Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Regulation of conformer-specific activation of the integrin LFA-1 by a chemokine-triggered Rho signaling module

Nature Immunology volume 10, pages 185–194 (2009)Cite this article

Abstract

Regulation of the affinity of the β2 integrin LFA-1 by chemokines is critical to lymphocyte trafficking, but the signaling mechanisms that control this process are not well understood. Here we investigated the signaling events controlling LFA-1 affinity triggering by chemokines in human primary T lymphocytes. We found that the small GTPase Rac1 mediated chemokine-induced LFA-1 affinity triggering and lymphocyte arrest in high endothelial venules. Unexpectedly, another Rho family member, Cdc42, negatively regulated LFA-1 activation. The Rho effectors PLD1 and PIP5KC were also critical to LFA-1 affinity modulation. Notably, PIP5KC was found to specifically control the transition of LFA-1 from an extended low–intermediate state to a high-affinity state, which correlated with lymphocyte arrest. Thus, chemokines control lymphocyte trafficking by triggering a Rho-dependent signaling cascade leading to conformer-specific modulation of LFA-1 affinity

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Rac1 mediates chemokine-induced LFA-1 affinity triggering and dependent adhesion in T lymphocytes.
Figure 2: Cdc42 is a negative regulator of chemokine-induced LFA-1 affinity triggering and dependent adhesion in T lymphocytes.
Figure 3: PLD1 activity mediates chemokine-induced LFA-1 affinity triggering and dependent adhesion in T lymphocytes.
Figure 4: RhoA-activated PLD1 mediates chemokine-induced LFA-1 affinity triggering and dependent adhesion in T lymphocyte.
Figure 5: PIP5KC selectively controls chemokine-induced LFA-1 high-affinity state triggering and mediated adhesion in T lymphocytes.
Figure 6: Regulation of PIP5KC activity by Rho small GTPases and PLD1.
Figure 7: Rac1, Cdc42, PLD1 and PIP5KC control human T lymphocyte in vivo arrest on HEVs.

Similar content being viewed by others

References

  1. Butcher, E.C. Leukocyte-endothelial cell recognition: three (or more) steps to specificity and diversity. Cell 67, 1033–1036 (1991).

    Article  CAS  PubMed  Google Scholar 

  2. Alon, R. & Dustin, M.L. Force as a facilitator of integrin conformational changes during leukocyte arrest on blood vessels and antigen-presenting cells. Immunity 26, 17–27 (2007).

    Article  CAS  PubMed  Google Scholar 

  3. Ley, K., Laudanna, C., Cybulsky, M.I. & Nourshargh, S. Getting to the site of inflammation: the leukocyte adhesion cascade updated. Nat. Rev. Immunol. 7, 678–689 (2007).

    Article  CAS  PubMed  Google Scholar 

  4. Laudanna, C. & Alon, R. Right on the spot. Chemokine triggering of integrin-mediated arrest of rolling leukocytes. Thromb. Haemost. 95, 5–11 (2006).

    Article  CAS  PubMed  Google Scholar 

  5. Kinashi, T. Intracellular signalling controlling integrin activation in lymphocytes. Nat. Rev. Immunol. 5, 546–559 (2005).

    Article  CAS  PubMed  Google Scholar 

  6. Carman, C.V. & Springer, T.A. Integrin avidity regulation: are changes in affinity and conformation underemphasized? Curr. Opin. Cell Biol. 15, 547–556 (2003).

    Article  CAS  PubMed  Google Scholar 

  7. Constantin, G. et al. Chemokines trigger immediate beta2 integrin affinity and mobility changes: differential regulation and roles in lymphocyte arrest under flow. Immunity 13, 759–769 (2000).

    Article  CAS  PubMed  Google Scholar 

  8. Cairo, C.W., Mirchev, R. & Golan, D.E. Cytoskeletal regulation couples LFA-1 conformational changes to receptor lateral mobility and clustering. Immunity 25, 297–308 (2006).

    Article  CAS  PubMed  Google Scholar 

  9. Giagulli, C. et al. RhoA and zeta PKC control distinct modalities of LFA-1 activation by chemokines: critical role of LFA-1 affinity triggering in lymphocyte in vivo homing. Immunity 20, 25–35 (2004).

