β-hydroxybutyryl-HIST1H3A (K122) Antibody

What is lysine β-hydroxybutyrylation and its significance in histone research?

Lysine β-hydroxybutyrylation (Kbhb) represents a relatively new histone post-translational modification identified in 2016. This modification involves the addition of a β-hydroxybutyryl group to lysine residues, creating a nuanced layer of regulation in cellular functions . Specifically for histone research, Kbhb is enriched in active gene promoters and has been found to modify numerous lysine residues across all four types of histones (H1, H2A, H2B, H3, H4) . Understanding this modification provides insight into epigenetic regulation mechanisms and potentially connects metabolic status with gene expression patterns.

What applications has the β-hydroxybutyryl-HIST1H3A (K122) antibody been validated for?

The β-hydroxybutyryl-HIST1H3A (K122) polyclonal antibody has been specifically validated for several key applications in molecular biology research:

• Enzyme-Linked Immunosorbent Assay (ELISA)

• Western Blotting (WB)

• Immunocytochemistry (ICC)

These validated applications make this antibody a versatile tool for detecting and studying β-hydroxybutyrylation at the K122 position of human Histone H3.1 across multiple experimental contexts, from protein quantification to cellular localization studies.

What is the target specificity of the β-hydroxybutyryl-HIST1H3A (K122) antibody?

The β-hydroxybutyryl-HIST1H3A (K122) polyclonal antibody specifically recognizes human histone H3.1 that has been β-hydroxybutyrylated at the lysine 122 position . The antibody was generated using an immunogen consisting of a peptide sequence surrounding the β-hydroxybutyryl-Lys (122) site derived from Human Histone H3.1 (accession number P68431) . This site-specific antibody enables researchers to distinguish this particular modification from other post-translational modifications that may occur on histone H3.1.

How can I verify the specificity of my β-hydroxybutyryl-specific antibody in my experimental system?

Verifying antibody specificity is crucial for accurate interpretation of results, especially given the documented concerns about cross-reactivity with some Kbhb antibodies . A comprehensive approach to verify specificity includes:

• Treatment comparison tests: Treat cells with β-hydroxybutyrate (BHB), structurally similar compounds (like butyrate), and histone deacetylase inhibitors (e.g., Trichostatin A). Properly specific antibodies should show strong signals only in BHB-treated samples .

• Immunoprecipitation followed by mass spectrometry:

  • Immunoprecipitate proteins using your antibody of interest

  • Confirm enrichment via Western blot

  • Perform mass spectrometry to identify the precipitated peptides

  • Calculate the percentage of Kbhb-containing peptides to assess specificity

• Peptide competition assays: Pre-incubate the antibody with β-hydroxybutyrylated peptides versus unmodified or alternatively modified peptides before immunoblotting or immunoprecipitation to test for signal blocking.

When analyzing results, consider that true Kbhb-specific antibodies should not cross-react with non-modified lysine residues, unmodified peptides, 2-hydroxyisobutyrylated peptides, or acetylated peptides .

What are the optimal sample preparation methods for detecting β-hydroxybutyryl-HIST1H3A (K122) modifications?

For optimal detection of β-hydroxybutyryl-HIST1H3A (K122) modifications, follow this detailed protocol:

• Cell/tissue preparation:

  • For enhanced Kbhb signal, consider treating cells with β-hydroxybutyrate (4-10 mM) for 12-24 hours

  • Harvest cells and wash with ice-cold PBS

• Histone extraction protocol:

  • Lyse cells in Triton Extraction Buffer (PBS containing 0.5% Triton X-100, 2 mM PMSF, 0.02% NaN₃)

  • Centrifuge at 6,500 x g for 10 minutes at 4°C

  • For histone-specific extraction, resuspend the nuclear pellet in 0.2 N HCl overnight at 4°C

  • Centrifuge at 6,500 x g for 10 minutes

  • Neutralize the supernatant with 20% NaOH

• Sample preparation for Western blotting:

  • Determine protein concentration using the DC Protein Assay Kit

  • Mix sample with 4x Laemmli buffer/10% beta-mercaptoethanol

  • Heat at 95°C for 5 minutes

  • Resolve proteins by SDS-PAGE and transfer to nitrocellulose membrane

  • Block with 5% milk in TBS-T for 30-60 minutes

  • Incubate with β-hydroxybutyryl-HIST1H3A (K122) antibody at the recommended dilution

Including deacetylase inhibitors and β-hydroxybutyrylation-preserving reagents in your buffers can help maintain modification integrity throughout the extraction process.

