Immunological adjuvants boost T cell responses. When a T cell is activated in the presence of an adjuvant, antigen specific T cell responses are prolonged and there is a greater proportion of T cells that enter the memory pool. This leads to productive immune responses as opposed to T cells that are activated in the absence of adjuvant which undergo abortive immune responses due to a decline in the number of antigen specific T cells. Natural adjuvants are materials associated with infectious organisms and have a conserved molecular pattern that is recognized by receptors on antigen presenting cells (MC) and T cells. Adjuvants are known to provide distinct set of signals via the APC or directly to the T cell which confers a survival and functional advantage which is intrinsic to the T cell.
Bcl-3 is an IκB/NFκB like factor whose transcript levels increased as a function of adjuvant stimulation in T cells. Further, enforced expression of Bcl-3 rescued activated T cells from death. Hence Bcl-3 was hypothesized to be an important pro-survival factor in adjuvanted T cells. How Bcl-3 confers survival in T cells is largely undefined and the main aim of this dissertation was to elucidate the Bcl-3 dependent biochemical pathway that is involved in exerting survival in activated CD4 T cells. We have previously shown that GSK-3β activity leads to clonal contraction of CD4 T cells. Pharmacological inhibition of GSK-3β by SB2616763 led to increased survival in non-adjuvanted CD4 T cells. In addition, it was inferred that non-adjuvanted CD4 T cells had increased GSK-3β activity and decreased protein levels of Mcl-1 a pro-survival factor that is primed for proteasomal degradation by GSK-3β. Since Bcl-3 is an important adjuvant induced prosurvival factor in CD4 T cells, it was hypothesized that Bcl-3 is required for the inhibition of GSK-3β activity to confer adjuvant induced survival. To test this hypothesis, adjuvant induced survival and signaling outcomes were assessed in CD4 T cells from Wild type (WT) or Bcl-3 knockout (KO) mice which were activated with Ova peptide323-339 (OvaP) in the presence or absence of a natural adjuvant lipopolysaccharide (LPS).
In adjuvanted CD4 T cells, the expression of Bcl-3 was increased both at the transcript and the protein level. It was seen that in the absence of bcl-3, adjuvant activation failed to rescue activated CD4 T cells from death at the peak of clonal expansion. Further, more apoptosing CD4 T cells were found in Bcl-3 KO CD4 T cells. Bcl-3 or adjuvant activation was also needed to enable the activated CD4 T cells to survive in a congenic host. The defective survival could not be correlated to the decreased expression of Bcl-2 or Bcl-x L but a failure to maintain the phospho-inhibited form of GSK-3β and Mcl-1. Pharmacological inhibition of GSK-3β increased survival in Bcl-3 KO CD4 T cells confirming GSK-3β activity as the cause of death. It could therefore be inferred that Bcl-3 is required for the maintenance of the phospho-inhibited form of GSK-3β which in turn prevents Mcl-1 from being rapidly turned over.
To gain an insight into the mechanism of Bcl-3's regulation of GSK-3β, gene array analysis was performed on adjuvanted CD4 T cells from WT or KO. It was found that the transcript level of a gene coding for calcineurin was significantly increased in Bcl-3 KO CD4 T cells. Calcineurin is one of the phosphatases that are responsible for the reactivation of GSK-3β activity. It was seen that Bcl-3 KO CD4 T cells had increased expression and activity of calcineurin and that calcineurin inhibition led to increased survival and maintenance of phospho-inhibited form of GSK-3β. Overexpression of Bcl-3 led to increased expression of phospho-inhibited GSK-3β and Mcl-1 and also decreased calcineurin activity. The above data therefore support a model wherein Bcl-3 inhibits calcineurin activity to maintain the levels of phospho-inhibited GSK-3β and Mcl-1 to promote survival in adjuvanted CD4 T cells.
|School:||University of Louisville|
|School Location:||United States -- Kentucky|
|Source:||DAI-B 68/12, Dissertation Abstracts International|
|Keywords:||Adjuvanted T cells, Bcl-3, Calcineurin, Glycogen synthase kinase|
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