Cytotoxic T lymphocytes (CTLs) are essential for productive immune responses and recent studies have demonstrated their strong clinical potential as immunotherapeutic agents against established tumors. Cytotoxic responses in CTLs are induced by the T cell receptor, which recognizes cognate peptide-major histocompatibility complex molecules on the surfaces of infected or transformed cells, which induces the formation the immunological synapse. Synapse formation is accompanied by a series of actin and microtubule remodeling events, the most striking of which is the dramatic reorientation of the centrosome to a position just beneath the center of the contact site. It has been proposed that lytic granules containing perforin and granzyme cluster around the centrosome in activated CTLs. This polarization toward the synapse promotes the selective fusion of these granules with the synaptic membrane, leading to directional release of their contents toward the target cell.
In order to study more precisely how the centrosome influences secretory responses and how these contributions affect CTL activity, we have developed an approach using OT-I mice bearing a conditional allele of both p53 (p53flox) as well as SAS4 (SAS4flox), a scaffolding protein that is absolutely required for the formation and maintenance of the centrosome. OT-I SAS4−/−p53−/− cells were generated by transducing SAS4flox/floxp53flox/flox cells with a retrovirus expressing Cre. Using gamma-tubulin and pericentrin staining, we were able to show that SAS4 deletion effectively eliminates interphase centrosomes in CTLs. OT-I SAS4−/− p53−/− cells exhibited a marked defect in target cell killing after 24h, implying a crucial role of the centrosome in cytotoxic responses. This killing defect was accompanied by a decrease in both granzyme B and perforin production. IFN-gamma production and secretion were not altered by the loss of the centrosome. Importantly, TCR induced signaling was normal in these cells.
In summary, we were able to show that centrosome deletion impairs target cell killing without altering their overall magnitude. Given the importance of centrosome polarization for lytic granules delivery, we can assume that the centrosome ablation selectively alters cytotoxic efficiency by disrupting the directional synaptic secretion.
Citation Format: Fella Tamzalit, Ariella Kepecs, Hisham Bazzi, Kathryn Anderson, Morgan Huse. The role of the centrosome in cytotoxic T cell function [abstract]. In: Proceedings of the Second CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; 2016 Sept 25-28; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(11 Suppl):Abstract nr A070.
- ©2016 American Association for Cancer Research.