How do I distinguish between true β-hydroxybutyrylation signals and potential cross-reactivity with other histone modifications?

Distinguishing true β-hydroxybutyrylation signals from cross-reactivity requires a multi-faceted approach:

• Control experiments:

  • Include parallel treatments with BHB, butyrate, and histone deacetylase inhibitors

  • True Kbhb-specific antibodies should show stronger signals in BHB-treated samples than with other treatments

  • Include untreated controls to establish baseline modification levels

• Mass spectrometry validation:

  • Follow immunoprecipitation with mass spectrometry analysis

  • Calculate the percentage of Kbhb-containing peptides in your samples

  • For true β-hydroxybutyrylation, BHB-treated samples should show significantly higher percentages of Kbhb-containing peptides compared to other treatments

• Cross-comparison with multiple antibodies:

  • Test your samples with pan-Kbhb antibodies alongside site-specific antibodies

  • Compare patterns between antibodies recognizing different sites (e.g., H3K122bhb vs. H4K8bhb)

  • Consistent patterns across different Kbhb-specific antibodies support true modification detection

Based on published research, caution is particularly warranted with H3K9bhb antibodies, which have shown cross-reactivity with acetylation and potentially other modifications .

What are the differences between various methodologies for studying β-hydroxybutyrylation patterns?

Multiple methodologies exist for studying β-hydroxybutyrylation patterns, each with distinct advantages:

For comprehensive β-hydroxybutyrylation research, integrating multiple methods provides the most robust understanding of this modification's biological significance and regulation .

How can I design experiments to investigate the functional significance of β-hydroxybutyrylation at H3K122?

Designing experiments to investigate the functional significance of β-hydroxybutyrylation at H3K122 requires a systematic approach:

• Manipulating β-hydroxybutyrylation levels:

  • Metabolic intervention: Treat cells with β-hydroxybutyrate (4-10mM) to increase Kbhb levels

  • Ketogenic conditions: Use low glucose/high fat medium or serum from fasting subjects

  • Genetic approaches: Manipulate enzymes potentially involved in β-hydroxybutyrylation (e.g., p300, CBP) using CRISPR-Cas9 or RNAi

• Functional readouts:

  • Transcriptional analysis: RNA-seq to identify genes affected by H3K122bhb changes

  • Chromatin accessibility: ATAC-seq or MNase-seq to determine if H3K122bhb affects nucleosome stability

  • Protein interactions: Identify readers of H3K122bhb using modified peptide pulldowns followed by mass spectrometry

• Site-specific manipulation:

  • Generate H3K122 mutants (K122R to prevent modification; K122Q to mimic acylation)

  • Express these mutants in cells with endogenous H3 depletion

  • Assess phenotypic and molecular consequences

• Integration with other histone modifications:

  • Perform sequential ChIP experiments (ChIP-reChIP) to determine co-occurrence with other modifications

  • Analyze spatial relationships between H3K122bhb and transcriptional machinery

When interpreting results, consider that H3K122 is located at the dyad axis of the nucleosome, potentially affecting DNA-histone interactions and chromatin accessibility.

What controls should I include when using β-hydroxybutyryl-HIST1H3A (K122) antibody in my experiments?

Rigorous controls are essential when working with β-hydroxybutyryl-HIST1H3A (K122) antibody:

• Antibody specificity controls:

  • Peptide competition assays: Pre-incubate antibody with β-hydroxybutyrylated and unmodified H3K122 peptides

  • Cross-reactivity assessment: Test against samples with known acetylation or other acylation modifications

  • Knockdown/knockout validation: Use H3.1-depleted cells as negative controls

• Treatment controls:

  • Positive control: Cells treated with β-hydroxybutyrate (BHB)

  • Negative controls: Untreated cells and cells treated with structurally similar compounds like butyrate

  • Process controls: Include histone deacetylase inhibitor (e.g., TSA) treated samples to distinguish from acetylation signals

• Technical controls:

  • Loading controls: Total H3 or housekeeping proteins

  • Secondary antibody-only controls: To assess non-specific binding

  • Isotype controls: Non-specific rabbit IgG for immunoprecipitation experiments

• Validation controls:

  • Mass spectrometry validation of immunoprecipitated proteins to confirm the presence of β-hydroxybutyrylated peptides

  • Alternative antibody validation: Use a separate antibody against total Kbhb or a different site-specific Kbhb antibody

Documenting all these controls systematically enhances the reliability and reproducibility of your research findings.

