KMT2E Antibody

What is KMT2E and why is it important in research?

KMT2E (lysine methyltransferase 2E) is a protein-coding gene located on chromosome 7 and a member of the myeloid/lymphoid or mixed-lineage leukemia (MLL) family. It encodes a protein with an N-terminal PHD zinc finger and a central SET domain . KMT2E is particularly important in research because it functions as:

• A key regulator of hematopoiesis, contributing to terminal myeloid differentiation and hematopoietic stem cell self-renewal through DNA methylation

• A crucial cell cycle regulator, influencing the G1/S transition, S phase progression, and mitotic entry

• A protein that binds to chromatin regions downstream of active gene start sites, regulating gene transcription through interaction with tri-methylated histone H3 at Lys-4 (H3K4me3)

Additionally, mutations in KMT2E have been associated with neurodevelopmental disorders including intellectual disability, autism, macrocephaly, hypotonia, gastrointestinal abnormalities, and epilepsy , making it relevant for both oncology and neuroscience research.

What types of KMT2E antibodies are available for research?

Current research-grade KMT2E antibodies include:

Most available KMT2E antibodies are raised in rabbits against human KMT2E, with cross-reactivity to mouse and rat in some cases. These antibodies are typically validated for Western blotting, immunohistochemistry, and immunofluorescence applications .

What is the molecular structure and function of the KMT2E protein?

KMT2E is a large protein of 1,858 amino acids with specific functional domains:

• N-terminal PHD zinc finger domain (amino acids 120-165)

• SET enzymatic domain (amino acids 282-445), which is predicted to be inactive

• Most of the protein has few scattered helices and strands

• Contains a disordered C-terminus

Functionally, KMT2E:

• Is recruited to E2F1 responsive promoters by HCFC1, promoting tri-methylation of histone H3 at Lys-4 (H3K4me3) and transcriptional activation

• Promotes the G1 to S phase transition in the cell cycle

• During myoblast differentiation, suppresses inappropriate expression of S-phase-promoting genes while maintaining the expression of determination genes in quiescent cells

How should I optimize Western blotting protocol for KMT2E detection?

For optimal Western blotting results with KMT2E antibodies:

• Sample preparation:

  • Use appropriate lysis buffers containing protease inhibitors to prevent degradation of this large protein (204 kDa)

  • Consider including proteasome inhibitors like MG132 in your experiments, as research indicates KMT2E protein stability can be proteasome-dependent

• Electrophoresis conditions:

  • Use low percentage (6-8%) SDS-PAGE gels to effectively resolve this high molecular weight protein

  • Run gels at lower voltage (80-100V) to prevent distortion of large proteins

• Transfer parameters:

  • Utilize wet transfer methods with cooled buffers

  • Extend transfer time (overnight at 30V) for complete transfer of large proteins

• Antibody dilutions:

  • Start with dilutions of 1:500-1:2000 as recommended for Western blotting

  • Optimize by testing several dilutions on positive control samples

• Controls:

  • Include positive controls from cell lines known to express KMT2E

  • Consider knockdown samples as negative controls to confirm specificity

Research has shown that KMT2E protein levels can be stabilized under hypoxic conditions, with short-term (4 hours) exposure to transcriptional inhibitor actinomycin D reducing KMT2E transcript but not protein expression , suggesting potential experimental considerations when studying protein stability.

What immunohistochemistry techniques work best for KMT2E detection in tissue samples?

