CBFB HUman
Description
Molecular Structure and Function of CBFB
CBFB functions as the non-DNA-binding β-subunit of the core-binding factor (CBF) complex, which partners with RUNX proteins (RUNX1, RUNX2, RUNX3) to regulate gene expression . Key features include:
Table 1: CBFB Characteristics
Core Binding Factor Acute Myeloid Leukemia (CBF-AML)
Chromosomal rearrangements such as inv(16)(p13q22) or t(16;16) fuse CBFB with MYH11, producing the oncogenic CBFβ-MYH11 protein . This fusion:
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Disrupts hematopoiesis: Sequesters RUNX1 in cytoplasmic aggregates, blocking myeloid differentiation .
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Requires secondary mutations: Additional genetic hits (e.g., FLT3, KIT) are necessary for leukemogenesis .
CBF-AML accounts for 5–8% of adult AML cases and is associated with eosinophilia .
Breast Cancer
CBFB is highly mutated in breast tumors, with loss-of-function mutations linked to:
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Loss of translational regulation: CBFB stabilizes RUNX1 mRNA and facilitates translation via eIF4B .
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Oncogenic transformation: CRISPR-Cas9 knockout in MCF10A cells induced tumorigenesis in xenograft models .
Cleidocranial Dysplasia (CCD)-Like Syndrome
Heterozygous CBFB pathogenic variants (e.g., nonsense, frameshift) cause skeletal abnormalities resembling CCD, including:
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Dental anomalies: Supernumerary teeth, delayed eruption .
Unlike RUNX2-related CCD, patients exhibit normal stature and neurocognitive deficits .
Dual Role in Transcription and Translation
CBFB regulates both nuclear transcription and cytoplasmic translation:
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Nuclear: Partners with RUNX1/2/3 to activate hematopoietic and osteogenic genes .
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Cytoplasmic: Binds mRNA via hnRNPK to enhance translation initiation of RUNX1 and other targets .
Therapeutic Targeting in AML
Recent advances include:
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Immunotherapy: Engineered T cells targeting CBFβ-MYH11 fusion peptides show efficacy in xenograft models, reducing tumor burden by >90% .
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Mechanistic insights: CBFβ-MYH11 disrupts CBF/RUNX1 nuclear localization, offering a pathway for small-molecule inhibitors .
Table 2: Clinically Significant CBFB Mutations
Future Directions
Research priorities include:
Product Specs
ASWQGEQRQT PSREYVDLER EAGKVYLKAP MILNGVCVIW KGWIDLQRLD GMGCLEFDEE RAQQEDALAQ QAFEEARRRT REFEDRDRSH
REEMEVRVSQ LLAVTGKKTT RP.
Q&A
What is CBFB and what are its key characteristics?
CBFB (Core Binding Factor Beta) is a widely-expressed 21-24 kDa protein that belongs to the CBFB family. It forms heterodimeric transcription factor complexes with RUNX proteins (RUNX1, RUNX2, RUNX3) . The human CBFB protein spans from Pro2 to Glu165 (Accession # Q13951) . Unlike typical transcription factors that function exclusively in the nucleus, CBFB has been discovered to have dual localization and function - regulating transcription in the nucleus and translation in the cytoplasm . This dual functionality makes CBFB unique among its class of proteins and suggests complex regulatory roles in cellular processes.
How can researchers detect CBFB in experimental systems?
CBFB can be detected through various experimental approaches:
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Western Blot Analysis: Using specific antibodies such as Sheep Anti-Human CBFB Antigen Affinity-purified Polyclonal Antibody. In western blots, CBFB appears as a specific band at approximately 22 kDa. Experiments should be conducted under reducing conditions using appropriate immunoblot buffers .
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Immunofluorescence: CBFB can be visualized in fixed cells using specific primary antibodies (e.g., at 10 μg/mL concentration) followed by fluorophore-conjugated secondary antibodies. Counterstaining with DAPI helps identify nuclei. This technique has revealed that CBFB localizes to both cytoplasm and nuclei in human cells .
