HIST1H1C (Ab-158) Antibody

What is HIST1H1C and what are its alternative designations in the literature?

HIST1H1C is a linker histone protein, specifically histone H1.2, which plays a crucial role in higher-order chromatin structure organization and epigenetic regulation. In the scientific literature, this protein is known by several synonyms, including:

• H1 histone family member 2

• H1.2

• H1F2

• H1s-1

• H12_HUMAN

• Histone H1c

• Histone H1d

• MGC3992

Understanding these alternative designations is essential when conducting literature searches and comparing experimental findings across different research groups.

What applications has the HIST1H1C (Ab-158) antibody been validated for?

The 2-hydroxyisobutyryl-HIST1H1C (K158) polyclonal antibody has been validated for multiple research applications:

• Enzyme-Linked Immunosorbent Assay (ELISA)

• Western Blotting (WB)

• Immunocytochemistry (ICC)

• Chromatin Immunoprecipitation (ChIP)

This diverse range of applications makes this antibody a versatile tool for investigating histone H1.2 modifications in various experimental contexts, from protein expression analysis to chromatin-DNA interaction studies.

What is the significance of the K158 position in HIST1H1C research?

The K158 position refers to the lysine residue at position 158 in the HIST1H1C protein sequence, which can undergo 2-hydroxyisobutyrylation, a post-translational modification (PTM). This specific modification is significant because:

• It represents one of the many PTMs that regulate histone function

• Site-specific modifications at different lysine residues can result in distinct functional outcomes

• The 2-hydroxyisobutyryl-HIST1H1C (K158) antibody specifically recognizes this modified form, allowing researchers to study this particular epigenetic mark

Research has demonstrated that site-specific modifications of histone H1 contribute to its regulatory functions in chromatin organization and gene expression control.

How should researchers design ChIP experiments using HIST1H1C (Ab-158) antibody?

When designing Chromatin Immunoprecipitation (ChIP) experiments with the HIST1H1C (Ab-158) antibody, researchers should follow these methodological considerations:

• Sample preparation: Use fresh tissue or cells with minimal processing time to preserve histone modifications

• Crosslinking: Optimize formaldehyde crosslinking time (typically 10-15 minutes) to effectively capture histone-DNA interactions

• Sonication: Adjust sonication conditions to generate DNA fragments of 200-500bp for optimal resolution

• Antibody concentration: Determine the optimal antibody concentration through titration experiments (typically starting with manufacturer recommendations)

• Controls: Include appropriate controls such as IgG negative control and a positive control targeting a well-characterized histone mark

• Validation: Confirm antibody specificity using Western blot on the same samples used for ChIP

This structured approach ensures reliable detection of 2-hydroxyisobutyrylated HIST1H1C at K158 in chromatin contexts.

What immunohistochemistry protocols work best with HIST1H1C antibodies?

For optimal immunohistochemistry (IHC) results with HIST1H1C antibodies, researchers should consider the following protocol elements:

• Fixation: Use 10% neutral buffered formalin for 24-48 hours

• Antigen retrieval: Heat-mediated antigen retrieval in citrate buffer (pH 6.0) is recommended

• Antibody dilution: A dilution range of 1:100-1:300 has been validated for IHC applications

• Detection system: Use a polymer-based detection system for enhanced sensitivity

• Counterstaining: Hematoxylin counterstaining provides optimal nuclear detail

• Positive control: Human thyroid cancer tissue has been successfully used as a positive control

This protocol has been optimized based on experimental validation with paraffin-embedded human tissue sections.

What are the validated sample types for HIST1H1C (Ab-158) antibody applications?

The HIST1H1C antibodies have been validated for use with various sample types:

Specific tissues with confirmed reactivity include:

• Mouse thymus tissue

• Mouse kidney tissue

• Mouse lung tissue

• Human thyroid cancer tissue

When working with new sample types, preliminary validation experiments are recommended to confirm antibody specificity and optimal working conditions.

How does ubiquitylation at different positions affect HIST1H1C function and interactions?

Recent research has revealed that site-specific ubiquitylation of histone H1 has profound effects on its function and interaction network:

• Distinct interactomes: Ubiquitylation at different positions results in overlapping but distinct interaction partners for H1.2

• Enzymatic regulation: Site-specific ubiquitylation modulates interactions with deubiquitylating enzymes and the deacetylase SIRT1

• Phase separation properties: Ubiquitylation at position K64 specifically impacts H1-induced phase separation and chromatosome assembly

• Conformational changes: K64 ubiquitylation induces conformational changes in the H1.2-SIRT1 complex, potentially affecting SIRT1's deacetylation activity

• Chromatin structure: Ubiquitylation at K64 leads to the formation of more numerous but less concentrated H1-dependent condensates, suggesting a reduction in chromatin compaction

These findings highlight that ubiquitylation serves as a critical regulator of H1 function beyond simple protein turnover, contributing to the modulation of chromatin structure and gene expression.

What is the relationship between HIST1H1C modifications and SIRT1 regulation?

The interaction between modified HIST1H1C and SIRT1 (Sirtuin 1) represents an important regulatory mechanism in chromatin biology:

• SIRT1 binding: HIST1H1C interacts with SIRT1, a NAD+-dependent deacetylase that targets various nuclear proteins including histones

• Ubiquitylation effects: Site-specific ubiquitylation of HIST1H1C at position K64 results in conformational changes within the HIST1H1C-SIRT1 complex

• Multiple conformational states: At least two main clusters of structural solutions exist for the H1.2 K64Ub-SIRT1 complex, suggesting multiple functional Ub-dependent conformations

• Deacetylation modulation: Ubiquitylation of H1.2 at K64 affects SIRT1's deacetylation capacity, potentially counteracting its transcriptional repressive function

• Epigenetic crosstalk: This interaction suggests crosstalk between different epigenetic modifications, as SIRT1 targets both core histones and linker histones

This complex interplay demonstrates how site-specific modifications of linker histones can influence broader epigenetic regulatory networks through protein-protein interactions.

