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B Activity in the Nucleus Through a Direct Interaction Involving a Novel Domain1
Laboratory of Gene Regulation, Division of Therapeutic Proteins, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD 20892
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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B/Rel
family of transcription factors are involved in the regulation of a
variety of genes important for immune function. The biological activity
of the NF-B transcription factors is controlled by IB proteins.
Our previous work demonstrated that an intracellular, constitutively
active form of human Notch-1/translocation-associated Notch homologue-1
(NotchIC) functions as an IB molecule with specificity
for the NF-B p50 subunit and physically interacts with NF-B in T
cells. In the current study, we investigated the roles of different
domains of NotchIC in the regulation of NF-B-directed
gene expression and NF-B DNA binding activity. We found that
NotchIC localizes to the nucleus and that a region in the
N-terminal portion of NotchIC, not the six ankyrin repeats,
is responsible for the inhibitory effects of Notch on NF-B-directed
gene expression and NF-B DNA binding activity. The N-terminal
portion of NotchIC inhibited p50 DNA binding and interacted
specifically with p50 subunit, not p65 of NF-B. The interaction
between Notch and NF-B indicates that in addition to its role in the
development of the immune system, Notch-1 may also have critical
functions in the immune response, inflammation, viral infection, and
apoptosis through control of NF-B-mediated gene
expression. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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.
Notch receptors are proteolytically processed by a furin-like protease
in the trans-Golgi network into the extracellular subunit
NotchEC and a transmembrane C-terminal subunit.
These two subunits are presented as a functional heterodimer on the
cell surface (6, 7). Ligands such as Delta or
Jagged/Serrate are thought to induce a second cleavage that requires
presenilins (
-amyloid precursor processing-related proteins) at a
site within the transmembrane region of Notch (8, 9). This
cleavage is suggested to release the intracellular fragment
NotchIC, which translocates to the nucleus
(9, 10, 11, 12). NotchIC binds to numerous
nuclear and cytoplasmic proteins (1, 13). The
best-characterized Notch signaling pathway is thought to be mediated by
binding to the transcription factors of the C promoter binding factor
(CBF)-1/suppressor of hairless/Lag-1 family. The mammalian member of
this family is CBF-1, also known as RBP-J. CBF-1 is a DNA binding
protein that recognizes the sequence CGTGGGAA. In the absence of
Notch, it functions as a repressor that recruits silencing complexes
containing various nuclear corepressors and histone deacetylase 1
(14, 15). Interaction with NotchIC
appears to displace the silencing complex from CBF-1, converting it
into a transcriptional activator (14). The nuclear
coactivator ski- interacting protein is recruited to the
CBF-1-Notch complex through an interaction with the ankyrin repeats,
and it may mediate transcriptional activation by the complex
(16). Downstream genes whose expression is up-regulated by
Notch activation include the Enhancer of Split group in
Drosophila and its mammalian homologues. This group includes
several basic helix-loop-helix proteins, such as Hairy/Enhancer of
Split-1 and -5 and Enhancer of Split-2 (the homologue of
Drosophila groucho) (1).
The mammalian Notch-1 gene is expressed in a broad range of tissues,
including the thymus and other lymphoid organs (17, 18, 19).
Notch-1 plays multiple nonmutually exclusive roles in T cell
development (20, 21). Constitutively active Notch-1
expression in hemopoietic progenitors blocks the development of
lymphoid precursor toward the B cell lineage and promotes the formation
of immature CD4+CD8+ T cell
lymphomas (22). Conversely, inducible deletion of the
Notch-1 gene in mice by cre/lox recombination
(23) leads to severe thymic atrophy due to a block of the
expansion of immature
CD4-CD8- T cell
precursors. Reduced Notch activity favors the 
T cell fate over
the 
T cell fate (24). Expression of an activated
form of Notch-1 in murine thymocytes under the lck promoter
leads to an increase in CD8+ T cells and a
parallel decrease in CD4+ T cells (21, 25). More recently, Yasutomo et al. (26) have shown
that Notch-1 is necessary for the postcommitment development of
CD8+ T cells rather than for the lineage choice
between CD4 and CD8.
