Inside-out regulation of ICAM-1 dynamics in TNF-alpha-activated endothelium.

<h4>Background</h4>During transendothelial migration, leukocytes use adhesion molecules, such as ICAM-1, to adhere to the endothelium. ICAM-1 is a dynamic molecule that is localized in the apical membrane of the endothelium and clusters upon binding to leukocytes. However, not much is kn...

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Autores principales:	Jaap D van Buul, Jos van Rijssel, Floris P J van Alphen, Mark Hoogenboezem, Simon Tol, Kees A Hoeben, Jan van Marle, Erik P J Mul, Peter L Hordijk
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2010
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/7193af2812dc4e36a4aa181fd4b28836
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spelling	oai:doaj.org-article:7193af2812dc4e36a4aa181fd4b288362021-12-02T20:20:27ZInside-out regulation of ICAM-1 dynamics in TNF-alpha-activated endothelium.1932-620310.1371/journal.pone.0011336https://doaj.org/article/7193af2812dc4e36a4aa181fd4b288362010-06-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/20596527/pdf/?tool=EBIhttps://doaj.org/toc/1932-6203<h4>Background</h4>During transendothelial migration, leukocytes use adhesion molecules, such as ICAM-1, to adhere to the endothelium. ICAM-1 is a dynamic molecule that is localized in the apical membrane of the endothelium and clusters upon binding to leukocytes. However, not much is known about the regulation of ICAM-1 clustering and whether membrane dynamics are linked to the ability of ICAM-1 to cluster and bind leukocyte integrins. Therefore, we studied the dynamics of endothelial ICAM-1 under non-clustered and clustered conditions.<h4>Principal findings</h4>Detailed scanning electron and fluorescent microscopy showed that the apical surface of endothelial cells constitutively forms small filopodia-like protrusions that are positive for ICAM-1 and freely move within the lateral plane of the membrane. Clustering of ICAM-1, using anti-ICAM-1 antibody-coated beads, efficiently and rapidly recruits ICAM-1. Using fluorescence recovery after photo-bleaching (FRAP), we found that clustering increased the immobile fraction of ICAM-1, compared to non-clustered ICAM-1. This shift required the intracellular portion of ICAM-1. Moreover, biochemical assays showed that ICAM-1 clustering recruited beta-actin and filamin. Cytochalasin B, which interferes with actin polymerization, delayed the clustering of ICAM-1. In addition, we could show that cytochalasin B decreased the immobile fraction of clustered ICAM-1-GFP, but had no effect on non-clustered ICAM-1. Also, the motor protein myosin-II is recruited to ICAM-1 adhesion sites and its inhibition increased the immobile fraction of both non-clustered and clustered ICAM-1. Finally, blocking Rac1 activation, the formation of lipid rafts, myosin-II activity or actin polymerization, but not Src, reduced the adhesive function of ICAM-1, tested under physiological flow conditions.<h4>Conclusions</h4>Together, these findings indicate that ICAM-1 clustering is regulated in an inside-out fashion through the actin cytoskeleton. Overall, these data indicate that signaling events within the endothelium are required for efficient ICAM-1-mediated leukocyte adhesion.Jaap D van BuulJos van RijsselFloris P J van AlphenMark HoogenboezemSimon TolKees A HoebenJan van MarleErik P J MulPeter L HordijkPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 5, Iss 6, p e11336 (2010)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Jaap D van Buul Jos van Rijssel Floris P J van Alphen Mark Hoogenboezem Simon Tol Kees A Hoeben Jan van Marle Erik P J Mul Peter L Hordijk Inside-out regulation of ICAM-1 dynamics in TNF-alpha-activated endothelium.
description	<h4>Background</h4>During transendothelial migration, leukocytes use adhesion molecules, such as ICAM-1, to adhere to the endothelium. ICAM-1 is a dynamic molecule that is localized in the apical membrane of the endothelium and clusters upon binding to leukocytes. However, not much is known about the regulation of ICAM-1 clustering and whether membrane dynamics are linked to the ability of ICAM-1 to cluster and bind leukocyte integrins. Therefore, we studied the dynamics of endothelial ICAM-1 under non-clustered and clustered conditions.<h4>Principal findings</h4>Detailed scanning electron and fluorescent microscopy showed that the apical surface of endothelial cells constitutively forms small filopodia-like protrusions that are positive for ICAM-1 and freely move within the lateral plane of the membrane. Clustering of ICAM-1, using anti-ICAM-1 antibody-coated beads, efficiently and rapidly recruits ICAM-1. Using fluorescence recovery after photo-bleaching (FRAP), we found that clustering increased the immobile fraction of ICAM-1, compared to non-clustered ICAM-1. This shift required the intracellular portion of ICAM-1. Moreover, biochemical assays showed that ICAM-1 clustering recruited beta-actin and filamin. Cytochalasin B, which interferes with actin polymerization, delayed the clustering of ICAM-1. In addition, we could show that cytochalasin B decreased the immobile fraction of clustered ICAM-1-GFP, but had no effect on non-clustered ICAM-1. Also, the motor protein myosin-II is recruited to ICAM-1 adhesion sites and its inhibition increased the immobile fraction of both non-clustered and clustered ICAM-1. Finally, blocking Rac1 activation, the formation of lipid rafts, myosin-II activity or actin polymerization, but not Src, reduced the adhesive function of ICAM-1, tested under physiological flow conditions.<h4>Conclusions</h4>Together, these findings indicate that ICAM-1 clustering is regulated in an inside-out fashion through the actin cytoskeleton. Overall, these data indicate that signaling events within the endothelium are required for efficient ICAM-1-mediated leukocyte adhesion.
format	article
author	Jaap D van Buul Jos van Rijssel Floris P J van Alphen Mark Hoogenboezem Simon Tol Kees A Hoeben Jan van Marle Erik P J Mul Peter L Hordijk
author_facet	Jaap D van Buul Jos van Rijssel Floris P J van Alphen Mark Hoogenboezem Simon Tol Kees A Hoeben Jan van Marle Erik P J Mul Peter L Hordijk
author_sort	Jaap D van Buul
title	Inside-out regulation of ICAM-1 dynamics in TNF-alpha-activated endothelium.
title_short	Inside-out regulation of ICAM-1 dynamics in TNF-alpha-activated endothelium.
title_full	Inside-out regulation of ICAM-1 dynamics in TNF-alpha-activated endothelium.
title_fullStr	Inside-out regulation of ICAM-1 dynamics in TNF-alpha-activated endothelium.
title_full_unstemmed	Inside-out regulation of ICAM-1 dynamics in TNF-alpha-activated endothelium.
title_sort	inside-out regulation of icam-1 dynamics in tnf-alpha-activated endothelium.
publisher	Public Library of Science (PLoS)
publishDate	2010
url	https://doaj.org/article/7193af2812dc4e36a4aa181fd4b28836
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_version_	1718374169934036992

Inside-out regulation of ICAM-1 dynamics in TNF-alpha-activated endothelium.

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