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The residue at position 5 of the N-terminal region of Src and Fyn modulates their myristoylation, palmitoylation, and membrane interactions

The interactions of Src family kinases (SFKs) with the plasma membrane are crucial for their activity. They depend on their fatty-acylated N-termini, containing N-myristate and either a polybasic cluster (in Src) or palmitoylation sites (e.g., Fyn). To investigate the roles of these moieties in SFK...

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Autores principales: Gottlieb-Abraham, Efrat, Gutman, Orit, Pai, Govind M., Rubio, Ignacio, Henis, Yoav I.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The American Society for Cell Biology 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5170614/
https://www.ncbi.nlm.nih.gov/pubmed/27733622
http://dx.doi.org/10.1091/mbc.E16-08-0622
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author Gottlieb-Abraham, Efrat
Gutman, Orit
Pai, Govind M.
Rubio, Ignacio
Henis, Yoav I.
author_facet Gottlieb-Abraham, Efrat
Gutman, Orit
Pai, Govind M.
Rubio, Ignacio
Henis, Yoav I.
author_sort Gottlieb-Abraham, Efrat
collection PubMed
description The interactions of Src family kinases (SFKs) with the plasma membrane are crucial for their activity. They depend on their fatty-acylated N-termini, containing N-myristate and either a polybasic cluster (in Src) or palmitoylation sites (e.g., Fyn). To investigate the roles of these moieties in SFK membrane association, we used fluorescence recovery after photobleaching beam-size analysis to study the membrane interactions of c-Src-GFP (green fluorescent protein) or Fyn-GFP fatty-acylation mutants. Our studies showed for the first time that the membrane association of Fyn is more stable than that of Src, an effect lost in a Fyn mutant lacking the palmitoylation sites. Unexpectedly, Src-S3C/S6C (containing cysteines at positions 3/6, which are palmitoylated in Fyn) exhibited fast cytoplasmic diffusion insensitive to palmitoylation inhibitors, suggesting defective fatty acylation. Further replacement of the charged Lys-5 by neutral Gln to resemble Fyn (Src-S3C/S6C/K5Q) restored Fyn-like membrane interactions, indicating that Lys-5 in the context of Src-S3C/S6C interferes with its myristoylation/palmitoylation. This was validated by direct myristoylation and palmitoylation studies, which indicated that the residue at position 5 regulates the membrane interactions of Src versus Fyn. Moreover, the palmitoylation levels correlated with targeting to detergent-resistant membranes (rafts) and to caveolin-1. Palmitoylation-dependent preferential containment of Fyn in rafts may contribute to its lower transformation potential.
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spelling pubmed-51706142017-02-16 The residue at position 5 of the N-terminal region of Src and Fyn modulates their myristoylation, palmitoylation, and membrane interactions Gottlieb-Abraham, Efrat Gutman, Orit Pai, Govind M. Rubio, Ignacio Henis, Yoav I. Mol Biol Cell Articles The interactions of Src family kinases (SFKs) with the plasma membrane are crucial for their activity. They depend on their fatty-acylated N-termini, containing N-myristate and either a polybasic cluster (in Src) or palmitoylation sites (e.g., Fyn). To investigate the roles of these moieties in SFK membrane association, we used fluorescence recovery after photobleaching beam-size analysis to study the membrane interactions of c-Src-GFP (green fluorescent protein) or Fyn-GFP fatty-acylation mutants. Our studies showed for the first time that the membrane association of Fyn is more stable than that of Src, an effect lost in a Fyn mutant lacking the palmitoylation sites. Unexpectedly, Src-S3C/S6C (containing cysteines at positions 3/6, which are palmitoylated in Fyn) exhibited fast cytoplasmic diffusion insensitive to palmitoylation inhibitors, suggesting defective fatty acylation. Further replacement of the charged Lys-5 by neutral Gln to resemble Fyn (Src-S3C/S6C/K5Q) restored Fyn-like membrane interactions, indicating that Lys-5 in the context of Src-S3C/S6C interferes with its myristoylation/palmitoylation. This was validated by direct myristoylation and palmitoylation studies, which indicated that the residue at position 5 regulates the membrane interactions of Src versus Fyn. Moreover, the palmitoylation levels correlated with targeting to detergent-resistant membranes (rafts) and to caveolin-1. Palmitoylation-dependent preferential containment of Fyn in rafts may contribute to its lower transformation potential. The American Society for Cell Biology 2016-12-01 /pmc/articles/PMC5170614/ /pubmed/27733622 http://dx.doi.org/10.1091/mbc.E16-08-0622 Text en © 2016 Gottlieb-Abraham et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0). “ASCB®,” “The American Society for Cell Biology®,” and “Molecular Biology of the Cell®” are registered trademarks of The American Society for Cell Biology.
spellingShingle Articles
Gottlieb-Abraham, Efrat
Gutman, Orit
Pai, Govind M.
Rubio, Ignacio
Henis, Yoav I.
The residue at position 5 of the N-terminal region of Src and Fyn modulates their myristoylation, palmitoylation, and membrane interactions
title The residue at position 5 of the N-terminal region of Src and Fyn modulates their myristoylation, palmitoylation, and membrane interactions
title_full The residue at position 5 of the N-terminal region of Src and Fyn modulates their myristoylation, palmitoylation, and membrane interactions
title_fullStr The residue at position 5 of the N-terminal region of Src and Fyn modulates their myristoylation, palmitoylation, and membrane interactions
title_full_unstemmed The residue at position 5 of the N-terminal region of Src and Fyn modulates their myristoylation, palmitoylation, and membrane interactions
title_short The residue at position 5 of the N-terminal region of Src and Fyn modulates their myristoylation, palmitoylation, and membrane interactions
title_sort residue at position 5 of the n-terminal region of src and fyn modulates their myristoylation, palmitoylation, and membrane interactions
topic Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5170614/
https://www.ncbi.nlm.nih.gov/pubmed/27733622
http://dx.doi.org/10.1091/mbc.E16-08-0622
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