Truncated forms of Notch receptors that lack all or most of the
extracellular region are constitutively active and have transforming
activity in vitro and in vivo (1, 27). Such a
constitutively active Notch-1 is produced by a 9:7 translocation in
about 10% of the cases of human T cell lymphoblastic leukemia
(17). This was defined translocation-associated Notch
homologue (TAN-1). Overexpression of apparently wild-type Notch-1 is
found in cervical carcinomas and various other solid tumors (28, 29). Depending upon the experimental model, Notch-1 activation
can delay terminal differentiation (30, 31), modulate cell
cycle progression (31), and regulate cell death
(32, 33, 34). Aside from CBF-1- or Deltex-mediated regulation
of basic helix-loop-helix transcription factors, other molecular
targets of Notch-1 have been identified. Notch-1 has been reported to
inhibit NF-B-induced gene expression (35), to regulate
the expression of transcription factor peroxisomal
proliferator-activated receptors (36), and to induce
NF-B2 (p100) promoter activity (37). The orphan nuclear
receptor nur77 (33) and the transcriptional regulator
EMB-5 (38) of Caenorhabditis elegans are
other putative mediators of Notch effects. These mediators have been
shown to bind to the Notch subunit in various experimental systems
(13).
NF-B is the prototype of a family of dimeric transcription factors
that play fundamental roles in the immune system (39, 40).
The nuclear translocation and DNA binding of Rel-NF-B complexes are
prevented by the IB family of proteins (40, 41, 42). These
include IB-, IB-, IB-
, Bcl-3, , and the
Drosophila protein cactus. In resting cells, IB proteins
sequester NF-B in the cytoplasm. Signal-induced activation of IB
kinase (43, 44, 45) by IB kinase kinase, NAK
(46), phosphorylates IBs and triggers their degradation
(47). IB phosphorylation and degradation lead to
transcriptional activation of NF-B and rapid resynthesis of IB.
Resynthesized IB translocates to the nucleus, inhibits NF-B DNA
binding, and promotes the transport of NF-B from the nucleus to the
cytoplasm (48, 49).
The NF-B transcription factors play multiple, crucial roles in the
inflammatory and immune responses as well as in the development of the
immune and hemopoietic systems. Regulations of cell survival,
proliferation, and cytokine production are some of the many functions
attributed to these transcription factors (40).
The functional interaction between the Notch signaling network and the
NF-B family is complex. We demonstrated that constitutively active
human Notch-1/TAN-1 functions as a novel IB molecule and regulates
NF-B-mediated gene expression through a direct interaction with the
NF-B p50 subunit. This prevents NF-B from binding to NF-B
recognition sites in DNA to regulate NF-B-dependent gene expression
(35). Our data have shown for the first time that Notch
directly interacted with the nuclear transcription factor NF-B
family member and revealed an important molecular basis for the pivotal
role of Notch in the immune system. Other investigators have shown
recently that Rel-NF-B can activate the Notch signaling pathway by
inducing the expression of Jagged 1, a ligand for Notch receptors
(21), and that NF-B2 (p100/p52) is a putative target
gene of activated Notch-1 via CBF-1/RBP-J (37).
The mechanism of the direct interaction of Notch-1 with p50 and its
possible physiological roles vis-à-vis the other Notch signaling
pathways remain unclear. By analogy with IB, the interaction of
Notch with NF-B may have been through the ankyrin repeats of
Notch.
In the present study, we have mapped the interaction of Notch/TAN-1
with NF-B. We found that this interaction is mediated by a domain of
Notch-1 that is N-terminal to the ankyrin repeats and appears to be
independent of these repeats. Thus, Notch-1 regulates NF-B through a
novel mechanism that does not involve ankyrin repeats interactions. The
possible significance of this mechanism is discussed.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Notch mutants were amplified by PCR using high fidelity PFU DNA
polymerase (Stratagene, La Jolla, CA). Specific primers for each mutant
are as follows: N + 6ANK, P1/G2; C + 6ANK, P2/G1; 6ANK, P2/G2; N,
P1/G5; P1 (sense), TCG GAT CCA TGG GCT TCA AAG TGT CTG AG; P2 (sense),
TCG GAT CCA TGG GGC CTG ATG GCT TCA CCC CG; G1 (antisense), GTG AAT
TCG AGG GGG CAC GGA CGG AGA C; G2 (antisense), GTG AAT TCC ATA TGA TCC
GTG ATG TC; and G5 (antisense), GTG AAT TCA GGC CCG CGG ACA TTG AC.
