Limits...
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.

Show MeSH

Related in: MedlinePlus

Localization of integrin-associated proteins by  immunoelectron microscopy.  The tetraspan proteins CD53  (A) and CD63 (B), the αvβ3-associated protein CD47 (C),  and CD32 (D) were localized  by immunoelectron microscopy using 12-nm (A, C, and  D) or 6-nm (B) gold particles  (arrows). The tetraspan proteins and CD47 localized primarily to microvilli, whereas  CD32 was found in substantial amounts on both the cell  body and microvilli. Nontransfected K562 cells (A)  and K562-α4β7 (B), K562-αVβ3 (C), and K562-αMβ2  (D) transfectants were used  for staining. Bar, 0.5 μm.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2139798&req=5

Figure 7: Localization of integrin-associated proteins by immunoelectron microscopy. The tetraspan proteins CD53 (A) and CD63 (B), the αvβ3-associated protein CD47 (C), and CD32 (D) were localized by immunoelectron microscopy using 12-nm (A, C, and D) or 6-nm (B) gold particles (arrows). The tetraspan proteins and CD47 localized primarily to microvilli, whereas CD32 was found in substantial amounts on both the cell body and microvilli. Nontransfected K562 cells (A) and K562-α4β7 (B), K562-αVβ3 (C), and K562-αMβ2 (D) transfectants were used for staining. Bar, 0.5 μm.

Mentions: Integrins have been shown to associate with a variety of intracellular and transmembrane proteins. Some of these interactions occur within the cell and are mediated by integrin cytoplasmic domains, whereas others are known or suspected to be extracellular. Since our results suggested that extracellular (and not transmembrane or cytoplasmic) domains determine integrin localization, we performed immunoelectron microscopy to localize several cell surface proteins known or suspected to interact with integrin extracellular domains. Several members of the tetraspan family of transmembrane proteins have been shown to associate with α4β1, α4β7, α6β1, and some other integrins (Berditchevski et al., 1996; Mannion et al., 1996). These associations are likely to involve the extracellular domains of tetraspans and integrins (see Discussion). Several tetraspans are constitutively expressed on K562 cells. CD53 (82 ± 5% on microvilli, Fig. 7 A), CD63 (92 ± 4% on microvilli, Fig. 7 B), and CD81 and CD82 (data not shown) are all localized predominantly to microvilli. Another transmembrane protein, CD47 (integrin-associated protein), has been shown to associate with αvβ3 via its extracellular domain. CD47, like αvβ3, was distributed mostly on microvilli (Fig. 7 C). CD32 is a transmembrane protein that has been reported to associate with αMβ2 (which is on the cell body; Fig. 5 C) and possibly with the tetraspan CD82 (found on microvilli, see above) (Lebel-Binay et al., 1995). CD32 was expressed on both microvilli (62 ± 15%) and the cell body (38 ± 15%) (Fig. 7 D).


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)

Localization of integrin-associated proteins by  immunoelectron microscopy.  The tetraspan proteins CD53  (A) and CD63 (B), the αvβ3-associated protein CD47 (C),  and CD32 (D) were localized  by immunoelectron microscopy using 12-nm (A, C, and  D) or 6-nm (B) gold particles  (arrows). The tetraspan proteins and CD47 localized primarily to microvilli, whereas  CD32 was found in substantial amounts on both the cell  body and microvilli. Nontransfected K562 cells (A)  and K562-α4β7 (B), K562-αVβ3 (C), and K562-αMβ2  (D) transfectants were used  for staining. Bar, 0.5 μm.
© Copyright Policy
Related In: Results  -  Collection

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

Figure 7: Localization of integrin-associated proteins by immunoelectron microscopy. The tetraspan proteins CD53 (A) and CD63 (B), the αvβ3-associated protein CD47 (C), and CD32 (D) were localized by immunoelectron microscopy using 12-nm (A, C, and D) or 6-nm (B) gold particles (arrows). The tetraspan proteins and CD47 localized primarily to microvilli, whereas CD32 was found in substantial amounts on both the cell body and microvilli. Nontransfected K562 cells (A) and K562-α4β7 (B), K562-αVβ3 (C), and K562-αMβ2 (D) transfectants were used for staining. Bar, 0.5 μm.
Mentions: Integrins have been shown to associate with a variety of intracellular and transmembrane proteins. Some of these interactions occur within the cell and are mediated by integrin cytoplasmic domains, whereas others are known or suspected to be extracellular. Since our results suggested that extracellular (and not transmembrane or cytoplasmic) domains determine integrin localization, we performed immunoelectron microscopy to localize several cell surface proteins known or suspected to interact with integrin extracellular domains. Several members of the tetraspan family of transmembrane proteins have been shown to associate with α4β1, α4β7, α6β1, and some other integrins (Berditchevski et al., 1996; Mannion et al., 1996). These associations are likely to involve the extracellular domains of tetraspans and integrins (see Discussion). Several tetraspans are constitutively expressed on K562 cells. CD53 (82 ± 5% on microvilli, Fig. 7 A), CD63 (92 ± 4% on microvilli, Fig. 7 B), and CD81 and CD82 (data not shown) are all localized predominantly to microvilli. Another transmembrane protein, CD47 (integrin-associated protein), has been shown to associate with αvβ3 via its extracellular domain. CD47, like αvβ3, was distributed mostly on microvilli (Fig. 7 C). CD32 is a transmembrane protein that has been reported to associate with αMβ2 (which is on the cell body; Fig. 5 C) and possibly with the tetraspan CD82 (found on microvilli, see above) (Lebel-Binay et al., 1995). CD32 was expressed on both microvilli (62 ± 15%) and the cell body (38 ± 15%) (Fig. 7 D).

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