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:

RhoH is required to maintain the integrin LFA-1 in a nonadhesive state on lymphocytes

Nature Immunology volume 5, pages 961–967 (2004)Cite this article

Abstract

Lymphocyte function–associated antigen 1 (LFA-1) is relatively nonadhesive on resting lymphocytes; however, the mechanisms underlying changes in its adhesiveness are poorly understood. In this study, we generated a Jurkat T cell clone, J+hi1.14, that contained low amounts of mRNA for RhoH, a leukocyte-specific inhibitory Rho family member. J+hi1.14 cells expressed constitutively adhesive LFA-1 and the cells bound spontaneously to intracellular adhesion molecules 1, 2 and 3. Reconstitution of RhoH mRNA expression in J+hi1.14 cells reverted the adhesion phenotype to that of wild-type. We obtained similar results using RNA interference in peripheral blood lymphocytes. These data demonstrate that RhoH is required for maintenance of lymphocyte LFA-1 in a nonadhesive state.

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: Isolation of constitutively adherent Jurkat clones.
Figure 2: One of the two provirus insertions in Jurkat clone J+hi1.14 is in RHOH.
Figure 3: Blot hybridization analysis of Jurkat clone RNA.
Figure 4: The adhesion of J+hi1.14 cells to ICAM-1–Fc is dependent on LFA-1.
Figure 5: An LFA-1-deficient clone derived from J+hi1.14 fails to bind to ICAM-1.
Figure 6: The adhesion phenotype of J+hi1.14 is reverted by cell fusion.
Figure 7: Transfection with RhoH cDNA reverts the J+hi1.14 phenotype to that of wild-type cells.
Figure 8: RhoH RNA interference in human peripheral blood T cells leads to a constitutively adherent phenotype.

Similar content being viewed by others

Accession codes

Accessions

GenBank/EMBL/DDBJ

References

  1. Springer, T.A. Adhesion receptors of the immune system. Nature 346, 425–434 (1990).

    Article  CAS  PubMed  Google Scholar 

  2. Marlin, S.D. & Springer, T.A. Purified intercellular adhesion molecule-1 (ICAM-1) is a ligand for lymphocyte function-associated antigen 1 (LFA-1). Cell 5, 813–819 (1987).

    Article  Google Scholar 

  3. Staunton, D.E., Dustin, M.L. & Springer, T.A. Functional cloning of ICAM-2, a cell adhesion ligand for LFA-1 homologous to ICAM-1. Nature 339, 61–64 (1989).

    Article  CAS  PubMed  Google Scholar 

  4. de Fougerolles, A.R., Stacker, S.A., Schwarting, R. & Springer, T.A. Characterization of ICAM-2 and evidence for a third counter-receptor for LFA-1. J. Exp. Med. 174, 253–267 (1991).

    Article  CAS  PubMed  Google Scholar 

  5. Ostermann, G., Weber, K.S., Zernecke, A., Schroder, A. & Weber, C. JAM-1 is a ligand of the β2 integrin LFA-1 involved in transendothelial migration of leukocytes. Nat. Immunol. 3, 151–158 (2002).

    Article  CAS  PubMed  Google Scholar 

  6. Tagaki, J. & Springer, T.A. Integrin activation and structural rearrangement. Immunol. Rev. 186, 141–163 (2002).

    Article  Google Scholar 

  7. Hogg, N., Henderson, R., Leitinger, B., Porter, A. & Stanley, P. Mechanisms contributing to the activity of integrins on leukocytes. Immunol. Rev. 186, 164–171 (2002).

    Article  CAS  PubMed  Google Scholar 

  8. Liddington, R.C. & Ginsberg, M.H. Integrin activation takes shape. J. Cell Biol. 158, 833–839 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  9. Arnaout, M.A. Integrin structure: new twists and turns in dynamic cell adhesion. Immunol. Rev. 186, 125–140 (2002).

    Article  CAS  PubMed  Google Scholar 

  10. Hynes, R.O. Integrins: Bidirectional, allosteric signaling machines. Cell 110, 673–687 (2002).

    Article  CAS  PubMed  Google Scholar 

  11. Rothlein, R. & Springer, T.A. The requirement for lymphocyte function-associated antigen 1 in homotypic leukocyte adhesion stimulated by phorbol ester. J. Exp. Med. 163, 1132–1149 (1986).

    Article  CAS  PubMed  Google Scholar 

  12. Chatila, T.A., Geha, R.S. & Arnaout, M.A. Constitutive and stimulus-induced phosphorylation of CD11/CD18 leukocyte adhesion molecules. J. Cell. Biol. 10, 3435–3444 (1989).

    Article  Google Scholar 

  13. van Kooyk, Y., Weder, P., Heije, K. & Figdor, C.G. Extracellular Ca2+ modulates leukocyte function-associated antigen-1 cell surface distribution on T lymphocytes and consequently affects cell adhesion. J. Cell Biol. 124, 1061–1070 (1994).