Amplified PCR products were purified, then digested with
BamHI and EcoRI and cloned into pcDNA3 vector for
expression in eukaryotic cells. The sequence of each mutant was
verified, and each mutant insert was transferred into pGEX1
T vector
for expression in bacteria. The reporter plasmid p-6B-luc (provided
by Dr. P. Baeuerle, Micromet, Martinsried, Germany) contains a
luciferase gene driven by six reiterated B sites. Plasmids
expressing p50 and p65 have been described previously
(50).
Bacterial expression and purification of GST-NotchIC and its mutants
Notch-GST fusion proteins were expressed in Escherichia coli BL21(DE3) and purified as previously described (35). Briefly, after mild sonication, cell debris was removed by centrifugation. The supernatant was loaded onto a Sepharose-glutathione affinity column, washed with solution A (0.5% Triton X-100 in 0.5 M NaCl and 10 mM Tris) and solution B (50 mM Tris (pH 8.0)), then eluted with 10 mM glutathione in 50 mM Tris (pH 8.0). Eluted proteins were aliquoted and stored at -20°C. The molecular mass and purity of each fusion protein were determined by 10% SDS-PAGE, followed by Coomassie blue staining.
Nuclear extracts and EMSA
Jurkat T cells were stimulated with PMA (50 ng/ml; Sigma, St.
Louis, MO) for 3 h. Nuclear extracts from stimulated Jurkat T
cells were prepared as described previously (35). Purified
GST-NotchIC or its derivatives (4 pmol each) were
added directly to DNA-protein binding reactions (10 mM Tris (pH 7.5),
10 mM EDTA, 0.5 mM DTT, 50 mM NaCl, 2 µg poly(dI:dC), 5% glycerol,
0.5 ng 32P-labeled NF-B oligonucleotides,
and 6 µg nuclear extracts) and kept at room temperature for 30 min.
The reaction products were run on 6% nondenaturing polyacrylamide gels
with 0.5x Tris-borate-EDTA buffer at room temperature. The gels were
dried and exposed to x-ray film at -70°C.
In vitro transcription and translation
Translation of full-length p50 and p65 was performed with in vitro transcription and translation system T7-coupled wheat-germ extract system in the presence of [35S]methionine according to the manufacturer’s protocols (Promega, Madison, WI).
Association of p50 with GST-NotchIC mutants
Purified GST-TAN-1 mutants were loaded on MicroSpin GST purification columns (Amersham, Indianapolis, IN) and incubated for 10 min. Columns were washed twice with PBS. [35S]Methionine-labeled p50 or p65 was added to each column and incubated at 4°C for 2 h. Columns were washed with PBS three times and eluted with 10 mM glutathione in PBS. Eluted samples were run on a 4–12% SDS-PAGE, followed by autoradiography.
Intracellular staining
Human NTera-2 embryonal carcinoma cells (American Type Culture Collection, Manassas, VA) were grown on Lab-Tek chamber slides (Nalge Nunc International, Naperville, IL) overnight, then transfected with LipofectAMINE (Life Technologies, Gaithersburg, MD) according to the manufacturer’s instructions. After 48 h, cells were permeabilized with 90% methanol at -20°C for 30 min, washed twice with PBS, twice with NGS (50% PBS (pH 7.0), 25% normal rabbit serum, 25% normal goat serum, and 0.1% azide), and then incubated with rabbit polyclonal anti-Notch serum as the first reagent (1/100 dilution) or normal rabbit serum (1/100 dilution) as a control at 37°C for 1 h. Cells were washed twice with NGS (4°C, 15 min/wash), then incubated at 37°C for 1 h with FITC-conjugated F(ab')2 of polyclonal goat anti-rabbit IgG (Caltag, South San Francisco, CA), followed by three washes at 37°C with NGS (15 min/wash). Cells were observed by epifluorescence microscopy.
Transient transfection and luciferase assay
Ntera-2 cells were cultured in DMEM containing 10% FCS and transfected with LipofectAMINE and the indicated plasmids for 48 h. Cells were lysed, and the lysates were clarified by centrifugation at 10,000 rpm for 5 min. An equal amount of supernatant from each lysate was assayed for luciferase activity using a kit from Promega and a Microtiter Plate Luminometer ML3000 with the Biolinx 2.10 program (Dynatech Laboratories, Chantilly, VA).
