An optimal range of transcription potency is necessary for efficient cell transformation by c-Rel to ensure optimal nuclear localization and gene-specific activation

Abstract

c-Rel is overexpressed in several B-cell lymphomas and c-rel gene overexpression can transform primary chicken lymphoid cells and induce tumors in animals. Although c-Rel is generally a stronger transcriptional activator than its viral derivative v-Rel, its oncogenic activity is significantly weaker. Among the mutations acquired during c-Rel's evolution into v-Rel are deletion of c-Rel's transactivation domain 2 (cTAD2) and mutations in cTAD1. Given the critical role of the Rel TADs in cell transformation, we investigated how mutations in c-Rel's cTAD1 and cTAD2 contribute to its oncogenicity and that of v-Rel. Mutations in cTAD2 noticeably increased c-Rel's transforming activity by promoting its nuclear localization and gene-specific transactivation, despite an overall decrease in jB site-dependent transactivation potency. Conversely, substitution of vTAD by cTAD1 increased v-Rel's transactivation and transforming efficiencies, whereas its substitution by the stronger cTAD2 compromised activation of mip-1b but not irf-4 and was detrimental to cell transformation. These results suggest that the Rel TADs differentially contribute to genespecific activation and that an optimal range of transcription potency is necessary for efficient transformation. These findings may have important implications for understanding how Rel TAD mutations can lead to a more oncogenic phenotype.

CAPERα Is a Novel Rel-TAD-Interacting Factor That Inhibits Lymphocyte Transformation by the Potent Rel/NF-κB Oncoprotein v-Rel

“…Indeed work from Gilmore's group and ours unveiled an inverse correlation between the strength of the v-Rel-and c-Rel TADs and their transforming efficiency (12,43,44). The fact that naturally occurring c-rel gene mutations in PMBCL and follicular lymphoma specimens decrease cRel's transactivation potency and enhance its transforming activity in chicken lymphocytes is also consistent with the idea that too much Rel transcriptional activity can be detrimental to cell transformation (45).…”

“…Additionally, some PMBCL and follicular lymphomas harbor c-rel gene mutations that decrease c-Rel's transactivation potency and enhance its transforming activity in primary chicken lymphocytes (45). This is consistent with the increased transforming phenotype conferred by certain mutations in v-Rel and c-Rel and indicates that modulation of Rel's transcriptional activity can significantly affect its oncogenicity (12,43,44).…”

“…Additionally, some PMBCL and follicular lymphomas harbor c-rel gene mutations that decrease c-Rel's transactivation potency and enhance its transforming activity in primary chicken lymphocytes (45). This is consistent with the increased transforming phenotype conferred by certain mutations in v-Rel and c-Rel and indicates that modulation of Rel's transcriptional activity can significantly affect its oncogenicity (12,43,44).The Rel proteins bind to B DNA sites as homo-or heterodimers with other NF-B family members via their N-terminal Rel homology domain (RHD), which also mediates nuclear localization and association with inhibitory IB subunits. …”

“…The 293T human embryonic kidney cell line, primary chicken embryo fibroblasts (CEFs), v-Rel-transformed chicken spleen cells (CSC), and the DT40 chicken pre-B-cell line were cultured as described previously (12,19). Luciferase assays were performed with extracts from 293T cells (1 ϫ 10 6 ) transfected with calcium phosphate using a total of 6.01 g DNA, including 0.01 g of hRL-null Renilla luciferase DNA as a control, as described previously (12). Extracts were prepared at 48 h posttransfection and analyzed for protein concentration and dual-luciferase activity (Promega Corp., Madison, WI).…”

Repression of B-Cell Linker (BLNK) and B-Cell Adaptor for Phosphoinositide 3-Kinase (BCAP) Is Important for Lymphocyte Transformation by Rel Proteins

“…Antibodies were against Rel, extracellular signal-regulated kinase (ERK), Akt (Santa Cruz), RelA-N (Rockland), actin (Sigma), phosphorylated ERK, phosphorylated Akt (Cell Signaling), Rel-RHD (2716; ref. 24), enhanced green fluorescent protein (EGFP; Torrey Pines), ch-BLNK or BCAP (gifts from T. Kurosaki, Kansai Medical University). c-Jun-NH 2 -kinase (JNK) activity was determined in in vitro kinase assays with cell lysates immunoprecipitated with anti-JNK1 (Santa Cruz), 32 P-gATP, and a glutathione S-transferasec-Jun substrate (Cell Signaling), followed by autoradiography and immunoblotting with anti-JNK1, as detailed in the Supplementary Materials.…”