Molecular determinants of PI3Kγ-mediated activation downstream of G-protein-coupled receptors (GPCRs)

Oscar Vadas, Hashem A. Dbouk, Aliaksei Shymanets, Olga Perisic, John E. Burke, Widian F Abi Saab, Bassem D. Khalil, Christian Harteneck, Anne R. Bresnick, Bernd Nürnberg, Jonathan M. Backer, Roger L. Williams

Research output: Contribution to journalArticle

57 Citations (Scopus)

Abstract

Phosphoinositide 3-kinase gamma (PI3Kγ) has profound roles downstream of G-protein-coupled receptors in inflammation, cardiac function, and tumor progression. To gain insight into how the enzyme's activity is shaped by association with its p101 adaptor subunit, lipid membranes, and Gβγ heterodimers, we mapped these regulatory interactions using hydrogen-deuterium exchange mass spectrometry. We identify residues in both the p110γ and p101 subunits that contribute critical interactions with Gβγ heterodimers, leading to PI3Kγ activation. Mutating Gβγ- interaction sites of either p110γ or p101 ablates G-protein-coupled receptor-mediated signaling to p110γ/p101 in cells and severely affects chemotaxis and cell transformation induced by PI3Kγ overexpression. Hydrogen-deuterium exchange mass spectrometry shows that association with the p101 regulatory subunit causes substantial protection of the RBD-C2 linker as well as the helical domain of p110γ. Lipid interaction massively exposes that same helical site, which is then stabilized by Gβγ. Membrane-elicited conformational change of the helical domain could help prepare the enzyme for Gβγ binding. Our studies and others identify the helical domain of the class I PI3Ks as a hub for diverse regulatory interactions that include the p101, p87 (also known as p84), and p85 adaptor subunits; Rab5 and Gβγ heterodimers; and the β-adrenergic receptor kinase.

Original languageEnglish (US)
Pages (from-to)18862-18867
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume110
Issue number47
DOIs
StatePublished - Nov 19 2013

Fingerprint

1-Phosphatidylinositol 4-Kinase
G-Protein-Coupled Receptors
Deuterium
Hydrogen
Mass Spectrometry
Heart Neoplasms
Chemotaxis
Enzymes
Membrane Lipids
Phosphatidylinositol 3-Kinases
Adrenergic Receptors
Phosphotransferases
Inflammation
Lipids
Membranes

ASJC Scopus subject areas

  • General

Cite this

Molecular determinants of PI3Kγ-mediated activation downstream of G-protein-coupled receptors (GPCRs). / Vadas, Oscar; Dbouk, Hashem A.; Shymanets, Aliaksei; Perisic, Olga; Burke, John E.; Saab, Widian F Abi; Khalil, Bassem D.; Harteneck, Christian; Bresnick, Anne R.; Nürnberg, Bernd; Backer, Jonathan M.; Williams, Roger L.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 110, No. 47, 19.11.2013, p. 18862-18867.

Research output: Contribution to journalArticle

Vadas, O, Dbouk, HA, Shymanets, A, Perisic, O, Burke, JE, Saab, WFA, Khalil, BD, Harteneck, C, Bresnick, AR, Nürnberg, B, Backer, JM & Williams, RL 2013, 'Molecular determinants of PI3Kγ-mediated activation downstream of G-protein-coupled receptors (GPCRs)', Proceedings of the National Academy of Sciences of the United States of America, vol. 110, no. 47, pp. 18862-18867. https://doi.org/10.1073/pnas.1304801110
Vadas, Oscar ; Dbouk, Hashem A. ; Shymanets, Aliaksei ; Perisic, Olga ; Burke, John E. ; Saab, Widian F Abi ; Khalil, Bassem D. ; Harteneck, Christian ; Bresnick, Anne R. ; Nürnberg, Bernd ; Backer, Jonathan M. ; Williams, Roger L. / Molecular determinants of PI3Kγ-mediated activation downstream of G-protein-coupled receptors (GPCRs). In: Proceedings of the National Academy of Sciences of the United States of America. 2013 ; Vol. 110, No. 47. pp. 18862-18867.
@article{5339c3a764764257919332eac710bd6e,
title = "Molecular determinants of PI3Kγ-mediated activation downstream of G-protein-coupled receptors (GPCRs)",
abstract = "Phosphoinositide 3-kinase gamma (PI3Kγ) has profound roles downstream of G-protein-coupled receptors in inflammation, cardiac function, and tumor progression. To gain insight into how the enzyme's activity is shaped by association with its p101 adaptor subunit, lipid membranes, and Gβγ heterodimers, we mapped these regulatory interactions using hydrogen-deuterium exchange mass spectrometry. We identify residues in both the p110γ and p101 subunits that contribute critical interactions with Gβγ heterodimers, leading to PI3Kγ activation. Mutating Gβγ- interaction sites of either p110γ or p101 ablates G-protein-coupled receptor-mediated signaling to p110γ/p101 in cells and severely affects chemotaxis and cell transformation induced by PI3Kγ overexpression. Hydrogen-deuterium exchange mass spectrometry shows that association with the p101 regulatory subunit causes substantial protection of the RBD-C2 linker as well as the helical domain of p110γ. Lipid interaction massively exposes that same helical site, which is then stabilized by Gβγ. Membrane-elicited conformational change of the helical domain could help prepare the enzyme for Gβγ binding. Our studies and others identify the helical domain of the class I PI3Ks as a hub for diverse regulatory interactions that include the p101, p87 (also known as p84), and p85 adaptor subunits; Rab5 and Gβγ heterodimers; and the β-adrenergic receptor kinase.",
author = "Oscar Vadas and Dbouk, {Hashem A.} and Aliaksei Shymanets and Olga Perisic and Burke, {John E.} and Saab, {Widian F Abi} and Khalil, {Bassem D.} and Christian Harteneck and Bresnick, {Anne R.} and Bernd N{\"u}rnberg and Backer, {Jonathan M.} and Williams, {Roger L.}",
year = "2013",
month = "11",
day = "19",
doi = "10.1073/pnas.1304801110",
language = "English (US)",
volume = "110",
pages = "18862--18867",
journal = "Proceedings of the National Academy of Sciences of the United States of America",
issn = "0027-8424",
number = "47",

}

TY - JOUR

T1 - Molecular determinants of PI3Kγ-mediated activation downstream of G-protein-coupled receptors (GPCRs)

AU - Vadas, Oscar

AU - Dbouk, Hashem A.

AU - Shymanets, Aliaksei

AU - Perisic, Olga

AU - Burke, John E.

AU - Saab, Widian F Abi

AU - Khalil, Bassem D.

AU - Harteneck, Christian

AU - Bresnick, Anne R.

AU - Nürnberg, Bernd

AU - Backer, Jonathan M.

AU - Williams, Roger L.

PY - 2013/11/19

Y1 - 2013/11/19

N2 - Phosphoinositide 3-kinase gamma (PI3Kγ) has profound roles downstream of G-protein-coupled receptors in inflammation, cardiac function, and tumor progression. To gain insight into how the enzyme's activity is shaped by association with its p101 adaptor subunit, lipid membranes, and Gβγ heterodimers, we mapped these regulatory interactions using hydrogen-deuterium exchange mass spectrometry. We identify residues in both the p110γ and p101 subunits that contribute critical interactions with Gβγ heterodimers, leading to PI3Kγ activation. Mutating Gβγ- interaction sites of either p110γ or p101 ablates G-protein-coupled receptor-mediated signaling to p110γ/p101 in cells and severely affects chemotaxis and cell transformation induced by PI3Kγ overexpression. Hydrogen-deuterium exchange mass spectrometry shows that association with the p101 regulatory subunit causes substantial protection of the RBD-C2 linker as well as the helical domain of p110γ. Lipid interaction massively exposes that same helical site, which is then stabilized by Gβγ. Membrane-elicited conformational change of the helical domain could help prepare the enzyme for Gβγ binding. Our studies and others identify the helical domain of the class I PI3Ks as a hub for diverse regulatory interactions that include the p101, p87 (also known as p84), and p85 adaptor subunits; Rab5 and Gβγ heterodimers; and the β-adrenergic receptor kinase.

AB - Phosphoinositide 3-kinase gamma (PI3Kγ) has profound roles downstream of G-protein-coupled receptors in inflammation, cardiac function, and tumor progression. To gain insight into how the enzyme's activity is shaped by association with its p101 adaptor subunit, lipid membranes, and Gβγ heterodimers, we mapped these regulatory interactions using hydrogen-deuterium exchange mass spectrometry. We identify residues in both the p110γ and p101 subunits that contribute critical interactions with Gβγ heterodimers, leading to PI3Kγ activation. Mutating Gβγ- interaction sites of either p110γ or p101 ablates G-protein-coupled receptor-mediated signaling to p110γ/p101 in cells and severely affects chemotaxis and cell transformation induced by PI3Kγ overexpression. Hydrogen-deuterium exchange mass spectrometry shows that association with the p101 regulatory subunit causes substantial protection of the RBD-C2 linker as well as the helical domain of p110γ. Lipid interaction massively exposes that same helical site, which is then stabilized by Gβγ. Membrane-elicited conformational change of the helical domain could help prepare the enzyme for Gβγ binding. Our studies and others identify the helical domain of the class I PI3Ks as a hub for diverse regulatory interactions that include the p101, p87 (also known as p84), and p85 adaptor subunits; Rab5 and Gβγ heterodimers; and the β-adrenergic receptor kinase.

UR - http://www.scopus.com/inward/record.url?scp=84888121509&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84888121509&partnerID=8YFLogxK

U2 - 10.1073/pnas.1304801110

DO - 10.1073/pnas.1304801110

M3 - Article

VL - 110

SP - 18862

EP - 18867

JO - Proceedings of the National Academy of Sciences of the United States of America

JF - Proceedings of the National Academy of Sciences of the United States of America

SN - 0027-8424

IS - 47

ER -