GO:0044666 MLL3/4 complex: Components, Assembly and Research Methods, Genes, Functions and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:0044666 (MLL3/4 complex) is a histone H3 lysine-4 (H3K4) methyltransferase complex defined by the presence of either MLL3 (KMT2C) or MLL4 (KMT2D) as its catalytic subunit.
• The complex is also known as the Trr/COMPASS-like complex and is conserved from Drosophila to mammals.
• MLL3/4 complexes establish active enhancer landscapes by depositing H3K4 mono- and di-methylation (H3K4me1/2) at enhancer regions.
• MLL3/4 complexes co-regulate H4K20 acetylation (H4K20ac) with the MiDAC complex to control specific gene expression programs.
• MLL3/4 subunits interact with ASXL proteins and BAP1 to recruit them to active enhancers, linking the complex to chromatin modification and deubiquitination.
• Dysregulation of MLL3/4 complex components is implicated in cancer and developmental disorders, and the complex is a target for menin-MLL inhibition strategies.
Description
The MLL3/4 complex (GO:0044666) is a multi-subunit protein complex that methylates lysine-4 of histone H3 and is defined by the presence of either MLL3 (KMT2C) or MLL4 (KMT2D) as its catalytic subunit in mammals, or their equivalent in other species. This complex is also referred to as the Trr/COMPASS-like complex and is conserved across metazoans. It plays a central role in establishing active enhancer landscapes, thereby influencing gene expression programs critical for development and cell identity. Researchers study GO:0044666 to understand how chromatin modifications are targeted to specific genomic regions and how disruptions in this process contribute to diseases such as cancer and developmental syndromes. The complex is not only a histone methyltransferase but also a platform for recruiting additional chromatin-modifying activities, including H4K20ac regulation via MiDAC and BAP1 recruitment through ASXL proteins. Understanding its composition, assembly, and regulation is essential for dissecting its roles in normal physiology and disease.
MLL3/4 complex At A Glance
| GO ID | GO:0044666 |
|---|---|
| GO term | MLL3/4 complex |
| Ontology | cellular_component |
| Synonym | Trr/COMPASS-like complex |
| Major function | Histone H3 lysine-4 methylation, enhancer activation |
| Catalytic subunits | MLL3 (KMT2C) or MLL4 (KMT2D) |
| Conservation | Conserved from Drosophila to mammals |
| Associated activities | H4K20ac co-regulation, BAP1 recruitment |
What Is GO:0044666?
According to the Gene Ontology, GO:0044666 (MLL3/4 complex) is a protein complex that can methylate lysine-4 of histone H3, and which contains either of the protein subunits MLL3 or MLL4 in mammals, or equivalent in other species. The complex is synonymous with the Trr/COMPASS-like complex and functions as a chromatin-modifying machine that deposits H3K4 methylation marks, particularly at enhancer regions.
Why Is MLL3/4 complex Important in Cell Biology?
The MLL3/4 complex is essential for establishing active enhancer landscapes that drive cell-type-specific gene expression programs. Its ability to methylate H3K4 and coordinate with other chromatin modifiers such as MiDAC and BAP1 places it at the center of transcriptional regulation. Dysregulation of MLL3/4 components is linked to cancer and developmental disorders, making the complex a focal point for understanding disease mechanisms and for therapeutic targeting, including menin-MLL inhibition strategies.
• Establishes active enhancer landscapes by depositing H3K4me1/2 marks.
• Co-regulates H4K20ac with MiDAC to control specific gene expression programs.
• Recruits BAP1 to active enhancers via ASXL proteins.
• Implicated in cancer through mutations in MLL3 (KMT2C) and MLL4 (KMT2D).
• Plays a role in developmental disorders such as Kabuki syndrome (MLL4/KMT2D mutations).
• Serves as a target for menin-MLL inhibition in leukemia.
• Contains PTIP-associated protein 1 (PA1), which has functions beyond the complex.
• Exhibits both enzymatic activity-dependent and -independent functions in gene regulation.
• Conserved function in Drosophila as the Trr/COMPASS-like complex.
• Potential biomarker for chemoresistance in BRCA-deficient cells through replication fork stability.
Structure and Composition of MLL3/4 complex
Catalytic core: MLL3 and MLL4
In simple terms: The complex has two possible catalytic engines, MLL3 or MLL4, which perform the actual methylation of histone H3.