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RNA Immunoprecipitation (RIP): For studying CBFB's interaction with mRNAs, RIP-seq has been successfully employed to identify transcripts bound by cytoplasmic CBFB .
How does CBFB function in transcriptional regulation?
CBFB does not directly bind to DNA but instead forms heterodimeric complexes with RUNX family proteins. These complexes function as transcription factors that regulate gene expression in various biological processes . The CBFB/RUNX1 complex specifically has been shown to transcriptionally repress the oncogenic NOTCH signaling pathway in breast cancer cells . This repressive function contributes to CBFB's role as a tumor suppressor in breast cancer.
Methodologically, ChIP-seq (Chromatin Immunoprecipitation followed by sequencing) experiments can be employed to identify genomic regions bound by CBFB/RUNX complexes, while RNA-seq analysis of cells with CBFB knockdown or knockout can reveal genes whose expression is regulated by CBFB-containing transcription complexes .
What is the relationship between CBFB and RUNX proteins?
CBFB can interact with all three RUNX proteins (RUNX1, RUNX2, RUNX3) to form heterodimeric transcription factors . This ability to partner with different RUNX proteins explains why CBFB mutations can affect multiple biological systems. For example:
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CBFB/RUNX1 complexes regulate hematopoiesis and are implicated in leukemia
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CBFB/RUNX2 interactions influence skeletal development, with CBFB mutations causing conditions resembling cleidocranial dysplasia (typically associated with RUNX2 mutations)
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The interaction with multiple RUNX proteins may explain phenotypic differences between CBFB-related and RUNX2-related skeletal disorders
How does CBFB regulate mRNA translation?
Recent research has revealed an unexpected function of CBFB in translation regulation. The cytoplasmic pool of CBFB:
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Binds to hundreds of mRNA transcripts via hnRNPK, an RNA-binding protein
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Enhances translation through interaction with eIF4B, a general translation initiation factor
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Specifically regulates translation of RUNX1 mRNA, demonstrating a feedback mechanism between transcriptional and translational regulation
This mechanism was discovered through RIP-seq (RNA Immunoprecipitation followed by sequencing), which identified CBFB-bound transcripts, and subsequent functional studies validated the translational impact .
What is the molecular mechanism of CBFB-mediated translation enhancement?
CBFB does not directly bind to mRNAs but rather interacts with hnRNPK, which directly binds to the transcripts. Experiments using recombinant proteins and RNA pull-down assays (RPA) demonstrated that while CBFB alone does not interact with mRNAs like RUNX1, it significantly enhances the binding of hnRNPK to these transcripts .
The binding specificity appears to be determined by a gapped motif containing poly-C sequences, which was identified in 86% of hnRNPK-bound transcripts using the GLAM2 algorithm . Once bound to mRNAs via hnRNPK, CBFB facilitates translation initiation through eIF4B recruitment, enhancing protein synthesis from these transcripts .
How is CBFB implicated in breast cancer?
Genome-wide sequencing studies have revealed that CBFB is highly mutated in human breast tumors, suggesting a critical role in breast cancer etiology . Research indicates that CBFB functions as a tumor suppressor in breast cancer through multiple mechanisms:
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In the nucleus, the CBFB/RUNX1 complex transcriptionally represses the oncogenic NOTCH signaling pathway
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In the cytoplasm, CBFB regulates the translation of numerous mRNAs that may include tumor suppressors
The dual functionality of CBFB suggests that breast cancer cells can evade both transcriptional and translational surveillance simultaneously through CBFB downregulation . This makes CBFB particularly important as a tumor suppressor, as its loss impacts multiple cellular processes simultaneously.
What experimental approaches are used to study CBFB in cancer?
Researchers investigating CBFB's role in cancer utilize several approaches:
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Genomic Analysis: Identifying mutations in CBFB in patient tumor samples
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Functional Studies: Knockdown or knockout of CBFB in cell lines to assess effects on proliferation, migration, and invasion
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Mechanistic Studies:
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Pathway Analysis: Examining effects of CBFB manipulation on specific signaling pathways like NOTCH
What role does CBFB play in male fertility and spermatogenesis?