    Article  CAS  PubMed  Google Scholar 

  14. Dustin, M.L. & Springer, T.A. T-cell receptor cross-linking transiently stimulates adhesiveness through LFA-1. Nature 341, 619–624 (1989).

    Article  CAS  PubMed  Google Scholar 

  15. Szabo, M.C., Butcher, E.C., McIntyre, B.W., Schall, T.J. & Bacon, K.B. RANTES stimulation of T lymphocyte adhesion and activation: role for LFA-1 and ICAM-3. Eur. J. Immunol. 27, 1061–1068 (1997).

    Article  CAS  PubMed  Google Scholar 

  16. Weber, K.S., Klickstein, L.B., Weber, P.C. & Weber, C. Chemokine-induced monocyte transmigration requires cdc42-mediated cytoskeletal changes. Eur. J. Immunol. 28, 2245–2251 (1998).

    Article  CAS  PubMed  Google Scholar 

  17. Larson, R.S., Hibbs, M.L. & Springer, T.A. The leukocyte integrin LFA-1 reconstituted by cDNA transfection in a nonhematopoietic cell line is functionally active and not transiently regulated. Cell Regul. 1, 359–367 (1990).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Johnston, S.C., Dustin, M.L., Hibbs, M.L. & Springer, T.A. On the species specificity of the interaction of LFA-1 with intercellular adhesion molecules. J. Immunol. 145, 1181–1187 (1990).

    CAS  PubMed  Google Scholar 

  19. O'Toole, T.E. et al. Integrin cytoplasmic domains mediate inside-out signal transduction. J. Cell Biol. 124, 1047–1059 (1994).

    Article  CAS  PubMed  Google Scholar 

  20. Weber, C., Alon, R., Moser, B. & Springer, T.A. Sequential regulation of α4β1 and α5β1 integrin avidity by CC chemokines in monocytes: implications for transendothelial chemotaxis. J. Cell Biol. 134, 1063–1073 (1996).

    Article  CAS  PubMed  Google Scholar 

  21. Li, X. et al. The hematopoiesis-specific GTP-binding protein RhoH is GTPase deficient and modulates activities of other Rho GTPases by an inhibitory function. Mol. Cell. Biol. 22, 1158–1171 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Dallery, E. et al. TTF, a gene encoding a novel small G protein, fuses to the lymphoma-associated LAZ3 gene by t(3,4) chromosomal translocation. Oncogene. 10, 2171–2178 (1995).

    CAS  PubMed  Google Scholar 

  23. Cherry, L.K., Weber, K.S.C. & Klickstein, L.B. A dominant Jurkat T cell mutation that inhibits LFA-1-mediated cell adhesion is associated with increased cell growth. J. Immunol. 167, 6171–6179 (2001).

    Article  CAS  PubMed  Google Scholar 

  24. Rubinson, D.A. et al. A lentivirus-based system to functionally silence genes in primary mammalian cells, stem cells and transgenic mice by RNA interference. Nat. Genet. 33, 401–406 (2003).

    Article  CAS  PubMed  Google Scholar 

  25. Weber, K.S.C., York, M.R., Springer, T.A. & Klickstein, L.B. Characterization of lymphocyte function-associated antigen 1 (LFA-1) deficient T cell lines. The αL and β2 subunits are interdependent for cell surface expression. J. Immunol. 158, 273–279 (1997).

    CAS  PubMed  Google Scholar 

  26. Liu, L., Schwartz, B.R., Lin, N., Winn, R.K. & Harlan, J.M. Requirement for Rho A kinase activation in leukocyte de-adhesion. J. Immunol. 169, 2330–2336 (2002).

    Article  CAS  PubMed  Google Scholar 

  27. 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 

  28. Weber, K.S. et al. Dual role of H-Ras in regulation of lymphocyte function antigen-1 activity by stromal cell-derived factor-1α: implications for leukocyte transmigration. Mol. Biol. Cell. 12, 3074–3086 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Geiger, C. et al. Cytohesin-1 regulates β-2 integrin-mediated adhesion through both ARF-GEF function and interaction with LFA-1. EMBO J. 19, 2525–2536 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Liu, L. et al. The GTPase Rap1 regulates phorbol 12-myristate 13-acetate-stimulated but not ligand-induced β1 integrin-dependent leukocyte adhesion. J. Biol. Chem. 277, 40893–40900 (2002).

    Article  CAS  PubMed  Google Scholar 

  31. Sebzda, E., Bracke, M., Tugal, T., Hogg, N. & Cantrell, D.A. Rap1A positively regulates T cells via integrin activation rather than inhibiting lymphocyte signaling. Nat. Immunol. 3, 251–258 (2002).

    Article  CAS  PubMed  Google Scholar 

  32. 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 

  33. Chang, W., Hubbard, S.C., Friedel, C. & Ruley, H.E. Enrichment of insertional mutants following retrovirus gene trap selection. Virology 193, 737–747 (1993).

    Article  CAS  PubMed  Google Scholar 

  34. Soriano, P., Cone, R.D., Mulligan, R.C. & Jaenisch, R. Tissue-specific and ectopic expression of genes introduced into trangenic mice by retroviruses. Science 234, 1409–1413 (1986).

    Article  CAS  PubMed  Google Scholar 

  35. Markowitz, D., Goff, S. & Bank, A. Construction and use of a safe and efficient amphotropic packaging cell line. Virology 167, 400–406 (1988).

    Article  CAS  PubMed  Google Scholar 

  36. Siebert, P.D., Chenchik, A., Kellogg, D.E., Lukyanov, K.A. & Lukyanov, S.A. An improved PCR method for walking in uncloned genomic DNA. Nucleic Acids Res. 23, 1087–1088 (1995).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Lennon, G., Auffray, C., Polymeropoulos, M. & Soares, M.B. The I.M.A.G.E. consortium: an integrated molecular analysis of genomes and their expression. Genomics. 33, 15–52 (1996).

    Article  Google Scholar 

  38. Otabor, I. et al. A role for lipid rafts in C1q-triggered O2− generation by human neutrophils. Mol. Immunol. 41, 185–190 (2004).

    Article  CAS  PubMed  Google Scholar 

  39. Changelian, P.S., Jack, R.M., Collins, L.A. & Fearon, D.T. PMA induces the ligand-independent internalization of CR1 on human neutrophils. J. Immunol. 134, 1851–1858 (1985).

    CAS  PubMed  Google Scholar 

  40. Petruzzelli, L., Maduzia, L. & Springer, T.A. Activation of lymphocyte function-associated molecule-1 (CD11a/CD18) and Mac-1 (CD11b/CD18) mimicked by an antibody directed against CD18. J. Immunol. 155, 854–866 (1995).

    CAS  PubMed  Google Scholar 

  41. Rothlein, R., Dustin, M.L., Marlin, S.D. & Springer, T.A. A human intercellular adhesion molecule (ICAM-1) distinct from LFA-1. J. Immunol. 137, 1270–1274 (1986).

    CAS  PubMed  Google Scholar 

  42. Landis, R.C., Bennett, R.I. & Hogg, N. A novel LFA-1 activation epitope maps to the I domain. J. Cell Biol. 120, 1519–1527 (1993).

    Article  CAS  PubMed  Google Scholar 

  43. Andrew, D. et al. KIM185 a monoclonal antibody to CD18 which induces a change in the conformation of CD18 and promotes both LFA-1- and CR3-dependent adhesion. Eur. J. Immunol. 23, 2217–2222 (1993).

    Article  CAS  PubMed  Google Scholar 

  44. Kozak, M. Point mutations define a sequence flanking the AUG initiator codon that modulates translation by eukaryotic ribosomes. Cell 44, 283–292 (1986).

    Article  CAS  PubMed  Google Scholar 

  45. Ory, D.S., Neugeboren, B.A. & Mulligan, R.C. A stable human-derived packaging cell line for production of high titer retrovirus/vesicular stomatitis virus G pseudotypes. Proc. Natl. Acad. Sci. USA 93, 11400–11406 (1996).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Southern, E.M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J. Mol. Biol. 98, 503–517 (1975).

    Article  CAS  PubMed  Google Scholar 

  47. Thomas, P.S. Hybridization of denatured RNA and small DNA fragments transferred to nitrocellulose. Proc. Natl. Acad. Sci. USA 77, 5201–5205 (1980).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We thank X. Bu for technical assistance. Supported by National Institutes of Health (F32AR08632 to L.K.C., R01DK047636 to B.L. and R01AR47243 to L.B.K.), the Leukemia and Lymphoma Society (B.L.), the Lymphoma Research Foundation (X.L.) and the Arthritis Foundation (P.S.).

Author information

Author notes

  1. Lisa K Cherry

    Present address: Genzyme Corporation, 1 Mountain Road, Framington, Massachusetts, 01701, USA

  2. Bing Lim

    Present address: Genome Institute of Singapore, 60 Biopolis Street, Singapore, 138672

Authors and Affiliations

  1. Division of Rheumatology, Department of Medicine, Immunology and Allergy, Brigham & Women's Hospital, Smith Building room 650, 1 Jimmy Fund Way, Boston, 02115, Massachusetts, USA

    Lisa K Cherry, Pascale Schwab & Lloyd B Klickstein

  2. Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Institutes of Medicine, 77 Avenue Louis Pasteur, Boston, 02215, Massachusetts, USA

    Xiaoyu Li & Bing Lim

Authors
  1. Lisa K Cherry
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Xiaoyu Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Pascale Schwab
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Bing Lim
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Lloyd B Klickstein
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Lloyd B Klickstein.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Cherry, L., Li, X., Schwab, P. et al. RhoH is required to maintain the integrin LFA-1 in a nonadhesive state on lymphocytes. Nat Immunol 5, 961–967 (2004). https://doi.org/10.1038/ni1103

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/ni1103

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