    Article  CAS  PubMed  Google Scholar 

  10. Kim, M., Carman, C.V., Yang, W., Salas, A. & Springer, T.A. The primacy of affinity over clustering in regulation of adhesiveness of the integrin αLβ2. J. Cell Biol. 167, 1241–1253 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Shamri, R. et al. Lymphocyte arrest requires instantaneous induction of an extended LFA-1 conformation mediated by endothelium-bound chemokines. Nat. Immunol. 6, 497–506 (2005).

    Article  CAS  PubMed  Google Scholar 

  12. Giagulli, C. et al. The Src family kinases Hck and Fgr are dispensable for inside-out, chemoattractant-induced signaling regulating beta 2 integrin affinity and valency in neutrophils, but are required for beta 2 integrin-mediated outside-in signaling involved in sustained adhesion. J. Immunol. 177, 604–611 (2006).

    Article  CAS  PubMed  Google Scholar 

  13. D'Ambrosio, D. et al. Quantitative differences in chemokine receptor engagement generate diversity in integrin-dependent lymphocyte adhesion. J. Immunol. 169, 2303–2312 (2002).

    Article  CAS  PubMed  Google Scholar 

  14. Pasvolsky, R. et al. RhoA is involved in LFA-1 extension triggered by CXCL12 but not in a novel outside-in LFA-1 activation facilitated by CXCL9. J. Immunol. 180, 2815–2823 (2008).

    Article  CAS  PubMed  Google Scholar 

  15. Shimonaka, M. et al. Rap1 translates chemokine signals to integrin activation, cell polarization, and motility across vascular endothelium under flow. J. Cell Biol. 161, 417–427 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Katagiri, K., Maeda, A., Shimonaka, M. & Kinashi, T. RAPL, a Rap1-binding molecule that mediates Rap1-induced adhesion through spatial regulation of LFA-1. Nat. Immunol. 4, 741–748 (2003).

    Article  CAS  PubMed  Google Scholar 

  17. Prochiantz, A. Homeodomain-derived peptides. In and out of the cells. Ann. NY Acad. Sci. 886, 172–179 (1999).

    Article  CAS  PubMed  Google Scholar 

  18. Zhao, X., Carnevale, K.A. & Cathcart, M.K. Human monocytes use Rac1, not Rac2, in the NADPH oxidase complex. J. Biol. Chem. 278, 40788–40792 (2003).

    Article  CAS  PubMed  Google Scholar 

  19. Hall, A. Rho GTPases and the actin cytoskeleton. Science 279, 509–514 (1998).

    Article  CAS  PubMed  Google Scholar 

  20. Zigmond, S.H. et al. Mechanism of Cdc42-induced actin polymerization in neutrophil extracts. J. Cell Biol. 142, 1001–1012 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Powner, D.J. & Wakelam, M.J. The regulation of phospholipase D by inositol phospholipids and small GTPases. FEBS Lett. 531, 62–64 (2002).

    Article  CAS  PubMed  Google Scholar 

  22. Henage, L.G., Exton, J.H. & Brown, H.A. Kinetic analysis of a mammalian phospholipase D: allosteric modulation by monomeric GTPases, protein kinase C, and polyphosphoinositides. J. Biol. Chem. 281, 3408–3417 (2006).

    Article  CAS  PubMed  Google Scholar 

  23. Bae, C.D., Min, D.S., Fleming, I.N. & Exton, J.H. Determination of interaction sites on the small G protein RhoA for phospholipase D. J. Biol. Chem. 273, 11596–11604 (1998).

    Article  CAS  PubMed  Google Scholar 

  24. Brown, H.A., Henage, L.G., Preininger, A.M., Xiang, Y. & Exton, J.H. Biochemical analysis of phospholipase D. Methods Enzymol. 434, 49–87 (2007).

    Article  CAS  PubMed  Google Scholar 

  25. Yamazaki, M. et al. Interaction of the small G protein RhoA with the C terminus of human phospholipase D1. J. Biol. Chem. 274, 6035–6038 (1999).

    Article  CAS  PubMed  Google Scholar 

  26. Cai, S. & Exton, J.H. Determination of interaction sites of phospholipase D1 for RhoA. Biochem. J. 355, 779–785 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Weernink, P.A. et al. Activation of type I phosphatidylinositol 4-phosphate 5-kinase isoforms by the Rho GTPases, RhoA, Rac1, and Cdc42. J. Biol. Chem. 279, 7840–7849 (2004).

