Characterization of the subunit Cβ2 of Protein Kinase A (PKA) DNA mapping, glucose uptake in KO mice and reaction in T cell activity as a response to incremental concentrations of glucose

Protein Kinase A (PKA) is a holoenzyme that consists of a regulatory (R) subunit dimer and two catalytic (C) subunits. Genes, which encode the catalytic subunit C, PRKACA and PRKACB, have several splice variants including Cβ2. Cβ2 subunit is highly expressed in T, B and Natural Killer cells. PKA regulates several functions including immune cell proliferation and glucose uptake and metabolism. We have used mice knocked out for the Cβ2 subunit of PKA. Polymerase Chain Reaction was used to clarify mice genotype and Western Blot analysis to verify ablation of Cβ2 protein in KO mice. Catalytic activity was significantly downregulated by 40 % in Cβ2 KO lymph node, spleen and thymus cells, suggesting that Cβ2 activity could be involved in the regulation of cell proliferation. We therefore used CD3/CD28 coated beads for stimulation of T cells and observed no difference in proliferation rates between Cβ2 KO lymphocytes and wild type cells from mice lymph nodes. Because the results could have been influenced by other cells, we repeated these experiments with positively isolated CD4+ T cells, which verified our previous result. There was, however, a significant increase in proliferation rate in Cβ2 KO spleen cells compared to wt and that was absent in cells isolated from lymph nodes. The biological significance of this observation is unclear. We also found unaltered RIα and RIIα subunit expressions in Cβ2 KO lymph node, spleen and thymus cells and that a mixed T cell population required glucose in order to proliferate. While investigating whether Cβ2 ablation could have an effect on glucose consumption we found this not to be the case. We did, however, find that Cβ2 could possess a regulatory link in the conversion and boosting effect of pyruvate. Taken into account that catalytic activity was reduced in all tissues but cells maintained their proliferation rate, even under different concentrations of glucose, Cβ2 does not appear to be important for cell proliferation or energy generation