For effective immunohistochemical detection of KMT2E:

• Fixation and embedding:

  • Use 10% neutral buffered formalin for tissue fixation (12-24 hours)

  • Standard paraffin embedding protocols are suitable

• Antigen retrieval:

  • Heat-induced epitope retrieval in citrate buffer (pH 6.0) is recommended

  • Pressure cooking for 20 minutes typically yields better results than microwave heating

• Antibody dilution and incubation:

  • Start with antibody dilutions of 1:50-1:200 as recommended

  • Incubate overnight at 4°C for optimal binding

• Detection systems:

  • HRP-conjugated secondary antibodies with DAB substrates provide good signal-to-noise ratio

  • For dual labeling studies, consider fluorescent secondary antibodies

• Validation approaches:

  • Compare staining patterns with Atlas Antibodies' validated Human Protein Atlas results

  • Include tissues from KMT2E-related syndrome patients as positive controls when available

  • Use standard blocking peptides to confirm specificity

Research has revealed increased expression of H3K9me3 (but not H3K27me3) in diseased pulmonary arterioles in human group 1 PAH and group 3 PH , which could be useful as a comparative marker in studies involving KMT2E.

How can I design experiments to study KMT2E and KMT2E-AS1 interactions?

Based on recent research findings, the following experimental approach is recommended:

• RNA-protein interaction studies:

  • Implement RNA-protein immunoprecipitation assays using antibodies against KMT2E followed by RT-qPCR of KMT2E-AS1

  • Use actinomycin D treatment to assess stability of RNA-protein complexes

  • Apply proteasomal inhibitors (e.g., MG132) to investigate protein stability mechanisms

• Proximity ligation assays:

  • Utilize this technique to study the interaction between H3K4me3 and KMT2E

  • Compare wild-type with deletion mutants missing conserved sequences

• Knockdown and overexpression experiments:

  • Design effective siRNAs targeting KMT2E-AS1 to assess effects on KMT2E protein expression

  • Create lentiviral constructs for forced expression of KMT2E-AS1 to study function

  • Generate deletion mutants of KMT2E-AS1 missing conserved sequences (~600-bp) to identify functional domains

• Histone modification analysis:

  • Assess H3K4me3 and H3K9me3 marks via immunoblotting after KMT2E-AS1 manipulation

  • Monitor changes under hypoxic versus normoxic conditions

Research has demonstrated that KMT2E-AS1 complexes with and stabilizes KMT2E protein to increase H3K4me3 histone trimethylation , and that KMT2E deficiency can induce a histone demethylase (LSD1) that specifically reduces H3K9me3 .

How does KMT2E function in epigenetic regulation of hematopoiesis?

KMT2E plays a critical role in hematopoietic regulation through several epigenetic mechanisms:

• Chromatin binding and modification:

  • KMT2E binds to chromatin regions downstream of active gene start sites

  • This binding is facilitated by interaction with tri-methylated histone H3 at Lys-4 (H3K4me3)

  • KMT2E promotes further tri-methylation of H3K4, creating a positive feedback loop for gene activation

• Hematopoietic stem cell maintenance:

  • Research indicates KMT2E contributes to hematopoietic stem cell self-renewal

  • This function appears to be mediated through DNA methylation patterns

• Terminal myeloid differentiation:

  • KMT2E regulates the expression of genes necessary for proper myeloid cell development

  • Disruption of KMT2E function may contribute to myeloid leukemias

• Cell cycle regulation in hematopoietic cells:

  • KMT2E is recruited to E2F1 responsive promoters by HCFC1

  • This recruitment promotes H3K4 trimethylation and transcriptional activation

  • These actions regulate the G1 to S phase transition in the cell cycle

Recent research has identified a genetic association between rs73184087, a single-nucleotide variant within a KMT2E intron, and disease risk in pulmonary arterial hypertension (PAH), suggesting potential pathogenic roles of KMT2E in vascular diseases beyond its known hematopoietic functions .

What is the role of KMT2E in neurodevelopmental disorders?

