Expression of the phagocytic receptors αMβ2 and αXβ2 is controlled by RIAM, VASP and Vinculin in neutrophildifferentiated HL-60 cells

ctivation of the integrin phagocytic receptors CR3 (αMβ2, CD11b/CD18) and CR4 (αXβ2, CD11c/CD18) requires Rap1 activation and RIAM function. RIAM controls integrin activation by recruiting Talin to β2 subunits, enabling the Talin- Vinculin interaction, which in term bridges integrins to the actin-cy...

Descripción completa

Detalles Bibliográficos
Autores: Torres-Gomez, Alvaro, Fiyouzi, Tara, Guerra-Espinosa, Claudia, Cardeñes, Beatriz, Clares, Irene, Toribio Serrano, Víctor, Reche, Pedro A, Cabañas, Carlos, Lafuente, Esther M.
Tipo de recurso: artículo
Fecha de publicación:2022
País:España
Institución:Universidad Autónoma de Madrid
Repositorio:Biblos-e Archivo. Repositorio Institucional de la UAM
Idioma:inglés
OAI Identifier:oai:repositorio.uam.es:10486/719220
Acceso en línea:http://hdl.handle.net/10486/719220
https://dx.doi.org/10.3389/fimmu.2022.951280
Access Level:acceso abierto
Palabra clave:Phagocytosis
cytoskeleton
VASP
integrin expression
CR3 (CD11b/CD18)
Vinculin
CR4 (CD11c/CD18)
RIAM
Biología y Biomedicina / Biología
id ES_3edbaf54173a9853cdf2ac7b8aee6bff
oai_identifier_str oai:repositorio.uam.es:10486/719220
network_acronym_str ES
network_name_str España
repository_id_str
spelling Expression of the phagocytic receptors αMβ2 and αXβ2 is controlled by RIAM, VASP and Vinculin in neutrophildifferentiated HL-60 cellsTorres-Gomez, AlvaroFiyouzi, TaraGuerra-Espinosa, ClaudiaCardeñes, BeatrizClares, IreneToribio Serrano, VíctorReche, Pedro ACabañas, CarlosLafuente, Esther M.PhagocytosiscytoskeletonVASPintegrin expressionCR3 (CD11b/CD18)VinculinCR4 (CD11c/CD18)RIAMBiología y Biomedicina / Biologíactivation of the integrin phagocytic receptors CR3 (αMβ2, CD11b/CD18) and CR4 (αXβ2, CD11c/CD18) requires Rap1 activation and RIAM function. RIAM controls integrin activation by recruiting Talin to β2 subunits, enabling the Talin- Vinculin interaction, which in term bridges integrins to the actin-cytoskeleton. RIAM also recruits VASP to phagocytic cups and facilitates VASP phosphorylation and function promoting particle internalization. Using a CRISPR-Cas9 knockout approach, we have analyzed the requirement for RIAM, VASP and Vinculin expression in neutrophilic-HL-60 cells. All knockout cells displayed abolished phagocytosis that was accompanied by a significant and specific reduction in ITGAM (aM), ITGAX (aX) and ITGB2 (β2) mRNA, as revealed by RT-qPCR. RIAM, VASP and Vinculin KOs presented reduced cellular F-actin content that correlated with aM expression, as treatment with the actin filament polymerizing and stabilizing drug jasplakinolide, partially restored aM expression. In general, the expression of aX was less responsive to jasplakinolide treatment than aM, indicating that regulatory mechanisms independent of F-actin content may be involved. The Serum Response Factor (SRF) was investigated as the potential transcription factor controlling αMβ2 expression, since its coactivator MRTF-A requires actin polymerization to induce transcription. Immunofluorescent MRTF-A localization in parental cells was primarily nuclear, while in knockouts it exhibited a diffuse cytoplasmic pattern. Localization of FHL-2 (SRF corepressor) was mainly sub-membranous in parental HL-60 cells, but in knockouts the localization was disperse in the cytoplasm and the nucleus, suggesting