The MLL3/4 complex is defined by the presence of either MLL3 (KMT2C) or MLL4 (KMT2D) as its catalytic subunit, both of which are histone H3 lysine-4 methyltransferases. These large proteins contain multiple domains that mediate interactions with other subunits and chromatin. The catalytic activity of MLL3/4 is responsible for depositing H3K4 methylation marks, primarily mono- and di-methylation, at enhancer regions.
ASXL proteins and BAP1 recruitment
In simple terms: ASXL proteins act as a bridge that brings the BAP1 enzyme to active enhancers.
ASXL proteins bind to the PHD2/3 fingers of MLL4, providing a mechanism for the recruitment of BAP1 to active enhancers. This interaction links the MLL3/4 complex to deubiquitination activities, which can influence chromatin states and gene expression.
PA1 and other accessory subunits
In simple terms: PA1 is a component of the MLL3/4 complex that also has independent roles.
PTIP-associated protein 1 (PA1) is a component of the MLL3/4 complex, but it also functions beyond the complex in other cellular processes. This suggests that the complex integrates multiple signaling and chromatin-modifying activities through its accessory subunits.
Co-regulation with MiDAC
In simple terms: The MLL3/4 complex works together with another complex called MiDAC to control histone acetylation.
The MLL3/4 complexes and MiDAC co-regulate H4K20ac to control a specific gene expression program. This functional interplay indicates that the MLL3/4 complex coordinates different histone modifications to fine-tune transcription.
Key Genes Involved in GO:0044666 MLL3/4 complex
The MLL3/4 complex comprises several key genes and proteins that contribute to its structure, regulation, and function.
| Gene | Major Role | Research Relevance |
|---|---|---|
| KMT2C (MLL3) | Catalytic subunit, H3K4 methyltransferase | Frequently mutated in cancer; defines the complex |
| KMT2D (MLL4) | Catalytic subunit, H3K4 methyltransferase | Mutations cause Kabuki syndrome; target for menin-MLL inhibition |
| ASXL1 | Accessory subunit, recruits BAP1 | Mutations in myeloid malignancies; links to chromatin modification |
| ASXL2 | Accessory subunit, recruits BAP1 | Part of the ASXL family that binds MLL4 PHD fingers |
| BAP1 | Deubiquitinase recruited by ASXL | Tumor suppressor; involved in enhancer regulation |
| PA1 (PAGR1) | Component of MLL3/4 complex | Has functions beyond the complex; potential signaling role |
| PTIP (PAXIP1) | Associated protein, DNA damage response | Links MLL3/4 to replication fork stability |
| MiDAC subunits | H4K20ac regulation | Co-regulates gene expression with MLL3/4 |
| MEN1 (Menin) | Interacts with MLL complexes | Target for menin-MLL inhibition in leukemia |
| KMT2A (MLL1) | Related H3K4 methyltransferase | Molecular switch with MLL3/4 dictates response to menin-MLL inhibition |
| KMT2B (MLL2) | Related H3K4 methyltransferase | Family member with distinct functions |
| SETD1A | H3K4 methyltransferase | COMPASS family member, not MLL3/4-specific |
| SETD1B | H3K4 methyltransferase | COMPASS family member |
| WDR5 | Common subunit of H3K4 methyltransferase complexes | Scaffold for complex assembly |
| RBBP5 | Common subunit of H3K4 methyltransferase complexes | Scaffold for complex assembly |
| ASH2L | Common subunit of H3K4 methyltransferase complexes | Stimulates methyltransferase activity |
| DPY30 | Common subunit of H3K4 methyltransferase complexes | Required for complex integrity |
How Is MLL3/4 complex Regulated?
The MLL3/4 complex is regulated at multiple levels. Its recruitment to chromatin is mediated by interactions with transcription factors and chromatin marks, and its activity can be modulated by associated proteins such as ASXL and BAP1. The complex also co-regulates H4K20ac with MiDAC, indicating that its function is integrated with other histone modification pathways. Additionally, a molecular switch between mammalian MLL complexes dictates the response to menin-MLL inhibition, suggesting that the balance between MLL3/4 and other MLL complexes can be dynamically regulated.