KMT2E has been implicated in several neurodevelopmental processes and disorders:

• Genetic evidence:

  • As of 2025, more than 120 people with KMT2E-related syndrome have been identified in medical research

  • Heterozygous variants in KMT2E cause a spectrum of neurodevelopmental phenotypes

  • Both de novo variants and inherited variants from parents have been documented

• Clinical manifestations:

  • Developmental delay and/or intellectual disability

  • Autism spectrum disorder or autistic features

  • Low muscle tone (hypotonia)

  • Seizures and speech delay

  • Head size abnormalities (both macrocephaly and microcephaly reported)

  • Sleep issues and behavioral problems including self-injury, anxiety, and aggression

  • Gastrointestinal issues

  • Brain changes visible on MRI

• Molecular mechanisms:

  • KMT2E is thought to regulate neurodevelopmental genes through histone modifications

  • Studies suggest roles in neuronal differentiation and maturation

  • The protein's SET domain, though predicted to be inactive, may still influence other epigenetic regulators

• Phenotypic variability:

  • Missense variants in KMT2E show no clustering pattern; they can occur in the SET domain, PHD domain, or outside identified domains

  • This may explain the wide spectrum of neurodevelopmental manifestations

Research indicates that therapeutic approaches for KMT2E-related syndrome should begin as early as possible, ideally before a child begins school, though currently there are no medicines specifically designed to treat the syndrome .

How can KMT2E antibodies be used to study its interaction with HIF-2α in pulmonary hypertension?

Recent research has uncovered a significant role for KMT2E in pulmonary hypertension through its interaction with HIF-2α. The following experimental approaches are recommended:

• Co-immunoprecipitation studies:

  • Use KMT2E antibodies to pull down protein complexes from hypoxic pulmonary arterial endothelial cells

  • Probe for HIF-2α to confirm interaction

  • Compare normoxic versus hypoxic conditions to assess oxygen-dependent interactions

• ChIP-seq analysis:

  • Perform chromatin immunoprecipitation with KMT2E antibodies followed by sequencing

  • Identify genomic regions co-occupied by KMT2E and HIF-2α

  • Analyze enrichment for histone marks such as H3K4me3

• Functional assays:

  • Combine KMT2E antibody-based detection with KMT2E-AS1 manipulation

  • Research has shown that KMT2E-AS1 stabilizes KMT2E protein to increase H3K4me3, driving HIF-2α-dependent metabolic and pathogenic endothelial activity

  • Monitor changes in HIF-2α expression and activity across epigenetic, transcriptional, and posttranscriptional contexts

• Genetic variant analysis:

  • Assess the impact of rs73184087, a single-nucleotide variant within a KMT2E intron associated with PAH risk

  • This variant displays allele (G)-specific association with HIF-2α and engages in long-range chromatin interactions

Research has demonstrated that KMT2E-AS1 and KMT2E act in a positive feedback loop with HIF-2α to exacerbate pulmonary arterial hypertension through epigenetic and metabolic changes . In vivo studies have shown that KMT2E-AS1 deficiency protected against PAH in mice, as did pharmacologic inhibition of histone methylation in rats, while forced lncRNA expression promoted more severe pulmonary hypertension .

What are common issues with KMT2E antibody specificity and how can they be addressed?

When working with KMT2E antibodies, researchers may encounter several specificity challenges:

• High molecular weight detection issues:

  • Problem: Incomplete transfer of this large protein (204 kDa) during Western blotting

  • Solution: Extended transfer times, lower percentage gels, and wet transfer methods

• Non-specific bands:

  • Problem: Secondary bands appearing at unexpected molecular weights

  • Solution:

    • Titrate antibody dilutions (try 1:500-1:2000 range for Western blotting)

    • Include blocking peptides as controls

    • Validate with siRNA knockdown samples

• Cross-reactivity concerns:

  • Problem: Potential cross-reactivity with other MLL family proteins

  • Solution:

    • When possible, use antibodies validated on protein arrays (e.g., those tested on arrays of 364 human recombinant protein fragments)

    • Compare results using multiple antibodies targeting different epitopes

• Epitope masking:

  • Problem: Post-translational modifications may mask antibody binding sites

  • Solution:

    • Try multiple antibodies targeting different regions of KMT2E

    • Consider the impact of hypoxia, which has been shown to stabilize KMT2E protein

• Validation approaches:

  • Implement the "Enhanced Validation" methodology used by Atlas Antibodies

  • Utilize genetic models (knockdown/knockout) as negative controls

  • Compare staining patterns with published results in the Human Protein Atlas

Some antibodies, like those from Bio-Rad, have been validated for cross-reactivity with mouse KMT2E , which is important for translational studies between human and animal models.

