Chemotaxis, or directional movement of cells up a chemical gradient, plays important roles in a wide range of cellular behaviors. Naive, randomly moving cells, such as leukocytes and Dictyostelium cells must be able to directionally sense extremely shallow extracellular chemoattractant gradients and respond by establishing a polarized cytoskeleton with an F-actin-based protruding pseudopod or lamellipod at the leading edge and a contractile trailing edge. This polarization is achieved through the spatially restricted amplification of the external gradient through the localized activation of pathways at the site of the cortex closest to the chemoattractant source. While significant advancements have been made in identifying many components that control chemotaxis, we do not yet understand how cells sense the direction of the gradient and restrict responses to the leading edge, how newly polymerized F-actin networks are stabilized to form pseudopod, or what regulates the timing of these events. Furthermore, chemotaxing cells are able to communicate, within the cell population, to organize more global social responses, such as the autocrine aggregation response in Dictyostelium cells in which cells respond to and then secrete the chemoattractant cAMP leading to the coordinate aggregation of ~105 cells (signal relay) or the paracrine response system used by macrophages and mammary tumor cells, resulting in interdependent migration of the two cell types during metastasis. As many of the pathways are evolutionarily conserved, we will use Dictyostelium to achieve three important goals: 1) We have demonstrated that TORC2, through the activation of PKB and PKBR1, integrates multiple inputs to regulate chemotaxis by controlling directional sensing and F-actin polymerization, and to control signal relay through the regulation of adenylyl cyclase. Furthermore, we have found that cAMP, through the activation of PKA, acts as an important feedback loop to negatively regulate the TORC2/PKB:PKBR1 pathway. We will investigate the molecular basis of this key feedback loop. 2) We have found that GSK-3 is an important regulator of chemotaxis and by comparing the phosphoproteomes of wild-type and GSK-3 null cells have identified putative GSK-3 substrates, including known components of the Ras/Rap1/F-actin pathways that control chemotaxis. Our goal is to define key GSK- 3 substrates and understand how GSK-3 functions to control chemotaxis. 3) We have identified a novel regulatory pathway that plays a central role in forming and stabilizing the pseudopod during chemotaxis. We have shown that Rab1A binds to and is required to activate Roco2, a member of the Roco family of GTPase-containing protein kinases. Roco2 acts upstream of the F-actin cross- linking protein filamin and both are required for pseudopod extension. We will examine the mechanism by which this pathway is regulated and how it controls chemotaxis.

Public Health Relevance

The research will focus on understanding the basic mechanisms by which cells are able to directionally move in response to a chemical gradient or chemotaxis, a process that is used by cells during wound repair, bacterial infection, and metastasis of cancer cells. The findings illuminated by the proposed work will shed light on the molecular basis of human disease and may help, in the future, identify new targets for treatment.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM037830-28
Application #
8608529
Study Section
Special Emphasis Panel (ZRG1-CB-P (02))
Program Officer
Gaillard, Shawn R
Project Start
1986-07-01
Project End
2016-01-31
Budget Start
2014-02-01
Budget End
2015-01-31
Support Year
28
Fiscal Year
2014
Total Cost
$526,995
Indirect Cost
$186,998
Name
University of California San Diego
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
804355790
City
La Jolla
State
CA
Country
United States
Zip Code
92093
Khanna, Ankita; Lotfi, Pouya; Chavan, Anita J et al. (2016) The small GTPases Ras and Rap1 bind to and control TORC2 activity. Sci Rep 6:25823
Bastounis, Effie; Álvarez-González, Begoña; del Álamo, Juan C et al. (2016) Cooperative cell motility during tandem locomotion of amoeboid cells. Mol Biol Cell 27:1262-71
Álvarez-González, Begoña; Meili, Ruedi; Bastounis, Effie et al. (2015) Three-dimensional balance of cortical tension and axial contractility enables fast amoeboid migration. Biophys J 108:821-32
Alvarez-González, Begoña; Meili, Ruedi; Firtel, Richard et al. (2014) Cytoskeletal Mechanics Regulating Amoeboid Cell Locomotion. Appl Mech Rev 66:
Bastounis, Effie; Meili, Ruedi; Álvarez-González, Begoña et al. (2014) Both contractile axial and lateral traction force dynamics drive amoeboid cell motility. J Cell Biol 204:1045-61
Kölsch, Verena; Shen, Zhouxin; Lee, Susan et al. (2013) Daydreamer, a Ras effector and GSK-3 substrate, is important for directional sensing and cell motility. Mol Biol Cell 24:100-14
del Álamo, Juan C; Meili, Ruedi; Álvarez-González, Begoña et al. (2013) Three-dimensional quantification of cellular traction forces and mechanosensing of thin substrata by fourier traction force microscopy. PLoS One 8:e69850
Takeda, Kosuke; Shao, Danying; Adler, Micha et al. (2012) Incoherent feedforward control governs adaptation of activated ras in a eukaryotic chemotaxis pathway. Sci Signal 5:ra2
Alonso-Latorre, Baldomero; Del Álamo, Juan C; Meili, Ruedi et al. (2011) An Oscillatory Contractile Pole-Force Component Dominates the Traction Forces Exerted by Migrating Amoeboid Cells. Cell Mol Bioeng 4:603-615
Bastounis, Effie; Meili, Ruedi; Alonso-Latorre, Baldomero et al. (2011) The SCAR/WAVE complex is necessary for proper regulation of traction stresses during amoeboid motility. Mol Biol Cell 22:3995-4003

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