MLL3/4 complex and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| KMT2C (MLL3) | Cancer (various solid tumors and leukemia) | Knockout cell lines, xenograft models |
| KMT2D (MLL4) | Kabuki syndrome, cancer | Patient-derived iPSCs, knockout mice |
| ASXL1 | Myeloid malignancies | Knockout hematopoietic stem cells |
| BAP1 | Tumor predisposition, cancer | Knockout cell lines, mouse models |
| PTIP (PAXIP1) | Chemoresistance in BRCA-deficient cells | BRCA1/2-deficient cell lines with PTIP knockout |
Cancer
Mutations in KMT2C (MLL3) and KMT2D (MLL4) are frequently observed in various cancers, including leukemia, where the MLL3/4 complex is implicated in oncogenesis. The complex is a target for menin-MLL inhibition, and a molecular switch between MLL complexes can dictate therapeutic response. Additionally, replication fork stability conferred by MLL3/4-associated proteins like PTIP may influence chemoresistance in BRCA-deficient cells.
Developmental disorders
Germline mutations in KMT2D (MLL4) cause Kabuki syndrome, a developmental disorder characterized by intellectual disability and distinctive facial features. This highlights the critical role of the MLL3/4 complex in development and gene regulation.
Myeloid malignancies
ASXL1 mutations, which affect a subunit of the MLL3/4 complex, are common in myeloid malignancies and disrupt BAP1 recruitment to enhancers, contributing to leukemogenesis.
From MLL3/4 complex-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| What is the role of MLL3/4 in enhancer activation? | Knockout of KMT2C/KMT2D in cell lines followed by ChIP-seq |
| How does MLL4 mutation affect development? | Knock-in of patient mutations in iPSCs or mice |
| Does BAP1 recruitment depend on ASXL binding to MLL4? | Point mutations in MLL4 PHD fingers to disrupt ASXL binding |
| What is the function of PA1 outside the MLL3/4 complex? | Knockout of PAGR1 and rescue with tagged versions |
| How does MLL3/4 coordinate with MiDAC? | Double knockout of MLL3/4 and MiDAC subunits |
| Can menin-MLL inhibition be enhanced by modulating MLL3/4? | Overexpression or knockout of MLL3/4 in leukemia cells treated with inhibitors |
How to Study the MLL3/4 complex Process
| Method | What It Measures | Typical Application |
|---|---|---|
| ChIP-seq | Genomic binding of MLL3/4 and H3K4me1/2 | Mapping enhancer landscapes |
| RNA-seq | Gene expression changes | Identifying target genes upon complex disruption |
| Co-IP/MS | Protein-protein interactions | Discovering new subunits and modifications |
| CRISPR screen | Genetic dependencies and modifiers | Finding synthetic lethal partners |
| ATAC-seq | Chromatin accessibility | Assessing enhancer activity upon MLL3/4 loss |
| Western blot | Protein levels and histone modifications | Validating knockout efficiency |
| Immunofluorescence | Subcellular localization | Visualizing complex recruitment |
| Replication fork assays | Fork stability and degradation | Linking MLL3/4 to chemoresistance |
Chromatin immunoprecipitation sequencing (ChIP-seq)
ChIP-seq for H3K4me1/2 and MLL3/4 subunits is used to map the genomic binding sites of the complex and its histone marks, particularly at enhancers.
RNA sequencing (RNA-seq)
RNA-seq measures gene expression changes upon knockout or knockdown of MLL3/4 complex components to identify target genes and pathways.
Proteomics and co-immunoprecipitation
Affinity purification coupled with mass spectrometry identifies interacting partners and post-translational modifications of the complex.
CRISPR screening
Genome-wide CRISPR screens can identify synthetic lethal interactions and modifiers of MLL3/4 complex function in cancer cells.
How CRISPR Can Be Used to Study GO:0044666 MLL3/4 complex
Knockout
CRISPR knockout of KMT2C or KMT2D in cell lines is used to study loss-of-function phenotypes, including changes in enhancer marks and gene expression. Knockout of accessory subunits like ASXL1 can reveal their specific contributions to complex function.
Point Mutation
Point mutations can be introduced into catalytic residues of MLL3/4 to separate enzymatic activity from scaffolding functions. Mutations in the PHD fingers of MLL4 can disrupt ASXL binding and BAP1 recruitment.
Knock-in
Knock-in of patient-derived mutations, such as those found in Kabuki syndrome, allows modeling of disease-associated variants in isogenic cell lines. Tagged knock-in of MLL3/4 with fluorescent or epitope tags facilitates imaging and proteomics.