How can I interpret conflicting results in KMT2E expression studies?

When facing contradictory findings in KMT2E expression analysis:

• Consider cellular context and conditions:

  • Hypoxia significantly impacts KMT2E regulation; KMT2E protein is stabilized under hypoxic conditions even when transcript levels decrease

  • The KMT2E-AS1/KMT2E interaction varies by oxygen status and cell type

• Evaluate detection methods:

  • Compare transcript versus protein detection results

  • mRNA levels may not correlate with protein levels due to post-transcriptional regulation

  • Research shows short-term (4 hours) exposure to transcriptional inhibitor actinomycin D reduces KMT2E transcript but not protein expression

• Assess genetic background:

  • Single nucleotide polymorphisms, particularly rs73184087 in the KMT2E intron, affect expression patterns

  • This SNV displays allele (G)–specific association with HIF-2α and expression patterns

• Examine experimental timepoints:

  • KMT2E functions in cell cycle regulation, so expression may vary based on cell cycle phase

  • Compare acute versus chronic manipulation experiments

• Consider protein-protein interactions:

  • KMT2E is recruited to E2F1 responsive promoters by HCFC1

  • Differences in cofactor expression could explain varying KMT2E activity

Research has shown that proteasomal inhibition with MG132 can reverse the effect of KMT2E-AS1 knockdown on KMT2E protein levels , suggesting that protein stability mechanisms significantly impact observed expression levels and should be considered when interpreting conflicting results.

How should discrepancies between in vitro and in vivo KMT2E findings be reconciled?

To address discrepancies between in vitro cell culture and in vivo animal model results:

• Microenvironment factors:

  • In vivo models provide the complete physiological context including tissue-specific interactions

  • Consider how the three-dimensional tissue architecture affects KMT2E function

  • Research shows KMT2E-AS1 deficiency protected against PAH in mice, while forced expression promoted worse PH in vivo

• Developmental timing:

  • KMT2E has roles in development that may not be recapitulated in cell culture

  • KMT2E-related syndrome manifests with developmental abnormalities that progress over time

  • Animal models may capture developmental aspects missing from cell studies

• Compensatory mechanisms:

  • In vivo systems may activate compensatory pathways absent in vitro

  • Other MLL family proteins may partially compensate for KMT2E dysfunction in intact organisms

• Cell type heterogeneity:

  • Tissue samples contain multiple cell types with varying KMT2E expression

  • Compare results from homogeneous cell cultures with heterogeneous tissue samples

  • Consider single-cell approaches to resolve cell-specific effects

• Methodological reconciliation:

  • Use organoid cultures as an intermediate model system

  • Apply pharmacologic inhibition (e.g., of histone methylation) in both settings to compare responses

  • Validate antibody performance in both contexts separately

What are new approaches for studying KMT2E in hematological malignancies?

Emerging methodologies for investigating KMT2E in blood cancers include:

• Single-cell multi-omics:

  • Combine single-cell RNA-seq with ChIP-seq using KMT2E antibodies

  • Correlate KMT2E binding patterns with gene expression at single-cell resolution

  • This approach can identify heterogeneous cell populations with differential KMT2E activity

• CRISPR-based epigenome editing:

  • Target KMT2E to specific genomic loci using dCas9-KMT2E fusions

  • Assess the direct impact of KMT2E recruitment on gene expression and cell transformation

  • Compare with effects of other MLL family members to determine functional specificity

• Patient-derived xenograft (PDX) models:

  • Establish PDX models from hematological malignancies

  • Use KMT2E antibodies to monitor protein expression and localization

  • Test targeted therapies against the KMT2E pathway in these models

• Liquid biopsy approaches:

