G Protein Coupled Receptors-Mediated Chemotaxis in the Model Organism Dictyostelium Discoideum and Neutrophils

Xuehua Xu*
Chemotaxis Signaling Section, Laboratory of Immunogenetics, NIH/NIAID, USA

*Corresponding author: Xuehua Xu, Chemotaxis Signaling Section, Laboratory of Immunogenetics, NIH/NIAID, 12441 Parklawn Drive, Rockville, MD 20852, USA

Published: 23 Feb, 2017
Cite this article as: Xu X. G Protein Coupled Receptors- Mediated Chemotaxis in the Model Organism Dictyostelium Discoideum and Neutrophils. Ann Infect Dis Epidemiol. 2017; 2(1): 1011.


Chemotaxis, a directional cell migration guided by extracellular chemoattractant gradients, plays critical roles in many physiological processes, recruitment of neutrophils to sites of inflammation, metastasis of cancer cells, and development of model organism Dictyostelium discoideum [1-4]. Both D. discoideum and mammalian neutrophils sense chemoattractants using G protein-coupled receptors (GPCRs) and share remarkable similarities in the signaling pathways by which regulate cell migration Jin et al. [5]. It has been proven that D. discoideum is a powerful model organism to establish new concepts and identify new components essential for chemotaxis [6,7]. We developed and applied state-of-the-art live cell/single molecule imaging technologies to visualize spatiotemporal dynamics of GPCR-mediated signaling network in D. discoideum [8,9]. We also built up chemosensing signaling network for computational modeling [10]. By interplaying computational simulation and experimental verification (Figure1), we revealed locally-controlled inhibitory mechanism upstream of PI3K that is essential for chemosensing [9,11,12]. Recently, we are focusing on identifying the inhibitors, such as negative regulators of Ras, for chemosensing in both D. discoideum and mammalian neutrophils. Our long-term goal is to identify novel components and signaling pathways essential for chemotaxis to provide new therapeutic targets and strategies for inflammatory diseases and metastasis of cancer.


All eukaryotic cells sense chemoattractants by G protein-coupled receptors (GPCRs) and share remarkable similarities in the signaling pathways which mediate chemotaxis. The knowledge of GPCR-mediated signaling pathways leading to chemotaxis are mostly from D. discoideum and neutrophils. In both system, the binding of chemoattractants to their receptors induces the dissociation of heterotrimeric G-proteins into Gα and Gβγ subunits, which, in turn, activate multiple pivotal effectors, such as small GTPase Ras, phosphatidylinositol (PtdIn)-3 kinases (PI3K), and phospholipase C (PLC) [13-21]. PI3K phosphorylates membrane phospholipid PIP2 to PIP3. The generated PIP3 mediates intracellularly polarized localization and activation of the proteins with Pleckstrin Homology (PH) domains, such as cytosolic regulator of adenylyl (CRAC), protein kinase B (PKB), and myosin I proteins (actin motors) [13,14,17,18,20,22]. PIP3-indepdedent pathways involving PLA2 and cGC have also been implicated in D. discoideum chemotaxis [23,24]. All together, they control the reorganization of the actin-myosin cytoskeleton for a directed cell migration [25- 30]. The key feature of chemotaxis in both D. discoideum and neutrophils is that cells sense a large range of chemoattractant concentrations (about 10-9 to 10-5 M) by applying a mechanism called adaptation [31,32]. Adaptive cell no longer responds to the present stimuli but still remains sensitive for higher concentration stimuli Hoeller et al. [31]. GPCR-mediated adaptive behavior occurs in many steps of GPCR-mediated signaling pathway of chemotaxis, such as activation of PLCβ2/3 and Ras [18,21,33-37] PI3K/PTEN-mediated transient PIP3 production [15,17,18,37,38] Rho/Rac activation and actin polymerization [39-41] indicating that adaptation is an fundamental strategy cell apply to chemotax across huge range of chemoattractant concentrations. To explain adaptation (Figure 2), different models agree upon the temporal dynamics of adaptation: an increase in receptor occupancy activates two antagonistic signaling processes: a rapid “excitation” that triggers cellular responses and a temporally delayed “inhibition” that terminates the responses to reach adaptation [2,6,12,26,42,31]. Many excitatory components has been identified, however, the inhibitors and their function in chemosensing and directed cell migration are still largely elusive.
We developed and applied the state-of-art live cell imaging techniques to monitor spatiotemporal dynamics of GPCR-mediated signaling events in live single cells. In both D. discoideum and neutrophils, gradient sensing is able to be uncoupled with initial polarization and cell migration by actin polymerization inhibitor such as latrunculin [2,28]. Nonpolarized immobile cells, hence, provides a simplified cell system for simultaneous monitoring multiple signaling events at subcellular level in real time [8]. We systematically measured the binding of cAMP binding to its receptor cAR1 by visualizing single receptor using total internal reflection (TIRF) microscopy [11], cAR1- induced dissociation of heterotrimeric G protein using Fluorescence Resonance Energy Transfer (FRET) technique [9], the dynamic membrane translocation of PI3K, PTEN, and PHcrac, a biosensor for PIP3 [11,9,12]. The spatiotemporal dynamics of signaling components have provided perimeters for computational simulation. Combining these dynamics with computational simulation led to a better understanding of GPCR-signaling network at a system level. Our findings suggest that a locally-regulated inhibitory mechanism, downstream of heterotrimeric G protein but upstream or at PI3K/ PTEN, is essential for gradient sensing. We postulate two types of inhibitors for chemosensing: 1) negative regulators for Ras signaling; 2) components regulates the redistribution of PTEN dynamics. Recently, we focus on understanding the molecular mechanism of Ras adaption and PTEN membrane localization.

