Alternative titles; symbols
HGNC Approved Gene Symbol: RBPJ
Cytogenetic location: 4p15.2 Genomic coordinates (GRCh38) : 4:26,105,449-26,435,131 (from NCBI)
| Location | Phenotype |
Phenotype MIM number |
Inheritance |
Phenotype mapping key |
|---|---|---|---|---|
| 4p15.2 | Adams-Oliver syndrome 3 | 614814 | Autosomal dominant | 3 |
The vast diversity of specificities in antigen recognition is primarily generated by the somatic combinatorial assembly of multiple germline DNA segments called variable (V), diversity (D), and joining (J) in immunoglobulin and T-cell receptor genes. Each segment of V, D, and J is flanked by a recombination-recognition sequence (RS) consisting of a palindromic heptanucleotide sequence and an AT-rich nonameric sequence separated by either 12 or 23 bp of spacer sequence. The heptamer and nonamer sequences are highly conserved during evolution, whereas the spacer sequences are not. Several lines of evidence indicate that the V(D)J recombination processes are directed by the recognition of the RS. The genes involved in V(D)J recombination include RAG1 (179615) and RAG2 (179616) in the mouse. The Igkjrb protein, identified in nuclear extracts from a mouse pre-B-cell line as a binding protein specific to Jk-type RS, may also be involved. The IGKJRB gene is well conserved among many species such as human, mouse, Xenopus, and Drosophila. Using cDNA fragments of the mouse Igkjrb, Amakawa et al. (1993) isolated its human counterpart, IGKJRB. Three types of cDNA with different 5-prime sequences were isolated, suggesting the presence of 3 protein isoforms. Two of these may be specific to human cells because the mouse counterparts were not detected. The amino acid sequences of the human and mouse genes are 98% identical in exons 2 through 11, whereas the homology of the human and mouse exon 1 sequences were 75%. A possible wider role for the RBP-Jk protein is suggested by the fact that in Drosophila the homologous gene is involved in the development of the peripheral nervous system.
Gross (2011) mapped the RBPJ gene to chromosome 4p15.2 based on an alignment of the RBPJ sequence (GenBank AK302230) with the genomic sequence (GRCh37).
Pseudogenes
By fluorescence in situ hybridization (FISH), Amakawa et al. (1993) demonstrated that the functional IGKJRB gene is localized at 3q25 and the 2 pseudogenes at 9p13 and 9q13. (The 2 pseudogenes, located on different arms of chromosome 9, showed 91% homology with the cDNA for 1 of the 3 IGKJRB proteins. The processed pseudogenes differ from each other. Amakawa et al. (1993) suggested the following evolutionary mechanism for their generation: the primordial pseudogene evolved from mRNA which was incorporated into a region at chromosome 9p13. The next processes could be recent events, including duplication of the primordial pseudogene, chromosomal inversion involving 9p13 and 9q13, and subsequent divergence by point mutations, nucleotide insertions, and deletions. Examination of peripheral lymphocytes by Southern blot analysis demonstrated that the genomes of 8 out of 22 individuals lacked pseudogene 2 located at 9q13.) By FISH, Tang et al. (1997) failed to find a locus for the functional gene on 3q25 and found instead that the functional gene was localized to 9p13-p12 and 9q13, 2 sites where pseudogenes had previously been assigned by Amakawa et al. (1993).
Henkel et al. (1994) determined that CBF1 is identical to IGKJRB1. Christensen et al. (1996) found that the product of the worm Lag1 gene, like CBF1 and the Drosophila homolog 'suppressor of hairless,' binds specifically to the consensus DNA sequence RTGGGAA (see also Tun et al. (1994)). This family of DNA-binding factors that mediate transcriptional activation or repression is designated CSL.
RBPJ functions immediately downstream of Notch signaling (see 190198). Han et al. (2002) used a conditional gene knockout strategy to inactivate the DNA-binding domain of Rbpj in mouse bone marrow and found that Rbpj was required for T-cell development. In the absence of Rbpj, there was an increase in thymic B-cell development. Han et al. (2002) proposed that RBPJ, by mediating Notch signaling, controls T- versus B-cell fate decisions in lymphoid progenitors.
Thymocytes can be divided into 4 subsets based on CD4 (186940) and CD8 (see 186910) expression, with double-negative (DN) cells being the least mature. The DN population can be further subdivided into 4 subsets, DN1 through DN4. Tanigaki et al. (2004) used a conditional knockout strategy to inactivate Rbpj at the DN2 and DN4 stages in mice. Inactivation at DN2 resulted in severe developmental arrest of alpha-beta T cells at the DN3 stage and enhanced generation of gamma-delta T cells. Inactivation at DN4 caused no abnormalities in CD4/CD8 lineage commitment, but it resulted in enhanced Th1 responses and reduced T-cell proliferation. Tanigaki et al. (2004) concluded that Notch/RBPJ signaling regulates not only the T-cell developmental process, but also the direction and magnitude of immune responses via regulation of peripheral T cells.