    Article  CAS  PubMed  Google Scholar 

  28. Jarquin-Pardo, M., Fitzpatrick, A., Galiano, F.J., First, E.A. & Davis, J.N. Phosphatidic acid regulates the affinity of the murine phosphatidylinositol 4-phosphate 5-kinase-Ibeta for phosphatidylinositol-4-phosphate. J. Cell. Biochem. 100, 112–128 (2007).

    Article  CAS  PubMed  Google Scholar 

  29. Kanaho, Y., Kobayashi-Nakano, A. & Yokozeki, T. The phosphoinositide kinase PIP5K that produces the versatile signaling phospholipid PI4,5P(2). Biol. Pharm. Bull. 30, 1605–1609 (2007).

    Article  CAS  PubMed  Google Scholar 

  30. Martel, V. et al. Conformation, localization, and integrin binding of talin depend on its interaction with phosphoinositides. J. Biol. Chem. 276, 21217–21227 (2001).

    Article  CAS  PubMed  Google Scholar 

  31. Wegener, K.L. et al. Structural basis of integrin activation by talin. Cell 128, 171–182 (2007).

    Article  CAS  PubMed  Google Scholar 

  32. Ling, K., Doughman, R.L., Firestone, A.J., Bunce, M.W. & Anderson, R.A. Type I gamma phosphatidylinositol phosphate kinase targets and regulates focal adhesions. Nature 420, 89–93 (2002).

    Article  CAS  PubMed  Google Scholar 

  33. Di Paolo, G. et al. Recruitment and regulation of phosphatidylinositol phosphate kinase type 1 gamma by the FERM domain of talin. Nature 420, 85–89 (2002).

    Article  CAS  PubMed  Google Scholar 

  34. Barsukov, I.L. et al. Phosphatidylinositol phosphate kinase type 1gamma and beta1-integrin cytoplasmic domain bind to the same region in the talin FERM domain. J. Biol. Chem. 278, 31202–31209 (2003).

    Article  CAS  PubMed  Google Scholar 

  35. Battistini, L. et al. CD8+ T cells from patients with acute multiple sclerosis display selective increase of adhesiveness in brain venules: a critical role for P-selectin glycoprotein ligand-1. Blood 101, 4775–4782 (2003).

    Article  CAS  PubMed  Google Scholar 

  36. Piccio, L. et al. Efficient recruitment of lymphocytes in inflamed brain venules requires expression of cutaneous lymphocyte antigen and fucosyltransferase-VII. J. Immunol. 174, 5805–5813 (2005).

    Article  CAS  PubMed  Google Scholar 

  37. Garcia-Bernal, D. et al. Vav1 and Rac control chemokine-promoted T lymphocyte adhesion mediated by the integrin alpha4beta1. Mol. Biol. Cell 16, 3223–3235 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Colley, W.C. et al. Phospholipase D2, a distinct phospholipase D isoform with novel regulatory properties that provokes cytoskeletal reorganization. Curr. Biol. 7, 191–201 (1997).

    Article  CAS  PubMed  Google Scholar 

  39. Sampath, R., Gallagher, P.J. & Pavalko, F.M. Cytoskeletal interactions with the leukocyte integrin beta2 cytoplasmic tail. Activation-dependent regulation of associations with talin and alpha-actinin. J. Biol. Chem. 273, 33588–33594 (1998).

    Article  CAS  PubMed  Google Scholar 

  40. Stanley, P. et al. Intermediate-affinity LFA-1 binds alpha-actinin-1 to control migration at the leading edge of the T cell. EMBO J. 27, 62–75 (2008).

    Article  CAS  PubMed  Google Scholar 

  41. Iyer, S.S. & Kusner, D.J. Association of phospholipase D activity with the detergent-insoluble cytoskeleton of U937 promonocytic leukocytes. J. Biol. Chem. 274, 2350–2359 (1999).

    Article  CAS  PubMed  Google Scholar 

  42. Kam, Y. & Exton, J.H. Phospholipase D activity is required for actin stress fiber formation in fibroblasts. Mol. Cell. Biol. 21, 4055–4066 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Isenberg, G. & Goldmann, W.H. Peptide-specific antibodies localize the major lipid binding sites of talin dimers to oppositely arranged N-terminal 47 kDa subdomains. FEBS Lett. 426, 165–170 (1998).

