Dok-1 and Dok-2 Regulate the Formation of Memory CD8+ T Cells

Abstract

Diverse signals received by CD8 T cells are integrated to achieve the required magnitude of cell expansion and the appropriate balance of effector/memory CD8 T cell generation. Notably, the strength and nature of TCR signaling influence the differentiation and functional capacity of effector and memory CD8 T cells. Dok-1 and Dok-2, the two members of the Dok family expressed in T cells, negatively regulate TCR signaling in vitro. However, the role of Dok proteins in modulating T cell function in vivo has not yet studied. We studied the function of Dok-1 and Dok-2 proteins in the regulation of the CD8 T cell response to vaccinia virus infection. Comparison of responses to vaccinia virus expressing OVA peptide SIINFEKL by wild-type and Dok-1/2 CD8 OT-I cells showed that the absence of Dok-1 and Dok-2 slightly reduced the magnitude of virus-specific effector CD8 T cell expansion. This was not due to reduced proliferation or enhanced apoptosis of effector CD8 T cells. Dok-1/2-deficient effector CD8 T cells showed increased cell surface TCR expression following virus infection in vivo and increased expression of granzyme B and TNF upon stimulation with peptide Ag ex vivo. Finally, Dok-1/2-deficient effector CD8 T had a severe defect in survival that resulted in impaired generation of memory CD8 T cells. These results reveal the critical involvement of Dok-1 and Dok-2 in a negative-feedback loop that prevents overactivation of CD8 T cells and promotes memory formation.

Dok-1 and Dok-2 Are Required To Maintain Herpes Simplex Virus 1-Specific CD8+T Cells in a Murine Model of Ocular Infection

“…We found that Dok-1 and Dok-2 are important for the formation of long-lived HSV-1-specific CD8 ϩ T cells in lymphoid and nonlymphoid organs. We previously reported the importance of Dok-1/2 in memory T cell formation in response to vaccinia virus expressing OVA peptide using an adoptivetransfer system of DKO OT-I cells (37). Just as we showed for VV OVA infection, the number of HSV-specific CD8 ϩ T cells in response to HSV-1 infection was diminished at the peak of the effector response.…”

“…The virus was propagated and titrated on Vero cells as described previously (49). Dok-1-deficient (Dok-1 KO) mice, Dok-2-deficient (Dok-2 KO) mice, and Dok-1/Dok-2 double-deficient (DKO) mice have been described previously (37). All colonies were maintained under pathogen-free conditions at the INRS Centre for Experimental Biology.…”

“…Pooled TG (2 TG equivalents) and pooled eyeballs (4 eyeball equivalents) were incubated in DMEM containing 51 U/ml of collagenase type 1 (Invitrogen) for 1 to 2 h at 37°C, dispersed into single-cell suspension by pipetting, and filtered through 40-m-pore-size filters to remove debris prior to staining. Spleens and TG were treated with red blood cell lysis buffer prior to their use, as described previously (37). Single-cell suspensions of various organs were stained with gB tetramers for 30 min at room temperature (RT), followed by cell surface marker staining for 20 min at 4°C in fluoresence-activated cell sorter (FACS) buffer (phosphate-buffered saline [PBS], 1% fetal bovine serum, 0.05% sodium azide, 2 mM EDTA).…”

“…Using Dok-1-and -2 (Dok-1/2)-deficient mice, biological roles for Dok-1 and Dok-2 have been demonstrated in antibody responses to thymus-dependent antigens, NK and hematopoietic cell development and function, innate immune response to lipopolysaccharide (LPS), myeloid homeostasis, and leukemia suppression (29)(30)(31)(32)(33)(34)(35)(36). In addition, we recently demonstrated that Dok proteins regulate the CD8 ϩ T cell response to an exogenous epitope expressed by vaccinia virus (VV) (37). In particular, we showed that Dok proteins negatively regulate the effector function of CD8 ϩ effector T cells and play a critical role in the generation of CD8 ϩ memory T cells (37).…”

“…In addition, we recently demonstrated that Dok proteins regulate the CD8 ϩ T cell response to an exogenous epitope expressed by vaccinia virus (VV) (37). In particular, we showed that Dok proteins negatively regulate the effector function of CD8 ϩ effector T cells and play a critical role in the generation of CD8 ϩ memory T cells (37). As mentioned above, during HSV-1 infection, CD8 ϩ T cells are important in controlling the infection, in particular in maintaining HSV-1 latency (24,25).…”