Van Es et al. (2005) showed a rapid, massive conversion of proliferative crypt cells into postmitotic goblet cells after conditional removal of the common Notch pathway transcription factor CSL/RBP-J. The authors obtained a similar phenotype by blocking the Notch cascade with a gamma-secretase inhibitor. The inhibitor also induced goblet cell differentiation in adenomas in mice carrying a mutation of the Apc tumor suppressor gene (611731). Thus, maintenance of undifferentiated, proliferative cells in crypts and adenomas requires the concerted activation of the Notch and Wnt cascades.
Siekmann and Lawson (2007) demonstrated that Notch signaling is necessary to restrict angiogenic cell behavior to tip cells in developing segmental arteries in the zebrafish embryo. In the absence of the Notch signaling component Rbpsuh, Siekmann and Lawson (2007) observed excessive sprouting of segmental arteries, whereas Notch activation suppressed angiogenesis. Through mosaic analysis they found that cells lacking Rbpsuh preferentially localized to the terminal position in developing sprouts. In contrast, cells in which Notch signaling had been activated were excluded from the tip cell position. In vivo time-lapse analysis revealed that endothelial tip cells undergo a stereotypical pattern of proliferation and migration during sprouting. In the absence of Notch, nearly all sprouting endothelial cells exhibited tip cell behavior, leading to excessive numbers of cells within segmental arteries. Furthermore, Siekmann and Lawson (2007) found that Flt4 (136352) is expressed in segmental artery tip cells and became ectopically expressed throughout the sprout in the absence of Notch. Loss of Flt4 could partially restore normal endothelial cell number in Rbpsuh-deficient segmental arteries. Finally, loss of the Notch ligand Dll4 (605185) also led to an increased number of endothelial cells within segmental arteries. Siekmann and Lawson (2007) concluded that their studies taken together indicated that proper specification of cell identity, position, and behavior in a developing blood vessel sprout is required for normal angiogenesis, and implicated the Notch signaling pathway in this process.
PTF1A (607194) is a basic helix-loop-helix transcription factor required for pancreatic development. Masui et al. (2007) found that Ptf1a interacted with Rbpj within a stable trimeric DNA-binding complex (PTF1) during early pancreatic development in mouse. As acinar cell development began, Rbpj was swapped for Rbpjl, the constitutively active, pancreas-restricted Rbpj paralog, and Rbpjl was a direct target of the PTF1 complex. At the onset of acinar cell development, when the Rbpjl gene was first induced, a PTF1 complex containing Rbpj bound to the Rbpjl promoter. As development proceeded, Rbpjl gradually replaced Rbpj in the PTF1 complex bound to the Rbpjl promoter and appeared on the PTF1 complex-binding sites on the promoters of other acinar-specific genes, including those for secretory digestive enzymes. Introduction of a Ptf1a mutant unable to bind Rbpj truncated pancreatic development at an immature stage, without the formation of acini or islets. The action of Rbpj within the PTF1 complex was independent of its role in Notch signaling.
Mizutani et al. (2007) showed that both neural stem cells and intermediate neural progenitors respond to Notch receptor activation, but that neural stem cells signal through the canonic Notch effector CBF1, whereas intermediate neural progenitors have attenuated CBF1 signaling. Furthermore, whereas knockdown of CBF1 promotes the conversion of neural stem cells to intermediate neural progenitors, activation of CBF1 is insufficient to convert intermediate neural progenitors back to neural stem cells. Using both transgenic and transient in vivo reporter assays, Mizutani et al. (2007) showed that neural stem cells and intermediate neural progenitors coexist in the telencephalic ventricular zone of mice and that they can be prospectively separated on the basis of CBF1 activity. Furthermore, using in vivo transplantation, they showed that whereas neural stem cells generate neurons, astrocytes, and oligodendrocytes at similar frequencies, intermediate neural progenitors are predominantly neurogenic. Mizutani et al. (2007) concluded that their study, together with previous work on hematopoietic stem cells, suggested the use or blockade of the CBF1 cascade downstream of Notch as a general feature distinguishing stem cells from more limited progenitors in a variety of tissues.
