,

Kuo,

Dunning

et al. 2024

The FASEB Journal

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Caveolae are small flask‐shaped invaginations of the surface membrane which are proposed to recruit and co‐localize signaling molecules. The distinctive caveolar shape is achieved by the oligomeric structural protein caveolin, of which three isoforms exist. Aside from the finding that caveolin‐3 is specifically expressed in muscle, functional differences between the caveolin isoforms have not been rigorously investigated. Caveolin‐3 is relatively cysteine‐rich compared to caveolins 1 and 2, so we investigated its cysteine post‐translational modifications. We find that caveolin‐3 is palmitoylated at 6 cysteines and becomes glutathiolated following redox stress. We map the caveolin‐3 palmitoylation sites to a cluster of cysteines in its C terminal membrane domain, and the glutathiolation site to an N terminal cysteine close to the region of caveolin‐3 proposed to engage in protein interactions. Glutathiolation abolishes caveolin‐3 interaction with heterotrimeric G protein alpha subunits. Our results indicate that a caveolin‐3 oligomer contains up to 66 palmitates, compared to up to 33 for caveolin‐1. The additional palmitoylation sites in caveolin‐3 therefore provide a mechanistic basis by which caveolae in smooth and striated muscle can possess unique phospholipid and protein cargoes. These unique adaptations of the muscle‐specific caveolin isoform have important implications for caveolar assembly and signaling.

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Section: Resultsmentioning

confidence: 99%

Cysteine post‐translational modifications regulate protein interactions of caveolin‐3

Ashford,

Kuo,

Dunning

et al. 2024

The FASEB Journal

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“…Multiple zDHHC-PATs are capable of palmitoylating IFITM3 [ 41 , 46 ]. The recent description of cell-specific expression maps of palmitoylating and depalmitoylating enzymes will clearly be an important tool to identify appropriate cellular models to evaluate the efficacy of zDHHC-PAT inhibitors and degraders [ 47 ].…”

Section: Discussionmentioning

confidence: 99%

Targeted degradation of zDHHC-PATs decreases substrate S-palmitoylation

Bai,

Gallen,

Memarzadeh

et al. 2024

PLoS ONE

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Reversible S-palmitoylation of protein cysteines, catalysed by a family of integral membrane zDHHC-motif containing palmitoyl acyl transferases (zDHHC-PATs), controls the localisation, activity, and interactions of numerous integral and peripheral membrane proteins. There are compelling reasons to want to inhibit the activity of individual zDHHC-PATs in both the laboratory and the clinic, but the specificity of existing tools is poor. Given the extensive conservation of the zDHHC-PAT active site, development of isoform-specific competitive inhibitors is highly challenging. We therefore hypothesised that proteolysis-targeting chimaeras (PROTACs) may offer greater specificity to target this class of enzymes. In proof-of-principle experiments we engineered cell lines expressing tetracycline-inducible Halo-tagged zDHHC5 or zDHHC20, and evaluated the impact of Halo-PROTACs on zDHHC-PAT expression and substrate palmitoylation. In HEK-derived FT-293 cells, Halo-zDHHC5 degradation significantly decreased palmitoylation of its substrate phospholemman, and Halo-zDHHC20 degradation significantly diminished palmitoylation of its substrate IFITM3, but not of the SARS-CoV-2 spike protein. In contrast, in a second kidney derived cell line, Vero E6, Halo-zDHHC20 degradation did not alter palmitoylation of either IFITM3 or SARS-CoV-2 spike. We conclude from these experiments that PROTAC-mediated targeting of zDHHC-PATs to decrease substrate palmitoylation is feasible. However, given the well-established degeneracy in the zDHHC-PAT family, in some settings the activity of non-targeted zDHHC-PATs may substitute and preserve substrate palmitoylation.

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“…This could help explain why ABHD17 appears to act primarily on substrates at the plasma membrane (Remsberg et al ., 2021). How ABHD17 localization affects activity and substrate specificity, and whether ABHD17 proteins are also regulated by post-translational modification or expression levels (Abazari et al ., 2023; Wild et al ., 2023) are important areas of future research. A complete understanding of ABHD17 regulation will be helpful in developing therapeutic approaches that target ABHD17 and will increase our understanding of related thioesterases.…”

Section: Discussionmentioning

confidence: 99%

An acylated N-terminus and a conserved loop regulate the activity of the ABHD17 de-acylase

Holme,

Sapia,

Davey

et al. 2024

Preprint

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The dynamic addition and removal of long chain fatty acids modulates protein function and localization. The alpha/beta hydrolase domain-containing (ABHD) 17 enzymes remove acyl chains from membrane localized proteins such as the oncoprotein NRas, but how the ABHD17 proteins are regulated is unknown. Here, we used cell-based studies and molecular dynamics simulations to show that ABHD17 activity is controlled by two mobile elements - an S-acylated N-terminal helix and a loop - that flank the substrate-binding pocket. S-acylation at multiple sites buries the helix in the membrane, which allows hydrophobic residues in the loop to interact with the bilayer. This stabilizes the conformation of both helix and loop, alters the conformation of the binding pocket and optimally positions the enzyme for substrate engagement. S-acylation may be a general feature of acyl-protein thioesterases. By providing a mechanistic understanding of how the lipid modification of a lipid-removing enzyme promotes its enzymatic activity, this work contributes to our understanding of cellular S-acylation cycles.

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CellPalmSeq: A curated RNAseq database of palmitoylating and de-palmitoylating enzyme expression in human cell types and laboratory cell lines

Cited by 11 publications

References 60 publications

Cysteine post‐translational modifications regulate protein interactions of caveolin‐3

Cysteine post‐translational modifications regulate protein interactions of caveolin‐3

Targeted degradation of zDHHC-PATs decreases substrate S-palmitoylation

An acylated N-terminus and a conserved loop regulate the activity of the ABHD17 de-acylase

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