Figure 1

Another alt text

Figure 1
The interplay between computational stimulation and experimental vitrification proposes a locally regulated inhibitory process, which is required for GPCR-mediated chemosensing.

Figure 2

Another alt text

Figure 2
Two major inhibitory branches of chemosensing.


  1. Bravo-Cordero JJ, Hodgson L, Condeelis J. Directed cell invasion and migration during metastasis. Curr Opin Cell Biol. 2012; 24: 277-283.
  2. Parent CA, Devreotes PN. A cell's sense of direction. Science. 1999;284: 765-770.
  3. Wen Z, Zheng, JQ. Directional guidance of nerve growth cones. Curr Opin Neurobiol. 2006;16: 52-58.
  4. Zigmond SH. Chemotaxis by polymorphonuclear leukocytes. J Cell Biol. 1978; 77: 269-287.
  5. Jin T, Xu X, Fang J, Isik N, Yan J, Brzostowski JA, et al. How human leukocytes track down and destroy pathogens: lessons learned from the model organism Dictyostelium discoideum. Immunol Res. 2009; 43: 118-127.
  6. Devreotes P, Janetopoulos C. Eukaryotic chemotaxis: distinctions between directional sensing and polarization. J Biol Chem. 2003; 278: 20445-20448.
  7. Jin T, Xu X, Hereld D. Chemotaxis, chemokine receptors and human disease. Cytokine. 2008; 44: 1-8.
  8. Xu X, Jin T. Imaging G-protein coupled receptor (GPCR)-mediated signaling events that control chemotaxis of Dictyostelium discoideum. J Vis Exp. 2011.
  9. Xu X, Meier-Schellersheim M, Jiao X, Nelson LE, Jin T. Quantitative imaging of single live cells reveals spatiotemporal dynamics of multistep signaling events of chemoattractant gradient sensing in Dictyostelium. Mol Biol Cell. 2005; 16: 676-688.
  10. Meier-Schellersheim M, Xu X, Angermann B, Kunkel EJ, Jin T, Germain RN. Key role of local regulation in chemosensing revealed by a new molecular interaction-based modeling method. PLoS Comput Biol. 2006; 2: e82.
  11. Xu X, Meckel T, Brzostowski JA, Yan J, Meier-Schellersheim M, Jin T. Coupling mechanism of a GPCR and a heterotrimeric G protein during chemoattractant gradient sensing in Dictyostelium. Sci Signal. 2010; 3: ra71.
  12. Xu X, Meier-Schellersheim M, Yan J, Jin T. Locally controlled inhibitory mechanisms are involved in eukaryotic GPCR-mediated chemosensing. J Cell Biol. 2007; 178: 141-153.
  13. Funamoto S, Meili R, Lee S, Parry L, Firtel RA. Spatial and temporal regulation of 3-phosphoinositides by PI 3-kinase and PTEN mediates chemotaxis. Cell. 2002; 109: 611-623.
  14. Funamoto S, Milan K, Meili R, Firtel RA. Role of phosphatidylinositol 3' kinase and a downstream pleckstrin homology domain-containing protein in controlling chemotaxis in dictyostelium. J Cell Biol. 2001; 153: 795-810.
  15. Iijima M, Devreotes P. Tumor suppressor PTEN mediates sensing of chemoattractant gradients. Cell. 2002; 109: 599-610.
  16. Jin T, Zhang N, Long Y, Parent CA, Devreotes PN. Localization of the G protein betagamma complex in living cells during chemotaxis. Science. 2000; 287: 1034-1036.
  17. Li Z, Dong X, Wang Z, Liu W, Deng N, Ding Y, et al. Regulation of PTEN by Rho small GTPases. Nat Cell Biol. 2005; 7: 399-404.
  18. Li Z, Jiang H, Xie W, Zhang Z, Smrcka AV, Wu D. Roles of PLC-beta2 and -beta3 and PI3Kgamma in chemoattractant-mediated signal transduction. Science. 2000;287: 1046-1049.
  19. Murphy PM. The molecular biology of leukocyte chemoattractant receptors. Annu Rev Immunol. 1994; 12: 593-633.
  20. Sasaki AT, Chun C, Takeda K, Firtel RA. Localized Ras signaling at the leading edge regulates PI3K, cell polarity, and directional cell movement. J Cell Biol. 2004: 167: 505-518.
  21. Zhang S, Charest PG, Firtel RA. Spatiotemporal regulation of Ras activity provides directional sensing. Curr Biol. 2008;18: 1587-1593.
  22. Tang W, Zhang Y, Xu W, Harden TK, Sondek J, Sun L, et al. A PLCbeta/PI3Kgamma-GSK3 signaling pathway regulates cofilin phosphatase slingshot2 and neutrophil polarization and chemotaxis. Dev Cell. 2011; 21: 1038-1050.
