RBP-L, a Transcription Factor Related to RBP-Jκ

Abstract

RBP-J is a sequence-specific DNA binding protein which plays a central role in signalling downstream of the Notch receptor by physically interacting with its intracellular region. Although at least four Notch genes exist in mammals, it is unknown whether each Notch requires a specific downstream signalling molecule. Here we report isolation and characterization of a mouse RBP-J-related gene named RBP-L that is expressed almost exclusively in lung, in contrast to the ubiquitous expression of RBP-J. For simplicity, we propose to call RBP-J RBP-J. The RBP-L protein bound to a DNA sequence almost identical to that of RBP-J. Surprisingly, RBP-L did not interact with any of the known four mouse Notch proteins. Although we found that RBP-L and EBNA-2 cooperated in transcriptional activation, they did not show significantly strong protein-protein interaction that can be detected by several in vivo and in vitro assays. This is again in contrast to physical association of RBP-J with EBNA-2. Several models to explain functional interaction between RBP-L and EBNA-2 are discussed.RBP-J is a 60-kDa DNA binding protein recognizing a consensus sequence (C/T)GTGGGAA although it has no typical DNA binding motif (19,23,38,52). The structure of the RBP-J protein is strongly conserved during evolution among nematode, fruit fly, mouse, and human (1, 5, 13, 38).We and others have shown that the Drosophila RBP-J gene is identical to Suppressor of Hairless [Su(H)] (14, 46), a member of the neurogenic gene family including Notch, Delta, Enhancer of split [E(spl)], and Hairless. Genetic analyses have shown that the neurogenic genes, including Su(H), participate in lateral inhibition to single out a sensory mother cell from its precursor cells during peripheral nervous system development (2, 40). RBP-J/Su(H) binding sites were identified in the 5Ј-flanking regions of the E(spl) complex [E(spl)-C] neurogenic genes of Drosophila melanogaster, and the transactivation of the E(spl) m8 enhancer promoter by Su(H) was demonstrated in the Schneider cell line (16). Transactivation of the E(spl)-C promoters by Su(H) was also demonstrated in vivo by using transgenic flies carrying E(spl)-C promoter-␤-galactosidase gene constructs (3,15,16,33).A breakthrough in the elucidation of the RBP-J function in vertebrates came from studies on transcriptional regulation of human DNA tumor viruses. The adenovirus capsid protein polypeptide IX (pIX) promoter contains the RBP-J targetlike TGGGAAAGAA sequence between the SP1 binding site and the TATA box. Repression of the pIX promoter by RBP-J was shown by in vitro as well as in vivo experiments (9). Epstein-Barr virus (EBV) nuclear antigen 2 (EBNA-2) is essential for transformation of primary human B lymphocytes (7, 21, 58) and acts as a transcriptional activator of latent viral as well as cellular genes by interacting with 22,54,61). Thus, RBP-J is essential to B-lymphocyte transformation by EBV.RBP-J knockout mice die before 10.5 days of gestation and show severe developmental defects in somites and neural tube...

The Notch signaling pathway: Transcriptional regulation at Notch target genes

“…The DNA binding sequence was identified as 5 -CGTGGGAA-3 [64]. A paralogue of RBP-J, RBP-L, was also described that is almost exclusively expressed in the lung [65]. RBP-L binds to DNA identical to RBP-J.…”

Neoplastic transformation by Notch is independent of transcriptional activation by RBP-J signalling

“…Details on the construction of the expression plasmids are available on request. The luciferase reporter plasmids RBP-J(6x)-Luc (ERE-TP1-Luc/ Ga981-6) and RBP-J(0x)-Luc (Ga50-1) have been described previously (Minoguchi et al, 1997). The GAL4 responsive promoter construct (pCR276-3) was constructed by replacing the EBNA2 responsive region of the TP-1 promoter between 7258 and 7142 relative to the TP-1 RNA start site by ®ve GAL4 binding sites.…”

Stimulation of p53 Transactivation Ability by Nicastrin in Mouse Fibroblasts

“…The transactivation ability of Notch-IC can be estimated by luciferase assay using the pTP1-Luc reporter plasmid which contains the Notch-binding domains [23]. The luciferase activity was markedly elevated by transfection with the positive control Notch-ΔE (Figure 2(a)) which lacks the TACE-eliminated domain and can, therefore, be activated only by γ-secretase [22].…”

“…pCS2 + -Notch-ΔE [22] was provided by Dr. Tasuku Honjo (Kyoto University). pTP1-Luc [23] was provided by Dr. Lothar J. Strobl (GSF National Research Center). pGL3-Bax-Luc and pGL3-p21-Luc [24] were provided by Dr. Mian Wu (University of Science and Technology of China).…”