We have recently shown that PKC?-/- T cell-reconstituted Rag2-/- mice failed to reject fully MHC mismatched (H-2d) cardiac allografts. Sub-optimal blockade of costimulation, insufficient to prolong survival of cardiac allografts in wild-type (WT) mice, induced donor specific tolerance in PKC?-/- mice. The current results showed that cardiac allografts were rejected in PKC?-/- T cell-reconstituted Rag2-/- mice by an adoptive transfer of PKC?-/- B cells. Cardiac allografts were accepted in PKC? and B cell double knockout mice (PKC?-/-/µ MT) and PKC? and CD28 double knockout (PKC?-/-CD28-/-) mice, whereas acute rejection occurred in either CD28 or B cell single knockout mice. Bioluminescence imaging and flow cytometry analyses showed reduced NF-(B activation in the cardiac allografts, decreased frequency of Th1, Th17, and Treg subsets in the spleen cells, and diminished the percentage of IE-reactive V-beta11 subset in the peripheral blood lympocytes in PKC?-/-, and PKC?-/-CD28-/- mice. A single TLR engagement with CpG ODNs prevented tolerance induction in both PKC?-/-/µ MT and PKC?-/-CD28-/- mice, associated with the elevation of NF-(B activation and specific T cell subsets. We conclude that B cells have the ability to compensate for the deficiency of PKC?, which is CD28 dependent. CpG ODNs restore allograft rejection bypassing PKC?, CD28, and B cell signaling pathways.