  23. van Haastert PJ, Keizer-Gunnink I, Kortholt A. Essential role of PI3-kinase and phospholipase A2 in Dictyostelium discoideum chemotaxis. J Cell Biol. 2007; 177: 809-816.
  24. Veltman DM, van Haastert PJ. The role of cGMP and the rear of the cell in Dictyostelium chemotaxis and cell streaming. J Cell Sci. 2008; 121: 120-127.
  25. Charest PG, Shen Z, Lakoduk A, Sasaki AT, Briggs SP, Firtel RA. A Ras signaling complex controls the RasC-TORC2 pathway and directed cell migration. Dev Cell. 2010; 18: 737-749.
  26. Houk AR, Jilkine A, Mejean CO, Boltyanskiy R, Dufresne ER, Angenent SB, et al. Membrane tension maintains cell polarity by confining signals to the leading edge during neutrophil migration. Cell. 2012; 148: 175-188.
  27. Li H, Yang L, Fu H, Yan J, Wang Y, Guo H, et al. Association between Galphai2 and ELMO1/Dock180 connects chemokine signalling with Rac activation and metastasis. Nat Commun. 2013; 4: 1706.
  28. Wang MJ, Artemenko Y, Cai WJ, Iglesias PA, Devreotes PN. The directional response of chemotactic cells depends on a balance between cytoskeletal architecture and the external gradient. Cell Rep. 2014; 9: 1110-1121.
  29. Xu X, Gera N, Li H, Yun M, Zhang L, Wang Y, et al. GPCR-mediated PLCbetagamma/PKCbeta/PKD signaling pathway regulates the cofilin phosphatase slingshot 2 in neutrophil chemotaxis. Mol Biol Cell. 2015; 26: 874-886.
  30. Yan J, Mihaylov V, Xu X, Brzostowski JA, Li H, Liu L, et al. A Gbetagamma effector, ElmoE, transduces GPCR signaling to the actin network during chemotaxis. Dev Cell. 2012; 22: 92-103.
  31. Hoeller O, Gong D, Weiner OD. How to understand and outwit adaptation. Dev Cell. 2014; 28: 607-616.
  32. Van Haastert PJ. Down-regulation of cell surface cyclic AMP receptors and desensitization of cyclic AMP-stimulated adenylate cyclase by cyclic AMP in Dictyostelium discoideum. Kinetics and concentration dependence. J Biol Chem. 1987; 262: 7700-7704.
  33. Bolourani P, Spiegelman G, Weeks G. Determinants of RasC specificity during Dictyostelium aggregation. J Biol Chem. 2010; 285: 41374-41379.
  34. Bolourani P, Spiegelman GB, Weeks G. Delineation of the roles played by RasG and RasC in cAMP-dependent signal transduction during the early development of Dictyostelium discoideum. Mol Biol Cell. 2006; 17: 4543-4550.
  35. Insall RH, Borleis J, Devreotes PN. The aimless RasGEF is required for processing of chemotactic signals through G-protein-coupled receptors in Dictyostelium. Curr Biol. 1996; 6: 719-729.
  36. Suire S, Lecureuil C, Anderson KE, Damoulakis G, Niewczas I, Davidson K, et al. GPCR activation of Ras and PI3Kc in neutrophils depends on PLCb2/b3 and the RasGEF RasGRP4. EMBO J. 2012; 31: 3118-3129.
  37. Zheng L, Eckerdal J, Dimitrijevic I, Andersson T. Chemotactic peptide-induced activation of Ras in human neutrophils is associated with inhibition of p120-GAP activity. The Journal of biological chemistry. 1997;272: 23448-23454.
  38. Iijima M, Huang YE, Luo HR, Vazquez F, Devreotes PN. Novel mechanism of PTEN regulation by its phosphatidylinositol 4,5-bisphosphate binding motif is critical for chemotaxis. J Biol Chem. 2004; 279: 16606-16613.
  39. Benard V, Bohl BP, Bokoch GM. Characterization of rac and cdc42 activation in chemoattractant-stimulated human neutrophils using a novel assay for active GTPases. J Biol Chem. 1999; 274: 13198-13204.
  40. Li S, Yamauchi A, Marchal CC, Molitoris JK, Quilliam LA, Dinauer MC. Chemoattractant-stimulated Rac activation in wild-type and Rac2-deficient murine neutrophils: preferential activation of Rac2 and Rac2 gene dosage effect on neutrophil functions. J Immunol. 2002; 169: 5043-5051.
  41. Wong K, Pertz O, Hahn K, Bourne H. Neutrophil polarization: spatiotemporal dynamics of RhoA activity support a self-organizing mechanism. Proc Natl Acad Sci. 2006; 103: 3639-3644.
  42. Nakajima A, Ishihara S, Imoto D, Sawai S. Rectified directional sensing in long-range cell migration. Nat Commun. 2014: 5: 5367.