Western blot
Ntera-2 cells were transfected with expression vectors in a six-well plate in the presence of LipofectAMINE. After 48 h, the cells were lysed, and lysates were centrifuged. Supernatants were boiled with sample loading buffer under reducing conditions and then run on a 4–12% SDS-PAGE, followed by electrotransferring to polyvinylidene difluoride membrane. Membranes were blocked with 3% BSA-PBS for 1 h, and then sequentially incubated with rabbit anti-human Notch-1 Ab followed by alkaline phosphatase-conjugated anti-rabbit IgG. Bound Abs were detected in the presence of alkaline phosphatase substrate.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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B binding to DNA by NotchIC and its
mutants
We have previously shown that NotchIC
inhibited NF-B binding to DNA (35). To investigate
which domain(s) within Notch-1 was responsible for the interaction
between NotchIC and NF-B, we constructed a
series of NotchIC mutants, as described in Fig. 1
, and cloned into expression vector
pcDNA3. Additionally, NotchIC mutants were
expressed as GST fusion proteins in E. coli BL21(DE3) and
purified by Sepharose-glutathione affinity column as shown in Fig. 2A. GST fusion proteins of
NotchIC and its mutants migrated at the predicted
molecular masses on 10% SDS-PAGE. The effects of mutant proteins on
NF-B DNA binding were assessed by EMSA.
NotchIC completely inhibited NF-B binding to
DNA (Fig. 2B), in agreement with our previous report
(35). Interestingly, the six ankyrin repeats by themselves
showed no inhibitory effect on NF-B DNA binding activity. The
C-terminal region of NotchIC together with the
six ankyrin repeats showed weak inhibitory activity, whereas the
N-terminal region of NotchIC together with the
six ankyrin repeats abolished NF-B DNA binding. Interestingly, the
N-terminal portion of NotchIC without the six
ankyrin repeats also completely prevented NF-B binding to DNA (Fig. 2C). These data indicate that the N-terminal portion of
NotchIC is sufficient for inhibition of NF-B
binding to DNA in vitro, and this activity is independent of the
ankyrin repeats.
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We previously demonstrated that NotchIC
specifically inhibits DNA binding of p50 and directly interacts with
p50 subunit of NF-B but not that of p65 (35). To
determine the subunit specificity of NotchIC
mutants, EMSA was performed by using in vitro transcript and translated
p50 and p65. NotchIC, N + 6ANK, and N effectively
inhibited DNA binding of p50 dimers (Fig. 3A), but not that of p65
dimers (data not shown). The mutants without the N-terminal portion of
NotchIC, C + 6ANK and 6ANK, had no effect on p50 DNA binding.
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B binding to DNA in vitro, and this activity is
dependent on direct interaction of the N-terminal portion of
NotchIC with the NF-B p50 subunit.
Inhibition of NF-B mediated gene expression by
NotchIC and its mutants
After activation, NF-B migrates to the cell nucleus, where it
recognizes specific DNA elements and regulates gene expression. The
interaction between NF-B and inhibitory proteins of the IB family
occurs primarily in the cytoplasm, except for Bcl-3. To identify the
cellular compartment where NF-B interacts with Notch-1, we
investigated the subcellular location of transfected
NotchIC in NTera-2 cells by immunofluorescence.
As shown in Fig. 4, NotchIC immunoreactivity was localized in the
nuclei. This suggests that the primary site of interaction with NF-B
may be the nucleus.
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B-mediated gene expression. The basal level of luciferase
expression directed by six repeated B elements was very low. When
the cells were cotransfected with p50/p65, luciferase activity was
dramatically increased. As we previously reported,
NotchIC strongly inhibited p50/p65 induced
reporter gene expression. Construct N + 6ANK, which contains the
N-terminal region and the six ankyrin repeats, and N, which is 109 aa
long and contains only the N-terminal region of
NotchIC, strongly inhibited p50/p65-directed gene
expression (Fig. 5A). In
contrast, constructs C + 6ANK (which contains the six ankyrin repeats
and the C-terminal region of NotchIC) and 6ANK
(which contains only the six ankyrin repeats) had no effect on
p50/p65-directed gene expression. To exclude the possibility that
NotchIC mutants are not equally expressed,
parallel samples were lysed and assayed for
NotchIC expression by Western blot with a rabbit
polyclonal Ab generated against NotchIC protein.
As shown in Fig. 5B, all NotchIC
mutants were equally well expressed, and the lack of inhibition of
p50/p65-mediated gene expression by C + 6ANK and 6ANK is not because of
the absence of NotchIC mutant protein expression.
