Elevated expression of the adhesion GPCR ADGRL4/ELTD1 promotes endothelial sprouting angiogenesis without activating canonical GPCR signalling

Abstract ADGRL4/ELTD1 is an orphan adhesion GPCR (aGPCR) expressed in endothelial cells that regulates tumour angiogenesis. The majority of aGPCRs are orphan receptors. The Stachel Hypothesis proposes a mechanism for aGPCR activation, in which aGPCRs contain a tethered agonist (termed Stachel) C-ter...

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Autores principales:	David M. Favara, Ines Liebscher, Ali Jazayeri, Madhulika Nambiar, Helen Sheldon, Alison H. Banham, Adrian L. Harris
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/496495a89bd84dd4903cd2c57332c696
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spelling	oai:doaj.org-article:496495a89bd84dd4903cd2c57332c6962021-12-02T17:32:56ZElevated expression of the adhesion GPCR ADGRL4/ELTD1 promotes endothelial sprouting angiogenesis without activating canonical GPCR signalling10.1038/s41598-021-85408-x2045-2322https://doaj.org/article/496495a89bd84dd4903cd2c57332c6962021-04-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-85408-xhttps://doaj.org/toc/2045-2322Abstract ADGRL4/ELTD1 is an orphan adhesion GPCR (aGPCR) expressed in endothelial cells that regulates tumour angiogenesis. The majority of aGPCRs are orphan receptors. The Stachel Hypothesis proposes a mechanism for aGPCR activation, in which aGPCRs contain a tethered agonist (termed Stachel) C-terminal to the GPCR-proteolytic site (GPS) cleavage point which, when exposed, initiates canonical GPCR signalling. This has been shown in a growing number of aGPCRs. We tested this hypothesis on ADGRL4/ELTD1 by designing full length (FL) and C-terminal fragment (CTF) ADGRL4/ELTD1 constructs, and a range of potential Stachel peptides. Constructs were transfected into HEK293T cells and HTRF FRET, luciferase-reporter and Alphascreen GPCR signalling assays were performed. A stable ADGRL4/ELTD1 overexpressing HUVEC line was additionally generated and angiogenesis assays, signalling assays and transcriptional profiling were performed. ADGRL4/ELTD1 has the lowest GC content in the aGPCR family and codon optimisation significantly increased its expression. FL and CTF ADGRL4/ELTD1 constructs, as well as Stachel peptides, did not activate canonical GPCR signalling. Furthermore, stable overexpression of ADGRL4/ELTD1 in HUVECs induced sprouting angiogenesis, lowered in vitro anastomoses, and decreased proliferation, without activating canonical GPCR signalling or MAPK/ERK, PI3K/AKT, JNK, JAK/HIF-1α, beta catenin or STAT3 pathways. Overexpression upregulated ANTXR1, SLC39A6, HBB, CHRNA, ELMOD1, JAG1 and downregulated DLL4, KIT, CCL15, CYP26B1. ADGRL4/ELTD1 specifically regulates the endothelial tip-cell phenotype through yet undefined signalling pathways.David M. FavaraInes LiebscherAli JazayeriMadhulika NambiarHelen SheldonAlison H. BanhamAdrian L. HarrisNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-13 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q David M. Favara Ines Liebscher Ali Jazayeri Madhulika Nambiar Helen Sheldon Alison H. Banham Adrian L. Harris Elevated expression of the adhesion GPCR ADGRL4/ELTD1 promotes endothelial sprouting angiogenesis without activating canonical GPCR signalling
description	Abstract ADGRL4/ELTD1 is an orphan adhesion GPCR (aGPCR) expressed in endothelial cells that regulates tumour angiogenesis. The majority of aGPCRs are orphan receptors. The Stachel Hypothesis proposes a mechanism for aGPCR activation, in which aGPCRs contain a tethered agonist (termed Stachel) C-terminal to the GPCR-proteolytic site (GPS) cleavage point which, when exposed, initiates canonical GPCR signalling. This has been shown in a growing number of aGPCRs. We tested this hypothesis on ADGRL4/ELTD1 by designing full length (FL) and C-terminal fragment (CTF) ADGRL4/ELTD1 constructs, and a range of potential Stachel peptides. Constructs were transfected into HEK293T cells and HTRF FRET, luciferase-reporter and Alphascreen GPCR signalling assays were performed. A stable ADGRL4/ELTD1 overexpressing HUVEC line was additionally generated and angiogenesis assays, signalling assays and transcriptional profiling were performed. ADGRL4/ELTD1 has the lowest GC content in the aGPCR family and codon optimisation significantly increased its expression. FL and CTF ADGRL4/ELTD1 constructs, as well as Stachel peptides, did not activate canonical GPCR signalling. Furthermore, stable overexpression of ADGRL4/ELTD1 in HUVECs induced sprouting angiogenesis, lowered in vitro anastomoses, and decreased proliferation, without activating canonical GPCR signalling or MAPK/ERK, PI3K/AKT, JNK, JAK/HIF-1α, beta catenin or STAT3 pathways. Overexpression upregulated ANTXR1, SLC39A6, HBB, CHRNA, ELMOD1, JAG1 and downregulated DLL4, KIT, CCL15, CYP26B1. ADGRL4/ELTD1 specifically regulates the endothelial tip-cell phenotype through yet undefined signalling pathways.
format	article
author	David M. Favara Ines Liebscher Ali Jazayeri Madhulika Nambiar Helen Sheldon Alison H. Banham Adrian L. Harris
author_facet	David M. Favara Ines Liebscher Ali Jazayeri Madhulika Nambiar Helen Sheldon Alison H. Banham Adrian L. Harris
author_sort	David M. Favara
title	Elevated expression of the adhesion GPCR ADGRL4/ELTD1 promotes endothelial sprouting angiogenesis without activating canonical GPCR signalling
title_short	Elevated expression of the adhesion GPCR ADGRL4/ELTD1 promotes endothelial sprouting angiogenesis without activating canonical GPCR signalling
title_full	Elevated expression of the adhesion GPCR ADGRL4/ELTD1 promotes endothelial sprouting angiogenesis without activating canonical GPCR signalling
title_fullStr	Elevated expression of the adhesion GPCR ADGRL4/ELTD1 promotes endothelial sprouting angiogenesis without activating canonical GPCR signalling
title_full_unstemmed	Elevated expression of the adhesion GPCR ADGRL4/ELTD1 promotes endothelial sprouting angiogenesis without activating canonical GPCR signalling
title_sort	elevated expression of the adhesion gpcr adgrl4/eltd1 promotes endothelial sprouting angiogenesis without activating canonical gpcr signalling
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/496495a89bd84dd4903cd2c57332c696
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Elevated expression of the adhesion GPCR ADGRL4/ELTD1 promotes endothelial sprouting angiogenesis without activating canonical GPCR signalling

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