How does HIST1H1C contribute to phase separation and chromatin organization?

Recent research has uncovered the role of HIST1H1C in biomolecular condensate formation and chromatin organization:

• Condensate formation: H1.2 can undergo liquid-liquid phase separation, forming molecular condensates important for chromatin organization

• Modification effects: Site-specific ubiquitylation affects and modulates condensate formation of H1.2

• K64 ubiquitylation: Ubiquitylation at position K64 leads to the formation of more numerous but less concentrated H1-dependent condensates

• Chromatosome effects: The effects observed for H1-DNA condensates are mirrored in intact chromatosomes (nucleosome + H1)

• Chromatin compaction: The observed reduction in condensate concentration suggests ubiquitylation promotes a more open chromatin conformation

• Transcriptional implications: This more open conformation may counteract transcriptional repression, consistent with the idea that H1 ubiquitylation plays a role in transcriptional control

These findings highlight how post-translational modifications of linker histones regulate not only protein-protein interactions but also the biophysical properties that govern chromatin structure.

What are the optimal storage conditions for maintaining HIST1H1C antibody activity?

To maintain optimal activity of HIST1H1C antibodies, researchers should follow these storage recommendations:

• Short-term storage: Store at 4°C for up to two weeks

• Long-term storage: Store at -20°C or -80°C in small aliquots to minimize freeze-thaw cycles

• Buffer composition: Ensure the antibody is stored in a suitable buffer containing preservatives

• Avoid freeze-thaw cycles: Minimize repeated freezing and thawing as this can lead to antibody denaturation and loss of activity

• Working solutions: Diluted working solutions should be prepared fresh and used within 24 hours

• Shipping conditions: Note that antibodies may be shipped at ambient temperature but should be properly stored upon arrival

Following these guidelines will help preserve antibody specificity and sensitivity for experimental applications.

How can researchers distinguish between different HIST1H1C modifications in experimental settings?

Differentiating between various HIST1H1C modifications requires careful methodological approaches:

• Specific antibodies: Use modification-specific antibodies such as the 2-hydroxyisobutyryl-HIST1H1C (K158) antibody that recognize particular PTMs at defined residues

• Mass spectrometry: Employ high-resolution mass spectrometry to detect and quantify different modifications

• Sequential immunoprecipitation: Perform sequential IP with antibodies against different modifications to analyze co-occurrence patterns

• In vitro competition assays: Use synthetic peptides with specific modifications to test antibody specificity

• Mutational analysis: Generate point mutations at specific lysine residues to validate antibody specificity and functional importance

• Western blot controls: Include appropriate controls such as unmodified protein and proteins with other modifications to confirm specificity

These approaches enable researchers to precisely characterize and distinguish between the various post-translational modifications that regulate HIST1H1C function.

What potential cross-reactivity issues should researchers be aware of when using HIST1H1C antibodies?

When working with HIST1H1C antibodies, researchers should be mindful of several potential cross-reactivity considerations:

• Histone family homology: The high sequence homology between histone variants may lead to cross-reactivity with other H1 family members (H1.1, H1.3, H1.4, etc.)

• Modification specificity: Antibodies targeting specific modifications (e.g., 2-hydroxyisobutyrylation at K158) may recognize similar modifications at equivalent positions in other histone variants

• Species specificity: While many HIST1H1C antibodies show cross-reactivity across human, mouse, and rat samples, verification is recommended when working with other species

• Non-specific binding: High antibody concentrations may lead to non-specific binding, requiring careful titration

• Validation approaches: Western blotting with recombinant proteins or peptide competition assays should be used to confirm specificity

• Batch variability: Different lots of the same antibody may show slight variations in specificity and sensitivity

Researchers should perform appropriate validation experiments with their specific samples to ensure antibody specificity before proceeding with full-scale experiments.

What emerging research areas are utilizing HIST1H1C antibodies for novel discoveries?

HIST1H1C antibodies are increasingly being utilized in several cutting-edge research areas:

• Phase separation biology: Investigating how histone modifications regulate condensate formation and chromatin organization

• Single-cell epigenomics: Exploring heterogeneity in histone modifications at the single-cell level

• Epigenetic crosstalk: Studying how histone H1 modifications interact with and influence other epigenetic marks

• Disease mechanisms: Examining the role of linker histone modifications in cancer, heart disease, and kidney disorders

• Therapeutic targeting: Developing approaches to modulate specific histone modifications for therapeutic purposes

These emerging areas represent exciting frontiers where HIST1H1C antibodies serve as essential tools for uncovering new biological mechanisms and potential therapeutic targets.

How can researchers integrate HIST1H1C modification data with other epigenomic and transcriptomic datasets?

For comprehensive epigenetic analysis, researchers can integrate HIST1H1C modification data with other datasets using these methodological approaches:

• Multi-omics integration platforms: Utilize bioinformatic tools designed for integrating ChIP-seq, RNA-seq, ATAC-seq, and other genomic datasets

• Correlation analysis: Perform correlation analysis between HIST1H1C modification patterns and gene expression profiles

• Co-occurrence mapping: Map the co-occurrence of HIST1H1C modifications with other histone marks and DNA methylation patterns

• Functional enrichment: Conduct pathway and ontology enrichment analyses on genes associated with specific HIST1H1C modification patterns

• Machine learning approaches: Apply supervised and unsupervised learning algorithms to identify complex relationships between multiple epigenetic marks

• Visualization tools: Use genome browsers and specialized visualization software to display integrated multi-omics data