CBFB has been identified as a critical regulator of male germline development and spermatogenesis. Using conditional knockout (cKO) mouse models, researchers have demonstrated that:
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Inactivation of Cbfb in the male germline results in rapid degeneration of the germline during the onset of spermatogenesis, impaired sperm production, and adult infertility
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CBFB function appears to be limited to undifferentiated spermatogonia despite its expression in other germ cell types
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Within undifferentiated spermatogonia, CBFB regulates proliferation, survival, and maintenance of the undifferentiated spermatogonia population
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CBFB also distally regulates meiotic progression and spermatid formation
Spatial transcriptomics revealed that CBFB modulates cell cycle checkpoint control genes associated with both proliferation and meiosis, suggesting that core programs established within prepubertal undifferentiated spermatogonia are necessary for both germline maintenance and sperm production .
How is CBFB involved in skeletal development?
Heterozygous pathogenic variants in CBFB have been found to cause a skeletal disorder resembling cleidocranial dysplasia (CCD) . This condition shares similarities with RUNX2-related CCD, including dental and clavicular abnormalities, but also presents distinguishing features such as normal stature and neurocognitive problems .
Several types of CBFB variants have been identified in affected individuals:
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Splice site alterations
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Nonsense variants
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Small duplications resulting in premature stop codons
How does the CBFB-MYH11 fusion contribute to leukemogenesis?
The CBFB-MYH11 fusion gene, generated by inversion of chromosome 16 in human acute myeloid leukemia (AML), is causative for oncogenic transformation . While the fusion was traditionally thought to act primarily through dominant inhibition of RUNX1 and CBFB functions, research has revealed more complex mechanisms:
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CBFB-MYH11 causes defects in both primitive and definitive hematopoiesis that are independent of Cbfb/Runx1 repression
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During primitive hematopoiesis, CBFB-MYH11 delays differentiation, characterized by sustained expression of genes like Gata2, Il1rl1, and Csf2rb - a phenotype not observed in Cbfb and Runx1 knockout mice
This suggests that CBFB-MYH11 has gain-of-function properties beyond simply inhibiting normal CBFB or RUNX1 activity, which contributes to its leukemogenic potential.
What experimental models are available to study CBFB in hematopoiesis?
Researchers investigating CBFB in hematopoiesis employ several model systems:
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Conditional Knockout Mouse Models: Using tissue-specific Cre recombinase expression to delete Cbfb in specific lineages or developmental stages
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Knock-in Models: Introducing specific mutations or fusion genes (like CBFB-MYH11) to study their effects on hematopoiesis
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Cell Line Models: Human leukemia cell lines like Jurkat (T-cell leukemia) and M1 (mouse myeloid leukemia) express CBFB and can be used for mechanistic studies
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Primary Cell Cultures: Isolated hematopoietic stem and progenitor cells can be manipulated in vitro to study CBFB function
What antibodies and detection methods are optimal for CBFB research?
Researchers have successfully used several antibodies and detection methods for CBFB:
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For Western Blot: Sheep Anti-Human CBFB Antigen Affinity-purified Polyclonal Antibody (e.g., R&D Systems Catalog # AF7349) at 1 μg/mL concentration, followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody . Western blots should be conducted under reducing conditions using appropriate immunoblot buffers.
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For Immunofluorescence: The same primary antibody can be used at 10 μg/mL for 3 hours at room temperature, followed by fluorophore-conjugated secondary antibodies such as NorthernLights 557-conjugated Anti-Sheep IgG . DAPI counterstaining helps identify nuclei and confirm subcellular localization.
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For RIP-seq: Antibodies suitable for immunoprecipitation of CBFB-RNA complexes are essential for studying the translational regulatory functions of CBFB .
How can researchers effectively study CBFB-RNA interactions?
To investigate CBFB's interaction with mRNAs, several approaches have proven effective:
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RNA Immunoprecipitation (RIP): Immunoprecipitate CBFB from cytoplasmic extracts and identify bound RNAs by sequencing
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RNA Pull-down Assays (RPA): Using recombinant CBFB and hnRNPK proteins with biotin-labeled RNA to determine direct binding interactions
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Motif Analysis: Tools like the GLAM2 algorithm can identify RNA sequence motifs recognized by CBFB-hnRNPK complexes
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Polysome Profiling: To assess the impact of CBFB on translation efficiency of specific mRNAs by analyzing their distribution across polysome fractions
What are current challenges in understanding CBFB function?