These data indicate that the N-terminal region of
NotchIC inhibits NF-B-mediated gene expression
independently of the ankyrin repeats.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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B DNA binding and
NF-B-mediated gene expression independently of the ankyrin repeats,
through a region N-terminal to these repeats, namely, aa 1773–1881.
This 109-aa region of Notch-1 has 98, 98, 77, and 70% identities with
the corresponding regions of rat, mouse, Xenopus, and
zebrafish Notch, respectively (Fig. 6).
It overlaps with the mouse RAM domain (aa 1751–1806), except for the
first 11 amino acids (RRQHGQLWFPE, 1751–1761) of the RAM domain.
Mutation analysis (4) demonstrated that these 11 amino
acids are essential for RBP-J binding to Notch, whereas they are
dispensable for NF-B binding, because the
NotchIC used in this study does not contain the
corresponding 11 amino acids of human Notch-1. Therefore, we conclude
that the domain we identified here is a novel protein-protein
interaction domain and designated it the NF-B binding domain (NBD).
NBD and RAM are two partially overlapping, but different, domains. NBD
contains one of the NLS. The NLS is highly conserved among these five
Notch proteins. The high level of identity among NBDs of these four
Notch proteins with that of human Notch-1 suggests that these four
Notch proteins may also function as nuclear IB molecules. NBD of
human Notch-1 has 48, 30, 31, and 29% identity with the corresponding
region of mouse Notch-3 (accession number Q61982), mouse Notch-4
(accession number P31695), Drosophila melanogaster
(accession number P07207), and human Notch-4 (accession number
AAC32288). The putative NLS seems conserved in these four proteins with
three or four basic amino acids. No significant similarity was found
between NBD of human Notch-1 with human Notch-2 and Notch-3.
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B inhibition in NF-B-directed gene expression contain 1)
the NBD region and 2) at least one NLS. Thus, a possible interpretation
of our data is that the six ankyrin repeats are unable to interfere
with NF-B after transfection due to the lack of an NLS. However, the
ankyrin repeats do not interfere with NF-B DNA binding in EMSA
assays. Additionally, the construct C + 6ANK does have an NLS in its
C-terminal region, but it does not inhibit NF-B-mediated gene
expression. Thus, the most likely interpretation of our data is that
the NBD domain and its NLS are both necessary and sufficient for
NF-B inhibition by NotchIC in our experimental
model. Together with our immunofluorescence data showing that
transfected NotchIC localizes to the nucleus,
these data support a working model in which
NotchIC interacts with the p50 subunit of NF-B
in the nucleus through the NBD and blocks the binding of p50/p65
heterodimer to NF-B sites, thereby interfering with NF-B-induced
transcriptional regulation.
The N-terminal portion of NotchIC contains the
high affinity binding site (RAM) required for
NotchIC interaction with CBF-1-RBP-J, the
putative major effector of Notch signaling (4, 5). Thus,
CBF-1 and NF-B interact with two partially overlapping domains of
Notch-1. It is not clear whether these interactions are mutually
exclusive. In this case the relative intracellular concentrations and
the intracellular compartmentalization of Notch-1, CBF-1, and NF-B
may dictate which complexes are predominantly formed within the
nucleus.
The intracellular subunit of Notch receptors appears to contain several
functional modules that interact with a variety of intracellular
targets (Fig. 7). The ankyrin repeats are
involved in binding the C. elegans chromatin-remodeling
factor EMB-5 (38) and the cytoplasmic Notch
effector Deltex (54, 55), which, in turn, regulate the
transcription factor E47 (56). Additionally, activated
Notch-1 inhibits the myogenic activity of the myocyte enhancer factor
2C by direct interaction of myocyte enhancer factor 2C with the ankyrin
repeat region of Notch (57). Kurooka et al.
(5) reported recently that the ankyrin repeats together
with C-terminal sequences are required for the interaction with histone
acetyltransferases PCAF and GCN5. The same region of Notch-1 appears to
be sufficient for its transforming activity (27). Other
Notch binding proteins include the orphan nuclear receptor Nur77
(33) and the c-Abl accessory protein Disabled
(58). Notch binding proteins that are suggested to be
mediators of Notch signaling, and their putative functions are
summarized in Fig. 6
. It is clear that Notch resides at the center of a
complex multipathway signaling network involved in decision processes
that affect cell proliferation, differentiation, migration, and
apoptosis in multiple cellular and organ systems.