RIAM, VASP and Vinculin are required to maintain FHL-2 close to cytoplasmic membranes, reducing its nuclear localization and inhibiting its corepressor activity. Finally, reexpression of VASP in the VASP knockout resulted in a complete reversion of the phenotype, as knock-ins restored aM expression. Taken together, our results suggest that RIAM, VASP and Vinculin, are necessary for the correct expression of αMβ2 and αXβ2 during neutrophilic differentiation in the human promyelocytic HL-60 cell line, and strongly point to an involvement of these proteins in the acquisition of a phagocytic phenotypeEconomía y Competitividad (MINECO) grants: SAF2016- 77096-R and PID2021-123199OB-I00 (E.L. and C.C.), the CAM (Comunidad Autónoma de Madrid) research agency through grant IND2020/BMD-17364 (P.A.R. and T.F.), and fellowship from MINECO to A. T-G. (FPU15/05349)Frontiers Media SADepartamento de Biología MolecularFacultad de Ciencias20222022-09-27research articlehttp://purl.org/coar/resource_type/c_2df8fbb1VoRhttp://purl.org/coar/version/c_970fb48d4fbd8a85info:eu-repo/semantics/articleapplication/mswordapplication/pdfhttp://hdl.handle.net/10486/719220https://dx.doi.org/10.3389/fimmu.2022.951280reponame:Biblos-e Archivo. Repositorio Institucional de la UAMinstname:Universidad Autónoma de MadridInglésengopen accesshttp://purl.org/coar/access_right/c_abf2Attribution 4.0 Internationalhttp://creativecommons.org/licenses/by/4.0/info:eu-repo/semantics/openAccessoai:repositorio.uam.es:10486/7192202026-06-23T12:46:27Z
dc.title.none.fl_str_mv Expression of the phagocytic receptors αMβ2 and αXβ2 is controlled by RIAM, VASP and Vinculin in neutrophildifferentiated HL-60 cells
title Expression of the phagocytic receptors αMβ2 and αXβ2 is controlled by RIAM, VASP and Vinculin in neutrophildifferentiated HL-60 cells
spellingShingle Expression of the phagocytic receptors αMβ2 and αXβ2 is controlled by RIAM, VASP and Vinculin in neutrophildifferentiated HL-60 cells
Torres-Gomez, Alvaro
Phagocytosis
cytoskeleton
VASP
integrin expression
CR3 (CD11b/CD18)
Vinculin
CR4 (CD11c/CD18)
RIAM
Biología y Biomedicina / Biología
title_short Expression of the phagocytic receptors αMβ2 and αXβ2 is controlled by RIAM, VASP and Vinculin in neutrophildifferentiated HL-60 cells
title_full Expression of the phagocytic receptors αMβ2 and αXβ2 is controlled by RIAM, VASP and Vinculin in neutrophildifferentiated HL-60 cells
title_fullStr Expression of the phagocytic receptors αMβ2 and αXβ2 is controlled by RIAM, VASP and Vinculin in neutrophildifferentiated HL-60 cells
title_full_unstemmed Expression of the phagocytic receptors αMβ2 and αXβ2 is controlled by RIAM, VASP and Vinculin in neutrophildifferentiated HL-60 cells
title_sort Expression of the phagocytic receptors αMβ2 and αXβ2 is controlled by RIAM, VASP and Vinculin in neutrophildifferentiated HL-60 cells
dc.creator.none.fl_str_mv Torres-Gomez, Alvaro
Fiyouzi, Tara
Guerra-Espinosa, Claudia
Cardeñes, Beatriz
Clares, Irene
Toribio Serrano, Víctor
Reche, Pedro A
Cabañas, Carlos
Lafuente, Esther M.
author Torres-Gomez, Alvaro
author_facet Torres-Gomez, Alvaro
Fiyouzi, Tara
Guerra-Espinosa, Claudia
Cardeñes, Beatriz
Clares, Irene
Toribio Serrano, Víctor
Reche, Pedro A
Cabañas, Carlos
Lafuente, Esther M.
author_role author
author2 Fiyouzi, Tara
Guerra-Espinosa, Claudia
Cardeñes, Beatriz
Clares, Irene
Toribio Serrano, Víctor
Reche, Pedro A
Cabañas, Carlos
Lafuente, Esther M.