Several aspects of CBFB biology remain incompletely understood:
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Integration of Dual Functions: How cells coordinate the nuclear (transcriptional) and cytoplasmic (translational) functions of CBFB and how these functions influence each other
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Tissue-Specific Effects: Understanding why CBFB mutations affect some tissues more severely than others despite CBFB's widespread expression
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Therapeutic Targeting: Developing strategies to restore CBFB function in cancers where it is downregulated or to inhibit oncogenic CBFB fusions like CBFB-MYH11
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Partner-Dependent Functions: Delineating which functions of CBFB are dependent on specific partners (RUNX proteins, hnRNPK, eIF4B) and which might be independent
What emerging technologies could advance CBFB research?
Several cutting-edge approaches hold promise for future CBFB research:
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Spatial Transcriptomics: Already being applied to study CBFB in spermatogenesis, this approach can provide insights into the spatial distribution of CBFB-regulated gene expression in other tissues
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Single-Cell Technologies: Single-cell RNA-seq and ATAC-seq could reveal cell-type-specific functions of CBFB and heterogeneity in CBFB activity
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Proteomics Approaches: Mass spectrometry-based interactome studies could identify novel CBFB-interacting proteins in different cellular compartments
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CRISPR-Based Screening: Genome-wide or focused CRISPR screens could identify synthetic lethal interactions with CBFB mutations or dependencies in CBFB-mutant cancers
Product Science Overview
Core Binding Factor Beta (CBFβ) is a crucial component of the core-binding transcription factor complex, which plays a significant role in the regulation of gene expression related to hematopoiesis and osteogenesis. The human recombinant form of CBFβ is a synthesized version of this protein, produced through recombinant DNA technology, and is used extensively in research and therapeutic applications.
CBFβ is the beta subunit of the heterodimeric core-binding transcription factor, which also includes an alpha subunit (RUNX1, RUNX2, or RUNX3). Unlike the alpha subunit, CBFβ does not bind directly to DNA. Instead, it enhances the DNA-binding affinity of the alpha subunit, thereby facilitating the transcription of target genes .
CBFβ is involved in the regulation of various genes essential for hematopoiesis (the formation of blood cellular components) and osteogenesis (bone formation). It interacts with the alpha subunit to bind to the core site of various enhancers and promoters, including those of the murine leukemia virus, polyomavirus enhancer, T-cell receptor enhancers, and GM-CSF promoters .
Mutations and chromosomal rearrangements involving the CBFB gene are associated with several diseases. For instance, a pericentric inversion of chromosome 16 [inv (16) (p13q22)] results in a chimeric transcript that fuses the N terminus of CBFβ with the C-terminal portion of the smooth muscle myosin heavy chain 11. This rearrangement is commonly associated with acute myeloid leukemia (AML) of the M4Eo subtype .
The human recombinant form of CBFβ is produced using recombinant DNA technology. This involves inserting the CBFB gene into an expression vector, which is then introduced into a host cell (such as E. coli or yeast). The host cells are cultured, and the recombinant protein is expressed, harvested, and purified for use in research and therapeutic applications.
CBFβ, as part of the core-binding factor complex, participates in various biochemical interactions. It allosterically enhances the DNA-binding capability of the alpha subunit, allowing the complex to bind to specific DNA sequences and regulate gene transcription. Analytical techniques such as electrophoretic mobility shift assays (EMSAs), chromatin immunoprecipitation (ChIP), and reporter assays are commonly used to study these interactions and the regulatory mechanisms of CBFβ.
The activity of CBFβ is regulated through its interaction with the alpha subunit and other co-factors. Post-translational modifications, such as phosphorylation, can also influence its function. Additionally, the expression of CBFβ is tightly controlled at the transcriptional level, ensuring that it is produced in the right amounts and at the right times during cellular differentiation and development.
