Characterisation of NF-kappa B complexes in Theileria parva-transformed T cells

Transformation of T cells by the intracellular parasite Theileria parva is accompanied by constitutive I-kappaB degradation and NF-kappaB activation, a process which is essential to prevent the spontaneous apoptosis of these parasite-transformed cells. NF-kappaB-mediated responses are regulated by selective combinations of NF-kappaB proteins as homo- or heterodimers and by distinct kappaB motifs. We characterised the NF-kappaB complexes induced by T. parva infection in TpM(803)T cells. By western blot, we demonstrated that all members of the NF-kappaB/Rel family of proteins translocate to the nucleus of infected cells. Using two different kappaB oligonucleotides (kappaB-1 and kappaB-2), both containing the decameric consensus kappaB motif(GGGACTTTCC), clearly distinct patterns of DNA binding activities could be demonstrated in electrophoretic mobility shift assays. Supershift analysis and UV crosslinking assays showed that complexes binding to kappaB-1 consisted of p50, p65 and RelB home and/or heterodimers. We could also detect an association of ATF-2 and c-Fos with one of the complexes. The HIV-derived kappaB-2 oligo only bound p50 and p65. Additionally, several agents known to inhibit a wide range of NF-kappaB activation pathways had no inhibitory effect on the activation of NF-kappaB DNA binding in TpM(803)T cells. (C) 2000 Editions scientifiques et medicales Elsevier SAS.