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Presentation of integrins on leukocyte microvilli: a role for the extracellular domain in determining membrane localization.

Abitorabi MA, Pachynski RK, Ferrando RE, Tidswell M, Erle DJ - J. Cell Biol. (1997)

Bottom Line: Therefore, differences between the transmembrane and cytoplasmic domains of alpha4 and beta2 integrins do not account for differences in ability to support attachment under flow or in membrane localization.Integrins alpha4beta1, alpha5beta1, alpha6Abeta1, alphavbeta3, and alphaEbeta7 also localized to microvilli.Transmembrane proteins known or suspected to associate with extracellular domains of microvillous integrins, including tetraspans and CD47, were concentrated on microvilli as well.

View Article: PubMed Central - PubMed

Affiliation: The Lung Biology Center, Department of Medicine, University of California, San Francisco, California 94143, USA.

ABSTRACT
Adhesion of blood leukocytes to the endothelium involves multiple steps including initial attachment (tethering), rolling, and firm arrest. Presentation of adhesion molecules on leukocyte microvilli can substantially enhance tethering. Localization of L-selectin to microvilli and of CD44 to the planar cell body have been shown to depend upon their transmembrane and cytoplasmic domains. We investigated the role of leukocyte integrin transmembrane and cytoplasmic domains in initiating adhesion under flow and in microvillous localization. Integrins alpha4beta7, alphaLbeta2, and alphaMbeta2 were heterologously expressed in K562 cells. alpha4beta7 initiated adhesion under flow and localized to microvilli, whereas beta2 integrins did not initiate adhesion and localized to the cell body. Chimeric integrins were produced by replacing the alpha4beta7 cytoplasmic and/or transmembrane domains with the homologous domains of alphaLbeta2 or alphaMbeta2. Unexpectedly, these chimeras efficiently mediated adhesion to the alpha4beta7 ligand mucosal addressin cell adhesion molecule-1 under flow and localized to microvilli. Therefore, differences between the transmembrane and cytoplasmic domains of alpha4 and beta2 integrins do not account for differences in ability to support attachment under flow or in membrane localization. Integrins alpha4beta1, alpha5beta1, alpha6Abeta1, alphavbeta3, and alphaEbeta7 also localized to microvilli. Transmembrane proteins known or suspected to associate with extracellular domains of microvillous integrins, including tetraspans and CD47, were concentrated on microvilli as well. These findings suggest that interactions between the extracellular domains of integrins and associated proteins could direct the assembly of multimolecular complexes on leukocyte microvilli.

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Schematic representation of wild-type and chimeric  α4β7 integrins. The amino acid sequence at the splice site is shown  with conserved regions in bold, and the αLβ2 or αMβ2 sequences  underlined.
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Figure 1: Schematic representation of wild-type and chimeric α4β7 integrins. The amino acid sequence at the splice site is shown with conserved regions in bold, and the αLβ2 or αMβ2 sequences underlined.

Mentions: The pCDM8-integrin α4 cDNA plasmid (Kamata et al., 1995) was a gift from Y. Takada (Scripps Research Institute, La Jolla, CA). The cloning of the β7 cDNA has been previously described (Erle et al., 1991). Integrin β2 cDNA (Hickstein et al., 1988) was provided by D. Hickstein (University of Washington, Seattle, WA). Integrin αL (Larson et al., 1989) and αM (Corbi et al., 1988) cDNAs were provided by T. Springer (Center for Blood Research, Boston, MA). αE cDNA (Shaw et al., 1994) was a gift from G. Russell and M. Brenner (Harvard Medical School, Boston, MA). Chimeric integrin subunits were constructed using splice overlap extension PCR (Horton et al., 1989). The amino acid splice sites for each construct are shown in Fig. 1. Chimeric α subunits were subcloned into pCDM8 (Invitrogen, San Diego, CA) and chimeric β subunits were subcloned into pCEP4 (Invitrogen). The integrity of the constructs was confirmed by DNA sequencing.