How can I quantify changes in β-hydroxybutyrylation levels in response to metabolic perturbations?

Quantifying changes in β-hydroxybutyrylation levels requires appropriate techniques and analytical approaches:

• Western blot quantification:

  • Normalize β-hydroxybutyryl-HIST1H3A (K122) signal to total H3 levels

  • Use densitometry software (ImageJ, Image Lab) for quantification

  • Apply statistical analysis across biological replicates (minimum n=3)

  • Present data as fold-change relative to control conditions

• Mass spectrometry-based quantification:

  • Label-free quantification: Compare spectral counts or ion intensities of β-hydroxybutyrylated peptides

  • SILAC labeling: Differentially label control and treated cells for direct comparison

  • Calculate modification stoichiometry: Ratio of modified to unmodified peptides

  • Example calculation: % Kbhb = (Kbhb peptide intensity / (Kbhb peptide + unmodified peptide intensity)) × 100

• ChIP-seq quantification:

  • Normalize H3K122bhb ChIP-seq reads to input and total H3 ChIP

  • Calculate differential binding scores across conditions

  • Correlate changes with gene expression data

  • Focus analysis on promoters, enhancers, and gene bodies separately

• Immunofluorescence quantification:

  • Measure nuclear fluorescence intensity using confocal microscopy

  • Perform single-cell analysis across populations

  • Plot distribution shifts rather than just average changes

When interpreting results, consider that BHB treatment typically increases H3K122bhb levels, with observed fold-changes of 2-5× being biologically significant in published studies.

What are common issues with β-hydroxybutyryl-HIST1H3A (K122) antibody experiments and how can they be resolved?

Researchers commonly encounter several challenges when working with β-hydroxybutyryl-HIST1H3A (K122) antibodies:

• Weak or absent signals:

  • Problem: Insufficient β-hydroxybutyrylation levels in samples

  • Solution: Pre-treat cells with β-hydroxybutyrate (4-10mM) for 12-24 hours

  • Problem: Degradation of modifications during sample preparation

  • Solution: Include deacetylase inhibitors and protease inhibitors in all buffers; process samples quickly at 4°C

• Non-specific binding and high background:

  • Problem: Antibody cross-reactivity with other modifications

  • Solution: Increase blocking time/concentration (5% milk or BSA for 1-2 hours); optimize antibody dilution with titration experiments

  • Problem: Excessive secondary antibody binding

  • Solution: More stringent washing (additional washes with higher Tween-20 concentration)

• Inconsistent results between experiments:

  • Problem: Variability in cell metabolic state

  • Solution: Standardize cell density, passage number, and serum conditions

  • Problem: Antibody lot-to-lot variation

  • Solution: Validate each new lot against previous lots; consider creating an internal standard sample

• Discrepancies between detection methods:

  • Problem: Different results from WB vs. ChIP vs. IF

  • Solution: Verify antibody validation for each specific application; optimize protocols for each technique separately

• Conflicting mass spectrometry data:

  • Problem: Antibody enriches non-β-hydroxybutyrylated peptides

  • Solution: Assess antibody specificity through IP-MS as described in search result ; consider using alternative antibodies if cross-reactivity is confirmed

How do I optimize ChIP protocols specifically for β-hydroxybutyryl-HIST1H3A (K122) antibody?

Optimizing ChIP protocols for β-hydroxybutyryl-HIST1H3A (K122) antibody requires several specific considerations:

• Crosslinking optimization:

  • Test different formaldehyde concentrations (0.5-2%) and incubation times (5-15 minutes)

  • Consider dual crosslinking with formaldehyde followed by EGS (ethylene glycol bis-succinimidyl succinate) for better preservation of protein-protein interactions

• Chromatin fragmentation:

  • Optimize sonication conditions to achieve fragments of 200-500bp

  • Verify fragment size distribution by agarose gel electrophoresis

  • Consider using enzymatic digestion (MNase) as an alternative to sonication for more precise fragmentation

• Antibody incubation parameters:

  • Test different antibody amounts (2-10 μg per ChIP reaction)

  • Optimize incubation time (overnight at 4°C is standard, but test 4-16 hours)

  • Include an IgG control and a positive control antibody (e.g., H3K4me3) in parallel

• Washing stringency balancing:

