Cargando…

The S-palmitoylome and DHHC-PAT interactome of Drosophila melanogaster S2R+ cells indicate a high degree of conservation to mammalian palmitoylomes

Protein S-palmitoylation, the addition of a long-chain fatty acid to target proteins, is among the most frequent reversible protein modifications in Metazoa, affecting subcellular protein localization, trafficking and protein-protein interactions. S-palmitoylated proteins are abundant in the neurona...

Descripción completa

Detalles Bibliográficos
Autores principales: Porcellato, Elena, González-Sánchez, Juan Carlos, Ahlmann-Eltze, Constantin, Elsakka, Mahmoud Ali, Shapira, Itamar, Fritsch, Jürgen, Navarro, Juan Antonio, Anders, Simon, Russell, Robert B., Wieland, Felix T., Metzendorf, Christoph
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9374236/
https://www.ncbi.nlm.nih.gov/pubmed/35960718
http://dx.doi.org/10.1371/journal.pone.0261543
_version_ 1784767746005794816
author Porcellato, Elena
González-Sánchez, Juan Carlos
Ahlmann-Eltze, Constantin
Elsakka, Mahmoud Ali
Shapira, Itamar
Fritsch, Jürgen
Navarro, Juan Antonio
Anders, Simon
Russell, Robert B.
Wieland, Felix T.
Metzendorf, Christoph
author_facet Porcellato, Elena
González-Sánchez, Juan Carlos
Ahlmann-Eltze, Constantin
Elsakka, Mahmoud Ali
Shapira, Itamar
Fritsch, Jürgen
Navarro, Juan Antonio
Anders, Simon
Russell, Robert B.
Wieland, Felix T.
Metzendorf, Christoph
author_sort Porcellato, Elena
collection PubMed
description Protein S-palmitoylation, the addition of a long-chain fatty acid to target proteins, is among the most frequent reversible protein modifications in Metazoa, affecting subcellular protein localization, trafficking and protein-protein interactions. S-palmitoylated proteins are abundant in the neuronal system and are associated with neuronal diseases and cancer. Despite the importance of this post-translational modification, it has not been thoroughly studied in the model organism Drosophila melanogaster. Here we present the palmitoylome of Drosophila S2R+ cells, comprising 198 proteins, an estimated 3.5% of expressed genes in these cells. Comparison of orthologs between mammals and Drosophila suggests that S-palmitoylated proteins are more conserved between these distant phyla than non-S-palmitoylated proteins. To identify putative client proteins and interaction partners of the DHHC family of protein acyl-transferases (PATs) we established DHHC-BioID, a proximity biotinylation-based method. In S2R+ cells, ectopic expression of the DHHC-PAT dHip14-BioID in combination with Snap24 or an interaction-deficient Snap24-mutant as a negative control, resulted in biotinylation of Snap24 but not the Snap24-mutant. DHHC-BioID in S2R+ cells using 10 different DHHC-PATs as bait identified 520 putative DHHC-PAT interaction partners of which 48 were S-palmitoylated and are therefore putative DHHC-PAT client proteins. Comparison of putative client protein/DHHC-PAT combinations indicates that CG8314, CG5196, CG5880 and Patsas have a preference for transmembrane proteins, while S-palmitoylated proteins with the Hip14-interaction motif are most enriched by DHHC-BioID variants of approximated and dHip14. Finally, we show that BioID is active in larval and adult Drosophila and that dHip14-BioID rescues dHip14 mutant flies, indicating that DHHC-BioID is non-toxic. In summary we provide the first systematic analysis of a Drosophila palmitoylome. We show that DHHC-BioID is sensitive and specific enough to identify DHHC-PAT client proteins and provide DHHC-PAT assignment for ca. 25% of the S2R+ cell palmitoylome, providing a valuable resource. In addition, we establish DHHC-BioID as a useful concept for the identification of tissue-specific DHHC-PAT interactomes in Drosophila.
format Online
Article
Text
id pubmed-9374236
institution National Center for Biotechnology Information
language English
publishDate 2022
publisher Public Library of Science
record_format MEDLINE/PubMed
spelling pubmed-93742362022-08-13 The S-palmitoylome and DHHC-PAT interactome of Drosophila melanogaster S2R+ cells indicate a high degree of conservation to mammalian palmitoylomes Porcellato, Elena González-Sánchez, Juan Carlos Ahlmann-Eltze, Constantin Elsakka, Mahmoud Ali Shapira, Itamar Fritsch, Jürgen Navarro, Juan Antonio Anders, Simon Russell, Robert B. Wieland, Felix T. Metzendorf, Christoph PLoS One Research Article Protein S-palmitoylation, the addition of a long-chain fatty acid to target proteins, is among the most frequent reversible protein modifications in Metazoa, affecting subcellular protein localization, trafficking and protein-protein interactions. S-palmitoylated proteins are abundant in the neuronal system and are associated with neuronal diseases and cancer. Despite the importance of this post-translational modification, it has not been thoroughly studied in the model organism Drosophila melanogaster. Here we present the palmitoylome of Drosophila S2R+ cells, comprising 198 proteins, an estimated 3.5% of expressed genes in these cells. Comparison of orthologs between mammals and Drosophila suggests that