Presentation of integrins on leukocyte microvilli: a role for the extracellular domain in determining membrane localization.

Abitorabi MA, Pachynski RK, Ferrando RE, Tidswell M, Erle DJ - J. Cell Biol. (1997)

Schematic representation of wild-type and chimeric  α4β7 integrins. The amino acid sequence at the splice site is shown  with conserved regions in bold, and the αLβ2 or αMβ2 sequences  underlined.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2139798&req=5

Figure 1: Schematic representation of wild-type and chimeric α4β7 integrins. The amino acid sequence at the splice site is shown with conserved regions in bold, and the αLβ2 or αMβ2 sequences underlined.
Mentions: The pCDM8-integrin α4 cDNA plasmid (Kamata et al., 1995) was a gift from Y. Takada (Scripps Research Institute, La Jolla, CA). The cloning of the β7 cDNA has been previously described (Erle et al., 1991). Integrin β2 cDNA (Hickstein et al., 1988) was provided by D. Hickstein (University of Washington, Seattle, WA). Integrin αL (Larson et al., 1989) and αM (Corbi et al., 1988) cDNAs were provided by T. Springer (Center for Blood Research, Boston, MA). αE cDNA (Shaw et al., 1994) was a gift from G. Russell and M. Brenner (Harvard Medical School, Boston, MA). Chimeric integrin subunits were constructed using splice overlap extension PCR (Horton et al., 1989). The amino acid splice sites for each construct are shown in Fig. 1. Chimeric α subunits were subcloned into pCDM8 (Invitrogen, San Diego, CA) and chimeric β subunits were subcloned into pCEP4 (Invitrogen). The integrity of the constructs was confirmed by DNA sequencing.

Bottom Line: Therefore, differences between the transmembrane and cytoplasmic domains of alpha4 and beta2 integrins do not account for differences in ability to support attachment under flow or in membrane localization.Integrins alpha4beta1, alpha5beta1, alpha6Abeta1, alphavbeta3, and alphaEbeta7 also localized to microvilli.Transmembrane proteins known or suspected to associate with extracellular domains of microvillous integrins, including tetraspans and CD47, were concentrated on microvilli as well.

View Article: PubMed Central - PubMed

Affiliation: The Lung Biology Center, Department of Medicine, University of California, San Francisco, California 94143, USA.

ABSTRACT
Adhesion of blood leukocytes to the endothelium involves multiple steps including initial attachment (tethering), rolling, and firm arrest. Presentation of adhesion molecules on leukocyte microvilli can substantially enhance tethering. Localization of L-selectin to microvilli and of CD44 to the planar cell body have been shown to depend upon their transmembrane and cytoplasmic domains. We investigated the role of leukocyte integrin transmembrane and cytoplasmic domains in initiating adhesion under flow and in microvillous localization. Integrins alpha4beta7, alphaLbeta2, and alphaMbeta2 were heterologously expressed in K562 cells. alpha4beta7 initiated adhesion under flow and localized to microvilli, whereas beta2 integrins did not initiate adhesion and localized to the cell body. Chimeric integrins were produced by replacing the alpha4beta7 cytoplasmic and/or transmembrane domains with the homologous domains of alphaLbeta2 or alphaMbeta2. Unexpectedly, these chimeras efficiently mediated adhesion to the alpha4beta7 ligand mucosal addressin cell adhesion molecule-1 under flow and localized to microvilli. Therefore, differences between the transmembrane and cytoplasmic domains of alpha4 and beta2 integrins do not account for differences in ability to support attachment under flow or in membrane localization. Integrins alpha4beta1, alpha5beta1, alpha6Abeta1, alphavbeta3, and alphaEbeta7 also localized to microvilli. Transmembrane proteins known or suspected to associate with extracellular domains of microvillous integrins, including tetraspans and CD47, were concentrated on microvilli as well. These findings suggest that interactions between the extracellular domains of integrins and associated proteins could direct the assembly of multimolecular complexes on leukocyte microvilli.

Show MeSH
Related in: MedlinePlus