  • Implement a gradient of washing stringency to find optimal conditions

  • Example washing series: Low salt → High salt → LiCl → TE buffer

  • Monitor signal-to-noise ratio across different washing conditions

• Elution and reversal of crosslinking:

  • Test different elution buffers (SDS-based vs. acidic elution)

  • Optimize reversal of crosslinking (65°C for 4-16 hours)

  • Include RNase A and Proteinase K treatments

• Quality control metrics:

  • Calculate percent input recovery (should be >1% for histone marks)

  • Assess enrichment at known positive vs. negative genomic regions by qPCR

  • Measure signal-to-noise ratio compared to IgG control (should be >10-fold)

When optimizing, preserve sample aliquots at each step to troubleshoot specific stages if the final output is suboptimal.

What are the considerations for using β-hydroxybutyryl-HIST1H3A (K122) antibody in different cell types and tissues?

Using β-hydroxybutyryl-HIST1H3A (K122) antibody across different cell types and tissues requires careful consideration of several factors:

• Baseline β-hydroxybutyrylation variability:

  • Metabolically active tissues (liver, brain, heart) may have higher basal Kbhb levels

  • Proliferating cells often show different histone modification patterns than differentiated cells

  • Consider measuring β-hydroxybutyrate levels in different tissues to predict modification abundance

• Cell/tissue-specific extraction protocols:

  • Adjust lysis conditions based on tissue hardness and extracellular matrix composition

  • For tissues: Consider using a Dounce homogenizer followed by filtration

  • For difficult tissues: Might require additional mechanical disruption or enzymatic digestion

• Fixation considerations for immunohistochemistry:

  • Fixative choice affects epitope accessibility (4% PFA is standard, but test 1-4% range)

  • Fixation time should be optimized for each tissue type (10 min - 24 hours)

  • Antigen retrieval methods need tissue-specific optimization (citrate buffer pH 6.0 vs. Tris-EDTA pH 9.0)

• Antibody concentration adjustments:

  • Different tissues may require different antibody dilutions

  • Create a dilution series (1:250 to 1:2000) for each new cell type/tissue

  • Monitor signal-to-noise ratio to determine optimal concentration

• Confounding factors in specific tissues:

  • Tissues with high lipid content (brain, adipose) may have naturally elevated β-hydroxybutyrate levels

  • Autofluorescence in certain tissues (liver, kidney) may interfere with IF detection

  • Endogenous peroxidases can affect IHC staining

• Physiological state considerations:

  • Fasting/feeding state dramatically affects β-hydroxybutyrylation levels

  • Consider time of collection and nutritional status

  • In rodent models, standardize collection time relative to feeding cycle

When publishing, thoroughly document all cell/tissue-specific protocol adaptations to ensure reproducibility.

How does β-hydroxybutyrylation at H3K122 differ from other histone modifications in terms of function and genomic localization?

β-hydroxybutyrylation at H3K122 presents distinct characteristics compared to other histone modifications:

• Structural and functional significance:

  • H3K122 is located at the dyad axis of the nucleosome, where DNA makes strong contacts with the histone octamer

  • Modifications at this site potentially disrupt histone-DNA interactions more directly than modifications on histone tails

  • This disruption may facilitate DNA accessibility to transcription factors and the transcriptional machinery

• Genomic localization patterns:

  • Kbhb marks are generally enriched in active gene promoters

  • H3K122bhb may show distinct enrichment patterns compared to other Kbhb sites

  • Unlike H3K9bhb (which has antibody specificity concerns ), H3K122bhb detection appears more reliable

• Comparison with other modifications at H3K122:

  • H3K122 can also undergo acetylation (H3K122ac), which similarly associates with active chromatin

  • The chemical differences between β-hydroxybutyrylation and acetylation (additional hydroxyl group and longer carbon chain) may recruit different reader proteins

• Metabolic connections:

  • As a β-hydroxybutyrylation site, H3K122bhb likely responds to metabolic fluctuations, particularly ketone body metabolism

  • This creates a potential direct link between cellular metabolic state and gene regulation at specific genomic regions

• Relationship with other histone modifications:

  • May work cooperatively with activating marks like H3K4me3 or H3K27ac

  • Could have antagonistic relationships with repressive marks like H3K27me3

  • The combinatorial pattern with other modifications determines the ultimate functional outcome

Understanding these distinctive characteristics helps researchers interpret H3K122bhb data in the broader context of chromatin regulation and gene expression.

What is known about the enzymes that catalyze β-hydroxybutyrylation at H3K122 and their regulation?