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B-regulated gene expression
(35), while NF-B regulates Notch signaling through
inducing the expression of the Notch ligand Jagged-1 on the cell
surface (59) suggests that the cross-talk between Notch
and NF-B is bidirectional. The possible biological roles of this
cross-talk remain to be elucidated. In principle, this interaction
could be used in vivo to amplify the differentiation of two initially
equipotent cells into a ligand-expressing signaling cell and a
receptor-expressing receiving cell (see Fig. 8). This phenomenon is well known in
Drosophila development (1). In this
hypothetical model NF-B activation in a given cell triggers Jagged-1
expression, thus leading this cell to activate Notch in surrounding
cells. Jagged-1 activation is known to up-regulate Notch-1 expression
in Drosophila (60) and mammalian cells. In
turn, Notch-1 up-regulation would inhibit NF-B in the receiving
cells, thus down-regulating NF-B-inducible gene expression,
including Jagged-1 expression. Because individual cells often express
both Notch and Delta (61, 62, 63), such a feedback could also
be envisioned bidirectionally within the same subset of cells. Notch
and Delta may also act in cis as a cell-autonomous mechanism
for switching off NF-B-mediated gene expression after
activation.
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B-activating stimuli include cytokines, mitogens, bacteria, and
viruses. Recently, Sun et al. reported that protein kinase C-
is
required for TCR-induced activation of NF-B in mature T lymphocytes
(64). NF-B-responsive genes include chemokines (RANTES,
macrophage inflammatory protein-1, and monocyte chemoattractant
protein-1), cytokines (TNF-, IL-2, and TGF-2), growth factors
(GM-CSF), immunoreceptors (MHC II, CD25, and TCR -chain), adhesion
molecules (VCAM-1 and endothelial-leukocyte adhesion molecule-1),
oncogenes (c-myb and c-myc), apoptotic genes
(Fas/CD95 and Fas ligand), and viral genes (HIV-1 and CMV) (40, 50, 65, 66, 67). Thus, Notch signaling through its effect on NF-B
could play a role in regulating the expression of these genes and
viruses in vivo.
Like NF-B, Notch receptors are expressed in a wide range of
hemopoietic and immune system cells, including human
CD34+ hemopoietic stem cells
(68, 69, 70). Recent evidence suggests that Notch activation
in CD4 T cells may induce peripheral tolerance (71) and
cell death in monocytes (72). The possible roles of
Notch/NF-B cross-talk in the regulation of cell fate decisions
during hemopoiesis, inflammation, immune responses, viral infection,
and apoptosis warrant further investigation.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Drs. Jinhai Wang or Ennan Guan, Division of Therapeutic Proteins, Center for Biologics Evaluation and Research, Food and Drug Administration, National Institutes of Health, Building 29B, Room 4E12, HFM-541, 8800 Rockville Pike, Bethesda, MD 20892. E-mail addresses: wangj@cber.fda.gov or guan{at}cber.fda.gov
3 Current address: Cancer Immunology Program, Cardinal Bernardin Cancer Center, Loyola University Chicago, 2160 South First Avenue, Maywood, IL 60153.
4 Abbreviations used in this paper: NotchEC, extracellular domains of Notch proteins; NotchIC, intracellular regions of Notch receptors; NLS, nuclear localization signal; TAN-1, translocation-associated Notch homologue; NBD, NF-B binding domain; CBF, C promoter binding factor.
