Cytotoxic T lymphocytes (CTLs) play a central role in cellular immune responses by destroying infected or transformed target cells. Their potent anti-tumor activity has made them the centerpiece of several promising immunotherapeutic strategies to fight cancer. CTLs operate by forming a close, radially symmetric contact with their target cell known as an immunological synapse (IS). Then, they secrete cytolytic molecules into the synaptic space to induce target cell death. It is generally thought that the cytoskeletal framework of the IS potentiates this response. Cortical filamentous actin (F-actin) is enriched in the periphery of the IS and depleted from the center, forming a characteristic ring. Concomitantly, the centrosome, which serves as a focal point for intracellular vesicular cargo, reorients to a position just beneath the center of the IS. It has been proposed that these events focus secretion toward the target cell by bringing granules containing cytolytic factors close to the synaptic membrane. This model has not been rigorously tested, however. In addition, other possible roles for cytoskeletal dynamics at the IS, such as the transfer of mechanical signals to the target cell, remain unexplored. Over the past five years, we have defined two key mechanisms that shape the synaptic cytoskeleton, a diacylglycerol dependent pathway that guides the centrosome and a phosphoinositide 3-kinase (PI3K) dependent pathway that controls F-actin ring formation. We will now examine how these mechanisms contribute to CTL function. Our overall hypothesis is that centrosome polarization, central F-actin clearance, and peripheral F-actin dynamics together provide strength and specificity to CTL effector responses. The following Specific Aims will be pursued: 1) Determine the role of centrosome polarization in guiding CTL secretory responses; 2) Determine the mechanism and function of F-actin clearance; and 3) Determine how PI3K dependent force exertion potentiates cytotoxicity. For the first Aim, a novel genetic strategy based on conditional deletion of the scaffolding protein SAS4 will be used to remove the centrosome from CTLs. For the second Aim, gain- and loss-of-function experiments will be used to evaluate how actin- remodeling factors of the gelsolin family influence cytoskeletal polarization and CTL effector responses. For the third Aim, biophysical approaches will be used to quantify mechanotransduction at the IS and examine how force exertion potentiates target cell killing. This work is technically innovative because it incorporates state-of- the-art imaging modalities, genetic tools, and biophysical methods to explore lymphocyte function in new ways. It also advances innovative concepts about how the structure of the IS facilitates the transfer chemical and also mechanical information to the target cell. The proposed studies are important because they will lead to a comprehensive understanding of how synaptic architecture specifies CTL function, which will aid efforts to harness and control CTL activity in immunotherapeutic contexts. Hence, our proposal is relevant to the NIH mission in that it will contribute to the advancement of knowledge that could improve human health.

Public Health Relevance

Cytotoxic T lymphocytes (CTL) play a crucial role in immune responses by killing infected or cancerous target cells. To carry out these functions, CTLs form specific interactions with target cells called immunological synapses. This proposal seeks to determine how the structure of these synapses contributes to CTL responses. A better understanding of this issue could aid immunotherapeutic efforts that harness CTL activity to fight cancer and other diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI087644-09
Application #
9614265
Study Section
Cellular and Molecular Immunology - A Study Section (CMIA)
Program Officer
Mallia, Conrad M
Project Start
2010-02-15
Project End
2020-12-31
Budget Start
2019-01-01
Budget End
2019-12-31
Support Year
9
Fiscal Year
2019
Total Cost
Indirect Cost
Name
Sloan-Kettering Institute for Cancer Research
Department
Type
DUNS #
064931884
City
New York
State
NY
Country
United States
Zip Code
10065
Sanchez, Elisa; Huse, Morgan (2018) Spatial and Temporal Control of T Cell Activation Using a Photoactivatable Agonist. J Vis Exp :
Huse, Morgan (2017) Mechanical forces in the immune system. Nat Rev Immunol 17:679-690
Oyler-Yaniv, Alon; Oyler-Yaniv, Jennifer; Whitlock, Benjamin M et al. (2017) A Tunable Diffusion-Consumption Mechanism of Cytokine Propagation Enables Plasticity in Cell-to-Cell Communication in the Immune System. Immunity 46:609-620
Basu, Roshni; Huse, Morgan (2017) Mechanical Communication at the Immunological Synapse. Trends Cell Biol 27:241-254
Basu, Roshni; Whitlock, Benjamin M; Husson, Julien et al. (2016) Cytotoxic T Cells Use Mechanical Force to Potentiate Target Cell Killing. Cell 165:100-110
Le Floc'h, Audrey; Huse, Morgan (2015) Molecular mechanisms and functional implications of polarized actin remodeling at the T cell immunological synapse. Cell Mol Life Sci 72:537-556
Huse, Morgan (2014) Lipid-based patterning of the immunological synapse. Biochem Soc Trans 42:1506-11
Chauveau, Anne; Le Floc'h, Audrey; Bantilan, Niels S et al. (2014) Diacylglycerol kinase ? establishes T cell polarity by shaping diacylglycerol accumulation at the immunological synapse. Sci Signal 7:ra82
Wu, Kong-Yan; He, Miao; Hou, Qiong-Qiong et al. (2014) Semaphorin 3A activates the guanosine triphosphatase Rab5 to promote growth cone collapse and organize callosal axon projections. Sci Signal 7:ra81
Basu, Roshni; Chen, Yuedan; Quann, Emily J et al. (2014) The variable hinge region of novel PKCs determines localization to distinct regions of the immunological synapse. PLoS One 9:e95531

Showing the most recent 10 out of 16 publications