The enzymatic regulation of β-hydroxybutyrylation at H3K122 remains an evolving area of research:

• Putative "writers" (acetyltransferases with potential β-hydroxybutyrylation activity):

  • HATs like p300 and CBP may have promiscuous acyltransferase activity, potentially catalyzing β-hydroxybutyrylation

  • These enzymes typically use acyl-CoA as donors, suggesting β-hydroxybutyryl-CoA may serve as a substrate

  • Definitive identification of specific enzymes for H3K122bhb remains an active research question

• Regulation mechanisms:

  • Metabolic regulation: β-hydroxybutyryl-CoA levels, derived from β-hydroxybutyrate metabolism, likely influence modification rates

  • Enzymatic competition: Various acyl-CoA species (acetyl-CoA, butyryl-CoA, β-hydroxybutyryl-CoA) may compete for the same enzyme active sites

  • Spatial regulation: Nuclear localization of metabolic enzymes generating β-hydroxybutyryl-CoA affects local substrate availability

• Potential "erasers" (deacylases):

  • HDACs (particularly class I and II) and sirtuins may remove β-hydroxybutyryl groups

  • SIRT3 has been implicated in removing various acylations, potentially including β-hydroxybutyrylation

  • Enzymatic specificity for different acylations varies, potentially creating differential regulation

• "Readers" of β-hydroxybutyrylation:

  • Bromodomain-containing proteins that recognize acetylated lysines might also interact with β-hydroxybutyrylated residues

  • YEATS domain proteins, which read various acylations, are potential candidates

  • Specific readers for H3K122bhb have not been definitively characterized

This incomplete understanding highlights opportunities for researchers to make significant contributions to this field through targeted experiments identifying and characterizing the enzymatic machinery governing H3K122 β-hydroxybutyrylation.

How can I integrate β-hydroxybutyrylation data with other omics approaches to understand broader biological implications?

Integrating β-hydroxybutyrylation data with other omics approaches provides a comprehensive understanding of its biological significance:

• Multi-omics integration strategies:

  • Correlate H3K122bhb ChIP-seq data with:

    • Transcriptomics (RNA-seq) to associate modification with gene expression

    • Other histone modification ChIP-seq to identify co-occurrence patterns

    • Chromatin accessibility (ATAC-seq, DNase-seq) to assess functional impact

    • Proteomics to connect histone modifications with protein expression

• Metabolomics integration:

  • Measure β-hydroxybutyrate levels and related metabolites alongside Kbhb profiling

  • Correlate β-hydroxybutyrate/β-hydroxybutyryl-CoA concentrations with modification abundance

  • Create metabolic perturbation time series to track dynamic responses

• Bioinformatic approaches:

  • Implement supervised machine learning to identify predictive relationships between Kbhb and other datasets

  • Use network analysis to place H3K122bhb in broader regulatory networks

  • Apply integrative genomics approaches like WGCNA (Weighted Gene Co-expression Network Analysis)

• Biological context integration:

  • Connect β-hydroxybutyrylation changes to:

    • Cell cycle progression

    • Differentiation trajectories

    • Stress responses

    • Metabolic adaptations (fasting, ketogenic conditions)

• Visualization and analysis tools:

  • Generate multi-dimensional visualizations integrating various omics layers

  • Use genome browsers with multiple tracks to visualize spatial relationships

  • Implement pathway enrichment analyses incorporating multiple data types

The integration of these diverse data types helps position H3K122bhb within the broader cellular regulatory landscape, revealing its functional roles in various biological processes and potentially identifying new therapeutic targets or biomarkers.

What are the latest developments in understanding the role of H3K122 β-hydroxybutyrylation in gene regulation?

Recent research has expanded our understanding of H3K122 β-hydroxybutyrylation's role in gene regulation:

• Functional genomics insights:

  • H3K122bhb, like other Kbhb marks, appears enriched at active gene promoters

  • The strategic position of K122 at the nucleosome dyad axis suggests direct impacts on DNA accessibility

  • Recent studies indicate potential roles in regulating specific gene sets responsive to metabolic fluctuations

  • The modification may serve as a direct link between cellular metabolic state and transcriptional regulation

• Biological context discoveries:

  • Emerging evidence connects H3K122bhb to cellular adaptations during fasting/feeding cycles

  • Studies suggest potential roles in metabolic diseases, particularly those involving ketone body metabolism