    Article  CAS  PubMed  Google Scholar 

  44. Woolf, E. et al. Lymph node chemokines promote sustained T lymphocyte motility without triggering stable integrin adhesiveness in the absence of shear forces. Nat. Immunol. 8, 1076–1085 (2007).

    Article  CAS  PubMed  Google Scholar 

  45. Zarbock, A., Lowell, C.A. & Ley, K. Spleen tyrosine kinase Syk is necessary for E-selectin-induced alpha(L)beta(2) integrin-mediated rolling on intercellular adhesion molecule-1. Immunity 26, 773–783 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Hartwell, L.H., Hopfield, J.J., Leibler, S. & Murray, A.W. From molecular to modular cell biology. Nature 402, C47–C52 (1999).

    Article  CAS  PubMed  Google Scholar 

  47. Bhalla, U.S. & Iyengar, R. Functional modules in biological signalling networks. Novartis Found. Symp. 239, 4–13 discussion 13–5, 45–51 (2001).

    CAS  PubMed  Google Scholar 

  48. Pereira-Leal, J.B., Levy, E.D. & Teichmann, S.A. The origins and evolution of functional modules: lessons from protein complexes. Phil. Trans. R. Soc. Lond. B 361, 507–517 (2006).

    Article  CAS  Google Scholar 

  49. Segal, E., Friedman, N., Kaminski, N., Regev, A. & Koller, D. From signatures to models: understanding cancer using microarrays. Nat. Genet. 37 Suppl, S38–S45 (2005).

    Article  CAS  PubMed  Google Scholar 

  50. Robinson, M.K. et al. Antibody against the Leu-CAM beta-chain (CD18) promotes both LFA-1- and CR3-dependent adhesion events. J. Immunol. 148, 1080–1085 (1992).

    CAS  PubMed  Google Scholar 

  51. Shimaoka, M. et al. AL-57, a ligand-mimetic antibody to integrin LFA-1, reveals chemokine-induced affinity up-regulation in lymphocytes. Proc. Natl. Acad. Sci. USA 103, 13991–13996 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Lum, A.F., Green, C.E., Lee, G.R., Staunton, D.E. & Simon, S.I. Dynamic regulation of LFA-1 activation and neutrophil arrest on intercellular adhesion molecule 1 (ICAM-1) in shear flow. J. Biol. Chem. 277, 20660–20670 (2002).

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

Supported by the Italian Association for Cancer Research, the Italian Ministry of University and Scientific Research, Fondazione Cariverona (Verona, Italy), the National Multiple Sclerosis Society of New York and Fondazione Italiana Sclerosi Multipla.

Author information

Authors and Affiliations

  1. Department of Pathology, Division of General Pathology, University of Verona, School of Medicine, Verona, 37134, Italy

    Matteo Bolomini-Vittori, Alessio Montresor, Cinzia Giagulli, Barbara Rossi, Marianna Martinello, Gabriela Constantin & Carlo Laudanna

  2. The Center for Biomedical Computing, University of Verona, Verona, 37134, Italy

    Alessio Montresor, Cinzia Giagulli & Carlo Laudanna

  3. ICOS, Bothell, 98021, Washington, USA

    Donald Staunton

Authors
  1. Matteo Bolomini-Vittori
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Alessio Montresor
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Cinzia Giagulli
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Donald Staunton
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Barbara Rossi
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Marianna Martinello
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Gabriela Constantin
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Carlo Laudanna
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

M.B.-V. developed the Tat technology, set up the siRNA technology and did the adhesion assays; A.M. set up the siRNA technology and did adhesion assays and biochemical studies, ImageStream experiments and analysis; C.G. developed the P1 technology; D.S. provided monoclonal antibody 327C; B.R. and M.M. did the intravital microscopy studies; G.C. provided expertise in intravital microscopy and assistance with writing; and C.L. designed the study, analyzed the data and wrote the paper.

Corresponding author

Correspondence to Carlo Laudanna.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–6 and Supplementary Methods (PDF 718 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Bolomini-Vittori, M., Montresor, A., Giagulli, C. et al. Regulation of conformer-specific activation of the integrin LFA-1 by a chemokine-triggered Rho signaling module. Nat Immunol 10, 185–194 (2009). https://doi.org/10.1038/ni.1691

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/ni.1691

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing