Korean Journal of Agricultural Science (농업과학연구)

Institute of Agricultural Science, CNU (충남대학교 농업과학연구소)
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IQGAP1, a signaling scaffold protein, as a molecular target of a small molecule inhibitor to interfere with T cell receptor-mediated integrin activation

  • Li, Lin-Ying (College of Pharmacy and Pharmaceutical Science, Chungnam National University) ;
  • Nguyen, Thi Minh Nguyet (College of Pharmacy and Pharmaceutical Science, Chungnam National University) ;
  • Woo, Eui Jeon (Korea Research Institute of Bioscience and Biotechnology) ;
  • Park, Jongtae (Department of Food Science and Technology, College of Agriculture, Chungnam National University) ;
  • Hwang, Inkyu (College of Pharmacy and Pharmaceutical Science, Chungnam National University)
  • Received : 2020.03.16
  • Accepted : 2020.05.20
  • Published : 2020.06.01
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Abstract

Integrins such as lymphocyte function-associated antigen -1 (LFA-1) have an essential role in T cell immunity. Integrin activation, namely, the transition from the inactive conformation to the active one, takes place when an intracellular signal is generated by specific receptors such as T cell receptors (TCRs) and chemokine receptors in T cells. In an effort to explore the molecular mechanisms underlying the TCR-mediated LFA-1 activation, we had previously established a high-throughput cell-based assay and screened a chemical library deposited in the National Institute of Health in the United States. As a result, several hits had been isolated including HIKS-1 (Benzo[b]thiophene-3-carboxylic acid, 2-[3-[(2-carboxyphenyl) thio]-2,5-dioxo-1-pyrrolinyl]-4,5,6,7-tetrahydro-,3-ethyl ester). In an attempt to reveal the mode of action of HIKS-1, in this study, we did drug affinity responsive target stability (DARTS) assay finding that HIKS-1 interacted with the IQ motif containing GTPase activating protein 1 (IQGAP1), a 189 kDa multidomain scaffold protein critically involved in various signaling mechanisms. Furthermore, the cellular thermal shift assay (CETSA) provided compelling evidence that HIKS-1 also interacted with IQGAP1 in vivo. Taken together, it can be concluded that HIKS-1 interferes with the TCR-mediated LFA-1 activation by interacting with IQGAP1 and thereby disrupting the signaling pathway for LFA-1 activation.

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References

  1. Abram CL, Lowell CA. 2009. The ins and outs of leukocyte integrin signaling. Annual Review of Immunology 27:339-362. https://doi.org/10.1146/annurev.immunol.021908.132554
  2. An WF, Tolliday NJ. 2009. Introduction: Cell-based assays for high-throughput screening. Methods in Molecular Biology 486:1-12. https://doi.org/10.1007/978-1-60327-545-3_1
  3. Bahk YY, Kim SA, Kim JS, Euh HJ, Bai GH, Cho SN, Kim YS. 2004. Antigens secreted from Mycobacterium tuberculosis: identification by proteomics approach and test for diagnostic marker. Proteomics 4:3299-3307. https://doi.org/10.1002/pmic.200400980
  4. Choi S, Thapa N, Hedman AC, Li Z, Sacks DB, Anderson RA. 2013. IQGAP1 is a novel phosphatidylinositol 4,5 bisphosphate effector in regulation of directional cell migration. The Embo Journal 32:2617-2630. https://doi.org/10.1038/emboj.2013.191
  5. Ding M, Kaspersson K, Murray D, Bardelle C. 2017. High-throughput flow cytometry for drug discovery: Principles, applications, and case studies. Drug Discovery Today 22:1844-1850. https://doi.org/10.1016/j.drudis.2017.09.005
  6. Freyer MW, Lewis EA. 2008. Isothermal titration calorimetry: Experimental design, data analysis, and probing macromolecule/ligand binding and kinetic interactions. Methods in Cell Biology 84:79-113. https://doi.org/10.1016/S0091-679X(07)84004-0
  7. Gobom J, Nordhoff E, Mirgorodskaya E, Ekman R, Roepstorff P. 1999. Sample purification and preparation technique based on nano-scale reversed-phase columns for the sensitive analysis of complex peptide mixtures by matrixassisted laser desorption/ionization mass spectrometry. Journal of Mass Spectrometry 34:105-116. https://doi.org/10.1002/(SICI)1096-9888(199902)34:2<105::AID-JMS768>3.0.CO;2-4
  8. Hedman AC, Smith JM, Sacks DB. 2015. The biology of IQGAP proteins: Beyond the cytoskeleton. EMBO Reports 16:427-446. https://doi.org/10.15252/embr.201439834
  9. Hinman SS, McKeating KS, Cheng Q. 2018. Surface plasmon resonance: Material and interface design for universal accessibility 90:19-39. https://doi.org/10.1021/acs.analchem.7b04251
  10. Hogg N, Patzak I, Willenbrock F. 2011. The insider's guide to leukocyte integrin signalling and function. Nature Reviews Immunology 11:416-426. https://doi.org/10.1038/nri2986
  11. Hwang IHM, Hu C, Waller A, Carter M, Wu Y, Sklar L. 2009. Summary report of screening for developing T cell immune modulators. Accessed in on 10 June 2019.
  12. Hwang I, Kim K, Choi S, Lomunova M. 2017. Potentiation of T cell stimulatory activity by chemical fixation of a weak peptide-MHC complex. Molecules and Cells 40:24-36. https://doi.org/10.14348/molcells.2017.2218
  13. Jacquemet G, Morgan MR, Byron A, Humphries JD, Choi CK, Chen CS, Caswell PT, Humphries MJ. 2013. Rac1 is deactivated at integrin activation sites through an IQGAP1-filamin-A-RacGAP1 pathway. Journal of Cell Science 126:4121-4135. https://doi.org/10.1242/jcs.121988
  14. Jafari R, Almqvist H, Axelsson H, Ignatushchenko M, Lundback T, Nordlund P, Martinez Molina D. 2014. The cellular thermal shift assay for evaluating drug target interactions in cells. Nature Protocols 9:2100-2122. https://doi.org/10.1038/nprot.2014.138
  15. Kim K, Wang L, Hwang I. 2009a. Acute inhibition of selected membrane-proximal mouse T cell receptor signaling by mitochondrial antagonists. PLoS One 4:e7738. https://doi.org/10.1371/journal.pone.0007738
  16. Kim K, Wang L, Hwang I. 2009b. A novel flow cytometric high throughput assay for a systematic study on molecular mechanisms underlying T cell receptor-mediated integrin activation. PLoS One 4:e6044. https://doi.org/10.1371/journal.pone.0006044
  17. Lomenick B, Hao R, Jonai N, Chin RM, Aghajan M, Warburton S, Wang J, Wu RP, Gomez F, Loo JA, Wohlschlegel JA, Vondriska TM, Pelletier J, Herschman HR, Clardy J, Clarke CF, Huang J. 2009. Target identification using drug affinity responsive target stability (DARTS). Proceedings of National Academy of Sciences of USA 106:21984-21989. https://doi.org/10.1073/pnas.0910040106
  18. Mack NA, Georgiou M. 2014. The interdependence of the Rho GTPases and apicobasal cell polarity. Small GTPases 5:10.
  19. Martinez Molina D, Jafari R, Ignatushchenko M, Seki T, Larsson EA, Dan C, Sreekumar L, Cao Y, Nordlund P. 2013. Monitoring drug target engagement in cells and tissues using the cellular thermal shift assay. Science 341:84-87. https://doi.org/10.1126/science.1233606
  20. Martinez Molina D, Nordlund P. 2016. The cellular thermal shift assay: A novel biophysical assay for in situ drug target engagement and mechanistic biomarker studies. Annual Review of Pharmacology and Toxicology 56:141-161. https://doi.org/10.1146/annurev-pharmtox-010715-103715
  21. O' Connor CJ, Laraia L, Spring DR. 2011. Chemical genetics. Chemical Society Reviews 40:4332-4345. https://doi.org/10.1039/c1cs15053g
  22. Pai MY, Lomenick B, Hwang H, Schiestl R, McBride W, Loo JA, Huang J. 2015. Drug affinity responsive target stability (DARTS) for small-molecule target identification. Methods in Molecules Biology 1263:287-298. https://doi.org/10.1007/978-1-4939-2269-7_22
  23. Pathmanathan S, Hamilton E, Atcheson E, Timson DJ. 2011. The interaction of IQGAPs with calmodulin-like proteins. Biochemical Society Transactions 39:694-699. https://doi.org/10.1042/BST0390694
  24. Pelikan-Conchaudron A, Le Clainche C, Didry D, Carlier MF. 2011. The IQGAP1 protein is a calmodulin-regulated barbed end capper of actin filaments: Possible implications in its function in cell migration. Journal of Biological Chemistry 286:35119-35128. https://doi.org/10.1074/jbc.M111.258772
  25. Schenone M, Dancik V, Wagner BK, Clemons PA. 2013. Target identification and mechanism of action in chemical biology and drug discovery. Nature Chemical Biology 9:232-240. https://doi.org/10.1038/nchembio.1199
  26. Sha WC, Nelson CA, Newberry RD, Kranz DM, Russell JH, Loh DY. 1988. Selective expression of an antigen receptor on CD8-bearing T lymphocytes in transgenic mice. Nature 335:271-274. https://doi.org/10.1038/335271a0
  27. Sjoblom B, Ylanne J, Djinovic-Carugo K. 2008. Novel structural insights into F-actin-binding and novel functions of calponin homology domains. Current Opinion Structural Biology 18:702-708. https://doi.org/10.1016/j.sbi.2008.10.003
  28. Smith JM, Hedman AC, Sacks DB. 2015. IQGAPs choreograph cellular signaling from the membrane to the nucleus. Trends in Cell Biology 25:171-184. https://doi.org/10.1016/j.tcb.2014.12.005
  29. Verma NK, Kelleher D. 2017. Not just an adhesion molecule: LFA-1 contact tunes the T lymphocyte program. The Journal of Immunology 199:1213-1221. https://doi.org/10.4049/jimmunol.1700495
  30. Walling BL, Kim M. 2018. LFA-1 in T cell migration and differentiation. Frontiers in Immunology 9:952. https://doi.org/10.3389/fimmu.2018.00952
  31. Ziegler S, Pries V, Hedberg C, Waldmann H. 2013. Target identification for small bioactive molecules: Finding the needle in the haystack. Angewandte Chemie International Edition England 52:2744-2792. https://doi.org/10.1002/anie.201208749