Received for publication August 21, 2000. Accepted for publication April 24, 2001.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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B kinases essential for NF-B activation. Science 278:860.B kinase complex (IKK) contains two kinase subunits, IKK and IKK, necessary for IB phosphorylation and NF-B activation. Cell 91:243.[Medline]
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-peptide generation and endoproteolysis of Notch. Proc. Natl. Acad. Sci. USA 97:5913.B can trigger the Notch signaling pathway by inducing the expression of Jagged1, a ligand for Notch receptors. EMBO J. 18:2803.[Medline]
is required for TCR-induced NF-B activation in mature but not immature T lymphocytes. Nature 404:402.[Medline]
B-dependent Fas ligand expression. Eur. J. Immunol. 29:2948.[Medline]
B mediates apoptosis through transcriptional activation of Fas (CD95) in adenoviral hepatitis. J. Biol. Chem. 275:6421.B family transactivate the murine c-myb gene. J. Biol. Chem. 270:7661.This article has been cited by other articles:
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 C. L. Curry, L. L. Reed, E. Broude, T. E. Golde, L. Miele, and K. E. Foreman Notch inhibition in Kaposi's sarcoma tumor cells leads to mitotic catastrophe through nuclear factor-{kappa}B signaling Mol. Cancer Ther., July 1, 2007; 6(7): 1983 - 1992. [Abstract] [Full Text] [PDF]
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 Y.-T. Ahn, B. Huang, L. McPherson, C. Clayberger, and A. M. Krensky Dynamic Interplay of Transcriptional Machinery and Chromatin Regulates "Late" Expression of the Chemokine RANTES in T Lymphocytes Mol. Cell. Biol., January 1, 2007; 27(1): 253 - 266. [Abstract] [Full Text] [PDF]
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 G. Weinmaster and R. Kopan A garden of Notch-ly delights Development, September 1, 2006; 133(17): 3277 - 3282. [Abstract] [Full Text] [PDF]
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P. M. Pegman, S. M. Smith, B. N. D'Souza, S. T. Loughran, S. Maier, B. Kempkes, P. A. Cahill, M. J. Simmons, C. Gelinas, and D. Walls Epstein-Barr Virus Nuclear Antigen 2 trans-Activates the Cellular Antiapoptotic bfl-1 Gene by a CBF1/RBPJ{kappa}-Dependent Pathway. J. Virol., August 1, 2006; 80(16): 8133 - 8144. [Abstract] [Full Text] [PDF]
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 E. Monsalve, M. A. Perez, A. Rubio, M. J. Ruiz-Hidalgo, V. Baladron, J. J. Garcia-Ramirez, J. C. Gomez, J. Laborda, and M. J. M. Diaz-Guerra Notch-1 Up-Regulation and Signaling following Macrophage Activation Modulates Gene Expression Patterns Known to Affect Antigen-Presenting Capacity and Cytotoxic Activity J. Immunol., May 1, 2006; 176(9): 5362 - 5373. [Abstract] [Full Text] [PDF]
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L. Pan, S. T. Glenn, C. A. Jones, and K. W. Gross Activation of the Rat Renin Promoter by HOXD10{middle dot}PBX1b{middle dot}PREP1, Ets-1, and the Intracellular Domain of Notch J. Biol. Chem., May 27, 2005; 280(21): 20860 - 20866. [Abstract] [Full Text] [PDF]
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C. M. Witt, V. Hurez, C. S. Swindle, Y. Hamada, and C. A. Klug Activated Notch2 Potentiates CD8 Lineage Maturation and Promotes the Selective Development of B1 B Cells Mol. Cell. Biol., December 1, 2003; 23(23): 8637 - 8650. [Abstract] [Full Text] [PDF]
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T.-S. Yeh, Y.-M. Lin, R.-H. Hsieh, and M.-J. Tseng Association of Transcription Factor YY1 with the High Molecular Weight Notch Complex Suppresses the Transactivation Activity of Notch J. Biol. Chem., October 24, 2003; 278(43): 41963 - 41969. [Abstract] [Full Text] [PDF]
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 L. Espinosa, J. Ingles-Esteve, A. Robert-Moreno, and A. Bigas Ikappa Balpha and p65 Regulate the Cytoplasmic Shuttling of Nuclear Corepressors: Cross-talk between Notch and NFkappa B Pathways Mol. Biol. Cell, February 1, 2003; 14(2): 491 - 502. [Abstract] [Full Text] [PDF]
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J. Chu, S. Jeffries, J. E. Norton, A. J. Capobianco, and E. H. Bresnick Repression of Activator Protein-1-mediated Transcriptional Activation by the Notch-1 Intracellular Domain J. Biol. Chem., February 22, 2002; 277(9): 7587 - 7597. [Abstract] [Full Text] [PDF]
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P. Cheng, A. Zlobin, V. Volgina, S. Gottipati, B. Osborne, E. J. Simel, L. Miele, and D. I. Gabrilovich Notch-1 Regulates NF-{kappa}B Activity in Hemopoietic Progenitor Cells J. Immunol., October 15, 2001; 167(8): 4458 - 4467. [Abstract] [Full Text] [PDF]
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