Overexpression
Overexpression of MLL3/4 or its subunits can be used to study gain-of-function effects and to test whether increased complex activity alters enhancer landscapes or drug responses.
How EDITGENE Supports MLL3/4 complex Research
Researchers studying MLL3/4 complex-related genes often need to determine whether a candidate gene is causally involved in enhancer regulation, chromatin modification, or disease phenotypes. EDITGENE provides a comprehensive suite of CRISPR services to enable precise genetic manipulation of MLL3/4 complex components in relevant cell models.
Contact EDITGENE today to design your custom CRISPR model for MLL3/4 complex research.
Frequently Asked Questions About MLL3/4 complex
What is the MLL3/4 complex?
The MLL3/4 complex (GO:0044666) is a protein complex that methylates histone H3 lysine-4 and contains either MLL3 or MLL4 as its catalytic subunit.
What genes are involved in the MLL3/4 complex?
Key genes include KMT2C (MLL3), KMT2D (MLL4), ASXL1, ASXL2, BAP1, PA1 (PAGR1), and PTIP (PAXIP1).
What is the function of the MLL3/4 complex?
It establishes active enhancer landscapes by depositing H3K4 methylation marks and coordinating with other chromatin modifiers.
How is the MLL3/4 complex regulated?
It is regulated by interactions with ASXL proteins, BAP1, and MiDAC, and its balance with other MLL complexes affects drug responses.
What diseases are associated with MLL3/4 complex mutations?
Mutations in KMT2C and KMT2D are linked to cancer and Kabuki syndrome, while ASXL1 mutations occur in myeloid malignancies.
What is the synonym for GO:0044666?
The synonym is Trr/COMPASS-like complex.
How can I study the MLL3/4 complex in the lab?
Common methods include ChIP-seq, RNA-seq, co-immunoprecipitation, and CRISPR knockout or knock-in models.
What is the role of MLL4 in enhancer regulation?
MLL4 binds ASXL proteins and recruits BAP1 to active enhancers, linking histone methylation to deubiquitination.
Is the MLL3/4 complex conserved?
Yes, it is conserved from Drosophila to mammals, where it is known as the Trr/COMPASS-like complex.
How does the MLL3/4 complex affect cancer treatment?
It influences response to menin-MLL inhibition, and a molecular switch between MLL complexes can dictate sensitivity.
Conclusion
The MLL3/4 complex (GO:0044666) is a critical chromatin-modifying machine that establishes active enhancer landscapes and coordinates multiple histone modifications to control gene expression. Its subunits are frequently mutated in cancer and developmental disorders, making it a prime target for therapeutic intervention. Understanding its structure, regulation, and disease roles requires precise genetic tools, which EDITGENE provides through custom CRISPR services.
References
- 1. Wang LH et al.. 2021. The MLL3/4 H3K4 methyltransferase complex in establishing an active enhancer landscape.. Biochem Soc Trans 49(3):1041-1054 PMID: 34156443
- 2. Liu B et al.. 2022. PTIP-Associated Protein 1: More Than a Component of the MLL3/4 Complex.. Front Genet 13:889109 PMID: 35754824
- 3. Ray Chaudhuri A et al.. 2016. Replication fork stability confers chemoresistance in BRCA-deficient cells.. Nature 535(7612):382-7 PMID: 27443740
- 4. Tian T et al.. 2024. UFL1 triggers replication fork degradation by MRE11 in BRCA1/2-deficient cells.. Nat Chem Biol 20(12):1650-1661 PMID: 38649452
- 5. Zhang Y et al.. 2024. ASXLs binding to the PHD2/3 fingers of MLL4 provides a mechanism for the recruitment of BAP1 to active enhancers.. Nat Commun 15(1):4883 PMID: 38849395
- 6. Wang X et al.. 2022. The MLL3/4 complexes and MiDAC co-regulate H4K20ac to control a specific gene expression program.. Life Sci Alliance 5(11) PMID: 35820704
- 7. Van HT et al.. 2024. KMT2 Family of H3K4 Methyltransferases: Enzymatic Activity-dependent and -independent Functions.. J Mol Biol 436(7):168453 PMID: 38266981
- 8. Soto-Feliciano YM et al.. 2023. A Molecular Switch between Mammalian MLL Complexes Dictates Response to Menin-MLL Inhibition.. Cancer Discov 13(1):146-169 PMID: 36264143