author2_role author
author
author
author
author
author
author
author
dc.contributor.none.fl_str_mv Departamento de Biología Molecular
Facultad de Ciencias
dc.subject.none.fl_str_mv Phagocytosis
cytoskeleton
VASP
integrin expression
CR3 (CD11b/CD18)
Vinculin
CR4 (CD11c/CD18)
RIAM
Biología y Biomedicina / Biología
topic Phagocytosis
cytoskeleton
VASP
integrin expression
CR3 (CD11b/CD18)
Vinculin
CR4 (CD11c/CD18)
RIAM
Biología y Biomedicina / Biología
description ctivation of the integrin phagocytic receptors CR3 (αMβ2, CD11b/CD18) and CR4 (αXβ2, CD11c/CD18) requires Rap1 activation and RIAM function. RIAM controls integrin activation by recruiting Talin to β2 subunits, enabling the Talin- Vinculin interaction, which in term bridges integrins to the actin-cytoskeleton. RIAM also recruits VASP to phagocytic cups and facilitates VASP phosphorylation and function promoting particle internalization. Using a CRISPR-Cas9 knockout approach, we have analyzed the requirement for RIAM, VASP and Vinculin expression in neutrophilic-HL-60 cells. All knockout cells displayed abolished phagocytosis that was accompanied by a significant and specific reduction in ITGAM (aM), ITGAX (aX) and ITGB2 (β2) mRNA, as revealed by RT-qPCR. RIAM, VASP and Vinculin KOs presented reduced cellular F-actin content that correlated with aM expression, as treatment with the actin filament polymerizing and stabilizing drug jasplakinolide, partially restored aM expression. In general, the expression of aX was less responsive to jasplakinolide treatment than aM, indicating that regulatory mechanisms independent of F-actin content may be involved. The Serum Response Factor (SRF) was investigated as the potential transcription factor controlling αMβ2 expression, since its coactivator MRTF-A requires actin polymerization to induce transcription. Immunofluorescent MRTF-A localization in parental cells was primarily nuclear, while in knockouts it exhibited a diffuse cytoplasmic pattern. Localization of FHL-2 (SRF corepressor) was mainly sub-membranous in parental HL-60 cells, but in knockouts the localization was disperse in the cytoplasm and the nucleus, suggesting RIAM, VASP and Vinculin are required to maintain FHL-2 close to cytoplasmic membranes, reducing its nuclear localization and inhibiting its corepressor activity. Finally, reexpression of VASP in the VASP knockout resulted in a complete reversion of the phenotype, as knock-ins restored aM expression. Taken together, our results suggest that RIAM, VASP and Vinculin, are necessary for the correct expression of αMβ2 and αXβ2 during neutrophilic differentiation in the human promyelocytic HL-60 cell line, and strongly point to an involvement of these proteins in the acquisition of a phagocytic phenotype
publishDate 2022
dc.date.none.fl_str_mv 2022
2022-09-27
dc.type.none.fl_str_mv research article
http://purl.org/coar/resource_type/c_2df8fbb1
VoR
http://purl.org/coar/version/c_970fb48d4fbd8a85
dc.type.openaire.fl_str_mv info:eu-repo/semantics/article
format article
dc.identifier.none.fl_str_mv http://hdl.handle.net/10486/719220
https://dx.doi.org/10.3389/fimmu.2022.951280
url http://hdl.handle.net/10486/719220
https://dx.doi.org/10.3389/fimmu.2022.951280
dc.language.none.fl_str_mv Inglés
eng
language_invalid_str_mv Inglés
language eng
dc.rights.none.fl_str_mv open access
http://purl.org/coar/access_right/c_abf2
Attribution 4.0 International
http://creativecommons.org/licenses/by/4.0/
dc.rights.openaire.fl_str_mv info:eu-repo/semantics/openAccess
rights_invalid_str_mv open access
http://purl.org/coar/access_right/c_abf2
Attribution 4.0 International
http://creativecommons.org/licenses/by/4.0/
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/msword
application/pdf
dc.publisher.none.fl_str_mv Frontiers Media SA
publisher.none.fl_str_mv Frontiers Media SA
dc.source.none.fl_str_mv reponame:Biblos-e Archivo. Repositorio Institucional de la UAM
instname:Universidad Autónoma de Madrid
instname_str Universidad Autónoma de Madrid
reponame_str Biblos-e Archivo. Repositorio Institucional de la UAM
collection Biblos-e Archivo. Repositorio Institucional de la UAM
repository.name.fl_str_mv
repository.mail.fl_str_mv
_version_ 1869406587882307584
score 15,198674

Similares en LA Referencia