Notch signaling regulates gene expression for specification of cell fate in diverse tissues during development and adult tissue renewal. In response to ligand binding, the intracellular domain (ICD) of Notch is proteolytically released by the gamma-secretase complex (see 104311) and translocates to the nucleus, where it binds CSL and triggers its conversion from a repressor to an activator of Notch target gene expression. Engel et al. (2010) found that Mtg16 (CBFA2T3; 603870) -/- mouse hematopoietic progenitor cells showed elevated expression of Notch targets, in addition to impaired differentiation, in response to Notch signaling. The defect was reversed by restoration of Mtg16 expression. Using mouse and human cells, Engel et al. (2010) showed that all MTG family proteins bound CSL and that MTG16 bound the ICDs of all Notch receptor proteins. Binding of MTG16 to Notch ICD disrupted MTG16-CSL and MTG16-NCOR (see 600849) interactions and permitted Notch signaling. Mutation and coprecipitation analysis revealed that the N-terminal PST region of MTG16 interacted directly with Notch ICD and that binding was independent of the MTG16 NTR domains required for DNA, CSL, and histone deacetylase binding. The PST region of Mtg16 was also essential for Mtg16-dependent lineage specification in mouse hematopoietic progenitor cells. Engel et al. (2010) concluded that MTG16 is an integral component of Notch signaling that contributes to basal repression of canonical Notch target genes.
Tong et al. (2011) found that expression of human BOAT1 (ATXN1L; 614301) in Drosophila wing disrupted Notch signaling, leading to wing defects. Coimmunoprecipitation analysis of HEK293 cells revealed that both BOAT1 and ATXN1 (601556) precipitated CBF1, which functions as a transcriptional activator when associated with NICD. Protein pull-down and yeast 2-hybrid analyses confirmed the interactions and showed that BOAT1 and ATXN1 competed for CBF1 binding. Coimmunoprecipitation experiments showed that NICD disrupted CBF1-BOAT/ATXN1 interactions. Reporter gene assays revealed that both BOAT1 and ATXN1 inhibited CBF1 activity at the promoter for HEY1 (602953), a Notch target gene. Chromatin immunoprecipitation assays showed that Boat1 and Atxn1, in addition to Smrt (NCOR2; 600848), occupied the Hey1 promoter in differentiating mouse C2C12 myoblasts. Atxn1 bound the Hey1 promoter transiently, whereas Boat1 and Smrt remained bound to the Hey1 promoter under the same conditions. Tong et al. (2011) concluded that BOAT1 and ATXN1 are chromatin-binding factors that repress Notch signaling in the absence of NCID by acting as CBF1 corepressors.
The COX4I2 gene (607976) contains a conserved oxygen response element (ORE) that is maximally active at a concentration of 4% oxygen. Using a yeast 1-hybrid screen to identify transcription factors binding the 13-bp ORE of human COX4I2, followed by DNA binding assays, Aras et al. (2013) detected binding by CHCHD2 (616244), CXXC5 (612752), and RBPJ, but not by HIF1A (603348). Luciferase analysis showed that RBPJ and CHCHD2 functioned as activators of the ORE, whereas CXXC5 repressed it. Coimmunoprecipitation analysis showed that RBPJ interacted with both CHCHD2 and CXXC5. Treatment of rat primary lung cells with small interfering RNA to Chchd2 or Rbpj resulted in a significant decrease in Cox4i2 expression.
By immunohistochemical analysis, Kulic et al. (2015) observed frequent depletion of RBPJ in human breast tumors, confirming microarray data. Implantation of human tumor cells after RBPJ depletion into immunodeficient mice resulted in enhanced tumor growth. Knockdown of RBPJ caused significant upregulation of NOTCH target genes, such as HEY1, HES1 (139605), SNAIL1 (SNAI1; 604238), and GUCY1A3 (139396), as well as MMP1 (120353), suggesting that RBPJ deficiency results in a NOTCH-like gene signature with RBPJ activating a subset of NOTCH target genes. RBPJ depletion caused epigenetic changes corresponding to promoter activity, as shown by EMSA, chromatin immunoprecipitation, and real-time quantitative PCR. Functional studies showed that RBPJ deficiency increased tumor cell survival, possibly, by enabling MYC (190080) and NFKB (see 164011) activation. Kulic et al. (2015) concluded that loss of RBPJ derepresses target gene promoters, allowing NOTCH-independent activation by alternate transcription factors that promote tumorigenesis.