S-palmitoylated proteins are more conserved between these distant phyla than non-S-palmitoylated proteins. To identify putative client proteins and interaction partners of the DHHC family of protein acyl-transferases (PATs) we established DHHC-BioID, a proximity biotinylation-based method. In S2R+ cells, ectopic expression of the DHHC-PAT dHip14-BioID in combination with Snap24 or an interaction-deficient Snap24-mutant as a negative control, resulted in biotinylation of Snap24 but not the Snap24-mutant. DHHC-BioID in S2R+ cells using 10 different DHHC-PATs as bait identified 520 putative DHHC-PAT interaction partners of which 48 were S-palmitoylated and are therefore putative DHHC-PAT client proteins. Comparison of putative client protein/DHHC-PAT combinations indicates that CG8314, CG5196, CG5880 and Patsas have a preference for transmembrane proteins, while S-palmitoylated proteins with the Hip14-interaction motif are most enriched by DHHC-BioID variants of approximated and dHip14. Finally, we show that BioID is active in larval and adult Drosophila and that dHip14-BioID rescues dHip14 mutant flies, indicating that DHHC-BioID is non-toxic. In summary we provide the first systematic analysis of a Drosophila palmitoylome. We show that DHHC-BioID is sensitive and specific enough to identify DHHC-PAT client proteins and provide DHHC-PAT assignment for ca. 25% of the S2R+ cell palmitoylome, providing a valuable resource. In addition, we establish DHHC-BioID as a useful concept for the identification of tissue-specific DHHC-PAT interactomes in Drosophila. Public Library of Science 2022-08-12 /pmc/articles/PMC9374236/ /pubmed/35960718 http://dx.doi.org/10.1371/journal.pone.0261543 Text en © 2022 Porcellato et al https://creativecommons.org/licenses/by/4.0/This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
Porcellato, Elena
González-Sánchez, Juan Carlos
Ahlmann-Eltze, Constantin
Elsakka, Mahmoud Ali
Shapira, Itamar
Fritsch, Jürgen
Navarro, Juan Antonio
Anders, Simon
Russell, Robert B.
Wieland, Felix T.
Metzendorf, Christoph
The S-palmitoylome and DHHC-PAT interactome of Drosophila melanogaster S2R+ cells indicate a high degree of conservation to mammalian palmitoylomes
title The S-palmitoylome and DHHC-PAT interactome of Drosophila melanogaster S2R+ cells indicate a high degree of conservation to mammalian palmitoylomes
title_full The S-palmitoylome and DHHC-PAT interactome of Drosophila melanogaster S2R+ cells indicate a high degree of conservation to mammalian palmitoylomes
title_fullStr The S-palmitoylome and DHHC-PAT interactome of Drosophila melanogaster S2R+ cells indicate a high degree of conservation to mammalian palmitoylomes
title_full_unstemmed The S-palmitoylome and DHHC-PAT interactome of Drosophila melanogaster S2R+ cells indicate a high degree of conservation to mammalian palmitoylomes
title_short The S-palmitoylome and DHHC-PAT interactome of Drosophila melanogaster S2R+ cells indicate a high degree of conservation to mammalian palmitoylomes
title_sort s-palmitoylome and dhhc-pat interactome of drosophila melanogaster s2r+ cells indicate a high degree of conservation to mammalian palmitoylomes
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9374236/
https://www.ncbi.nlm.nih.gov/pubmed/35960718
http://dx.doi.org/10.1371/journal.pone.0261543
work_keys_str_mv AT porcellatoelena thespalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT gonzalezsanchezjuancarlos thespalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT ahlmanneltzeconstantin thespalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT elsakkamahmoudali thespalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT shapiraitamar thespalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT fritschjurgen thespalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT navarrojuanantonio thespalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT anderssimon thespalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT russellrobertb thespalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT wielandfelixt thespalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT metzendorfchristoph thespalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT porcellatoelena spalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT gonzalezsanchezjuancarlos spalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT ahlmanneltzeconstantin spalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT elsakkamahmoudali spalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT shapiraitamar spalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT fritschjurgen spalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT navarrojuanantonio spalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT anderssimon spalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT russellrobertb spalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT wielandfelixt spalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes
AT metzendorfchristoph spalmitoylomeanddhhcpatinteractomeofdrosophilamelanogasters2rcellsindicateahighdegreeofconservationtomammalianpalmitoylomes