  • Develop techniques to detect KMT2E protein or KMT2E-AS1 in circulation

  • Correlate with disease progression and treatment response

  • Early research suggests potential for biomarker development

• Targeting the KMT2E-AS1/KMT2E interaction:

  • Design therapeutic strategies to disrupt the stabilizing effect of KMT2E-AS1 on KMT2E

  • Research has shown this tandem pair operates as a functional unit

  • RNA-targeting approaches may provide novel therapeutic avenues

KMT2E has been implicated in myeloid/lymphoid or mixed-lineage leukemia, and aberrant levels have been shown to inhibit cell cycle progression , making it a promising target for therapeutic intervention in hematological malignancies.

How can KMT2E antibodies be used in studying neurodevelopmental disorders?

Innovative applications of KMT2E antibodies in neurodevelopmental research include:

• Brain organoid studies:

  • Use KMT2E antibodies for immunostaining of human brain organoids

  • Compare organoids derived from patients with KMT2E-related syndrome versus controls

  • Track KMT2E expression during organoid development to identify critical windows

• Genome-wide binding profiling:

  • Perform ChIP-seq with KMT2E antibodies in neural progenitor cells and mature neurons

  • Identify neuronal genes regulated by KMT2E

  • Compare binding profiles between cells with wild-type and mutant KMT2E

• Protein interaction networks:

  • Use KMT2E antibodies for co-immunoprecipitation followed by mass spectrometry

  • Map KMT2E interaction partners in neural tissues

  • Compare interactomes between normal and KMT2E-mutant samples

• Post-mortem tissue analysis:

  • Apply KMT2E antibodies for immunohistochemistry on brain sections

  • Examine expression patterns in individuals with neurodevelopmental disorders

  • Compare with genetic data on KMT2E mutations when available

• Therapeutic monitoring:

  • Use KMT2E antibodies to assess effects of potential therapeutics on KMT2E levels and activity

  • Monitor histone modifications (H3K4me3) as a readout of KMT2E function

  • Current research indicates therapies should begin as early as possible

Research has shown that KMT2E mutations are associated with a range of neurological disorders including epilepsy, autism, and abnormalities in gastrointestinal function , making KMT2E antibodies valuable tools for studying these conditions.

What potential exists for using KMT2E as a therapeutic target in pulmonary hypertension?

Recent research has revealed promising avenues for targeting KMT2E in pulmonary hypertension treatment:

• Inhibition of the KMT2E-AS1/KMT2E axis:

  • Research has demonstrated that KMT2E-AS1 and KMT2E act in a positive feedback loop with HIF-2α to exacerbate PAH

  • In vivo studies show that KMT2E-AS1 deficiency protected against PAH in mice

  • Developing antisense oligonucleotides targeting KMT2E-AS1 could disrupt this pathogenic pathway

• Histone methylation inhibition:

  • Pharmacologic inhibition of histone methylation reversed PAH pathology in rats

  • KMT2E antibodies can be used to monitor treatment effects on protein levels and localization

  • Screening for small molecule inhibitors of the KMT2E complex could yield therapeutic candidates

• Targeting the rs73184087 variant:

  • This single-nucleotide variant within a KMT2E intron is associated with PAH risk

  • It displays allele (G)–specific association with HIF-2α and engages in long-range chromatin interactions

  • Gene editing approaches targeting this variant could be explored in preclinical models

• Biomarker development:

  • KMT2E antibodies could be used to develop assays for monitoring disease progression

  • Changes in KMT2E protein levels or post-translational modifications might correlate with disease severity

  • Combined with genetic screening for the rs73184087 variant, this could enable personalized treatment approaches

• Combination therapies:

  • Targeting the KMT2E pathway alongside standard PAH therapies

  • KMT2E antibodies can be used to monitor pathway modulation in response to treatment

  • Synergistic approaches targeting both KMT2E and HIF-2α pathways may provide enhanced efficacy