  • Investigation into developmental contexts reveals dynamic regulation during cellular differentiation

  • Potential regulatory functions during stress responses that alter metabolic states

• Technological advancements:

  • Improved antibody specificity has enabled more reliable detection of H3K122bhb

  • Advanced mass spectrometry techniques allow better quantification of modification stoichiometry

  • Single-cell approaches are beginning to reveal cell-to-cell variation in H3K122bhb patterns

  • CRISPR-based approaches enable site-specific manipulation of H3K122 to assess functional consequences

• Mechanistic understanding:

  • Recent investigations into the "readers" of H3K122bhb have identified potential protein interactors

  • Structural studies provide insight into how β-hydroxybutyrylation may alter histone-DNA interactions

  • Emerging models integrate H3K122bhb into broader epigenetic regulatory networks

These developments collectively point to H3K122bhb as an important epigenetic regulator that connects cellular metabolism with gene expression programs.

How do results from β-hydroxybutyryl-HIST1H3A (K122) antibody studies compare with mass spectrometry-based approaches?

Comparing antibody-based and mass spectrometry-based approaches for studying β-hydroxybutyryl-HIST1H3A (K122) reveals important complementarities and discrepancies:

• Detection sensitivity comparison:

  • Antibody-based methods generally offer higher sensitivity for detecting low-abundance modifications

  • Mass spectrometry provides definitive chemical identification but may miss low-stoichiometry modifications

  • Studies using both approaches show antibodies can detect H3K122bhb in conditions where MS approaches may yield negative results

• Specificity considerations:

  • Mass spectrometry offers unambiguous identification of the exact chemical modification

  • Antibody specificity concerns exist, with documented cross-reactivity for some Kbhb antibodies

  • When using H3K122bhb antibodies, validation with mass spectrometry is recommended

• Quantification capabilities:

  • MS approaches provide more accurate absolute quantification of modification stoichiometry

  • Antibody methods offer better relative quantification across different samples/conditions

  • Studies combining both approaches show that antibody signal intensity doesn't always linearly correlate with modification abundance

• Coverage differences:

  • Antibody methods enable genome-wide localization studies (ChIP-seq)

  • MS approaches better identify co-occurring modifications on the same histone molecule

  • Integrated studies reveal unique insights not obtainable from either method alone

• Consensus and discrepancies:

  • Both methods confirm H3K122bhb increases upon BHB treatment

  • Potential discrepancies in baseline detection levels should be interpreted carefully

  • Researchers should be aware that MS-identified bhb sites may not always be detectable by available antibodies

The field is increasingly moving toward integrated approaches, using antibodies for high-sensitivity detection and localization, complemented by MS for definitive identification and accurate quantification.

What future research directions are most promising for understanding β-hydroxybutyrylation function?

Several promising research directions will advance our understanding of β-hydroxybutyrylation function:

• Molecular mechanism exploration:

  • Identifying and characterizing specific "writers," "erasers," and "readers" of H3K122bhb

  • Determining the structural consequences of β-hydroxybutyrylation at the nucleosome level

  • Developing specific inhibitors or activators of enzymes regulating β-hydroxybutyrylation

• Physiological and disease relevance:

  • Investigating β-hydroxybutyrylation dynamics during:

    • Aging and longevity

    • Metabolic diseases (diabetes, obesity)

    • Neurodegenerative disorders

    • Cancer metabolism reprogramming

  • Exploring therapeutic potential of manipulating β-hydroxybutyrylation through diet or pharmacological approaches

• Technological developments:

  • Creating more specific antibodies against various Kbhb sites

  • Developing live-cell imaging tools for tracking dynamic β-hydroxybutyrylation changes

  • Implementing CRISPR-based technologies for site-specific manipulation of lysine residues

  • Enhancing computational approaches to integrate Kbhb data with other omics datasets

• Evolutionary perspectives:

  • Comparative analysis of β-hydroxybutyrylation across species

  • Investigating conservation of regulatory mechanisms

  • Exploring species-specific adaptations in β-hydroxybutyrylation pathways

• Single-cell and spatial technologies:

  • Mapping β-hydroxybutyrylation heterogeneity at the single-cell level

  • Spatial transcriptomics integrated with histone modification data

  • In situ detection of β-hydroxybutyrylation in tissue contexts

These research directions collectively will provide a comprehensive understanding of β-hydroxybutyrylation's role in cellular physiology and potentially reveal new therapeutic opportunities for manipulating this modification in disease contexts.