By yeast 2-hybrid, coimmunoprecipitation, and pull-down analyses, Xu et al. (2017) found that human L3MBTL3 (618844) interacted directly with RBPJ. Mutation analysis showed that the interaction required the N-terminal region of L3MBTL3 and the beta-trefoil domain (BTD) of RBPJ. The Notch ICD also interacted with the BTD of RBPJ, allowing competition between L3MBTL3 and the Notch ICD for RBPJ binding. Thermodynamic analysis showed that the Notch ICD could outcompete L3MBTL3 for binding to RBPJ. However, in the absence of Notch signaling, interaction with L3MBTL3 allowed RBPJ to recruit L3MBTL3 on chromatin to repress expression of Notch target genes. L3MBTL3 also interacted with KDM1A (609132), a histone demethylase, and linked KDM1A to Notch-responsive elements. KDM1A interacted with RBPJ and promoted demethylation of dimethylated lys4 of histone H3 (see 602810), resulting in repression of Notch target gene expression. Genetic analysis in Drosophila and C. elegans demonstrated that the RBPJ-L3MBTL3 interaction was evolutionarily conserved in metazoans.
Using a variant-filtering strategy to perform exome resequencing in 2 unrelated families with Adams-Oliver syndrome (AOS3; 614814), Hassed et al. (2012) identified 2 different heterozygous missense mutations in the RBPJ gene (147183.0001 and 147183.0002) that segregated with disease in each family. Functional analysis confirmed impaired DNA binding of mutant RBPJ.
Amakawa et al. (1993) demonstrated that the functional IGKJRB gene contains 13 exons and spans at least 67 kb. The human genome contains 1 functional IGKJRB gene and 2 types of processed pseudogenes.
RBPSUH and DLL4 (605185) are both involved in Notch signaling. Krebs et al. (2004) showed that Dll4 haploinsufficiency or Rbpsuh knockout in mice resulted in severe vascular defects leading to embryonic lethality. Rbpsuh -/- embryos did not express several arterial-specific endothelial cell markers. Conditional inactivation of Rbpsuh function demonstrated that Notch activation was essential in the endothelial cell lineage. Dll4 and Rbpsuh mutant embryos also exhibited arteriovenous malformations, likely due to an inability to establish and maintain distinct arterial-venous vascular beds.
Wang et al. (2016) found that after fractures, mice with a conditional deletion of Rbpjk in skeletal progenitor cells had persistent callus formation along the periosteum without bridging between the cortices. Histologic analysis and assessment of mechanical competence indicated both delayed fracture repair and complete nonunion of fractured bone. Immunofluorescence microscopy demonstrated depletion of bone marrow stromal/stem cells (BMSCs) with altered differentiation potential rather than altered vascularization or osteoclast numbers in Rbpjk mutant mice. Wang et al. (2016) concluded that NOTCH signaling and BMSCs are required for fracture repair, irrespective of stability and vascularity.
In a father and daughter with Adams-Oliver syndrome (AOS3; 614814), Hassed et al. (2012) identified heterozygosity for a 188A-G transition in the RBPJ gene, resulting in a glu63-to-gly (E63G) substitution in the highly conserved DNA-binding domain. Functional analysis using an oligonucleotide corresponding to a canonical RBPJ binding site in the promoter of HES1 (139605) demonstrated that whereas wildtype RBPJ formed a specific complex with the probe, the E63G mutant did not exhibit any specific binding complex; similarly, in live cells, the E63G mutant showed decreased binding to the HES1 promoter compared to wildtype RBPJ.
In affected individuals from a 3-generation family with Adams-Oliver syndrome (AOS3; 614814), Hassed et al. (2012) identified heterozygosity for a 505A-G transition in the RBPJ gene, resulting in a lys169-to-glu (K169E) substitution in the highly conserved DNA-binding domain. Functional analysis using an oligonucleotide corresponding to a canonical RBPJ binding site in the promoter of HES1 (139605) demonstrated that whereas wildtype RBPJ formed a specific complex with the probe, the K169E mutant did not exhibit any specific binding complex; similarly, in live cells, the K169E mutant showed decreased binding to the HES1 promoter compared to wildtype RBPJ.
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Gross, M. B. Personal Communication. Baltimore, Md. 2/3/2011.
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Xu, T., Park, S.-S., Giaimo, B. D., Hall, D., Ferrante, F., Ho, D. M., Hori, K., Anhezini, L., Ertl, I., Bartkuhn, M., Zhang, H., Milon, E., and 15 others. RBPJ/CBF1 interacts with L3MBTL3/MBT1 to promote repression of Notch signaling via histone demethylase KDM1A/LSD1. EMBO J. 36: 3232-3249, 2017. [PubMed: 29030483] [Full Text: https://doi.org/10.15252/embj.201796525]