C17ORF49 Human

What is C17ORF49 and what are its known synonyms?

C17ORF49 (Chromosome 17 open reading frame 49) is a protein-coding gene that produces a component of chromatin complexes. The protein is also known by several synonyms:

• BPTF-associated protein of 18 kDa (BAP18)

• Chromatin complexes subunit BAP18

• MLL1/MLL complex subunit C17orf49

• MGC49942

• HEPIS

The protein functions as a component of chromatin complexes including the MLL1/MLL and NURF (Nucleosome Remodeling Factor) complexes, suggesting its importance in chromatin regulation and gene expression .

What are the known biological functions of C17ORF49?

C17ORF49 (BAP18) plays significant roles in chromatin regulation through its association with multiple chromatin-modifying complexes:

• It functions as a component of the NURF complex, which relaxes condensed chromatin to promote DNA accessibility and target gene activation .

• It serves as an interacting partner in the MLL1/MLL complex, suggesting involvement in the regulation of histone methylation, particularly H3K4me3 marks .

• Its association with both complexes suggests BAP18 could be a critical link between active chromatin reader and writer activities, potentially coordinating these functions at transcription start sites .

• The protein appears to play a role in the regulation of promoters at critical genomic positions, such as active chromatin domains, offering multiple looping possibilities and thus regulating neighboring transcriptional units .

What are the most effective methods for studying C17ORF49 protein interactions?

Several methodologies have proven effective for investigating C17ORF49 protein interactions:

• Yeast Two-Hybrid (Y2H) Assays: This approach has successfully identified interaction partners such as HMGXB4's potential association with C17ORF49, as seen in studies using HeLa cDNA libraries .

• Co-Immunoprecipitation (Co-IP): For confirming direct protein-protein interactions in cellular contexts, particularly with other chromatin complex components like BPTF, SNF2L/SMARCA1, or MLL1.

• Chromatin Immunoprecipitation (ChIP): Valuable for identifying genomic regions where C17ORF49 binds, especially around transcription start sites enriched with H3K4me3 marks.

• Proximity Ligation Assays: To visualize protein interactions in situ within the nuclear compartment.

• Mass Spectrometry Analysis: For comprehensive identification of C17ORF49-associated protein complexes.

When designing interaction studies, researchers should consider using antibodies specifically validated for human C17ORF49, such as HPA024457, SAB1408286, or HPA022961, which have been confirmed effective for immunofluorescence, immunohistochemistry, and western blot applications .

What expression systems are optimal for producing recombinant C17ORF49?

Based on current research protocols, the following expression systems have been successfully employed:

• E. coli Expression Systems: Most commonly used for producing recombinant C17ORF49, typically yielding proteins with >90% purity as determined by SDS-PAGE .

• Tag Selection: An N-terminal His-tag (6x histidine) facilitates efficient purification using nickel affinity chromatography.

• Purification Protocol:

  • Initial capture using immobilized metal affinity chromatography

  • Further purification through proprietary chromatographic techniques

  • Buffer optimization (20mM Tris-HCl buffer (pH 8.0), 0.1M NaCl, 1mM DTT, 10% glycerol)

• Storage Considerations:

  • Store at 4°C if using within 2-4 weeks

  • For longer storage, maintain at -20°C

  • Addition of carrier protein (0.1% HSA or BSA) is recommended for long-term stability

  • Avoid multiple freeze-thaw cycles

What knockdown approaches are most effective for studying C17ORF49 function?

For functional studies investigating the role of C17ORF49, several gene knockdown strategies have been developed:

• siRNA Approaches: Predesigned siRNAs targeting C17ORF49 are available, designed using proprietary algorithms like Rosetta Inpharmatics for optimal knockdown efficiency .

• shRNA Methods: Several validated shRNA constructs targeting C17ORF49 can be employed for stable knockdown studies in various cell types .

• Experimental Considerations:

  • Verification of knockdown efficiency using validated antibodies (HPA024457, SAB1408286, HPA022961) is essential

  • Assessment of phenotypic effects should focus on chromatin structure alterations, transcriptional changes at H3K4me3-enriched promoters, and potential impacts on NURF and MLL1 complex functions

  • Controls should include scrambled RNA sequences and rescue experiments with recombinant C17ORF49 to confirm specificity

How does C17ORF49 contribute to chromatin regulation through its association with NURF and MLL complexes?

C17ORF49 (BAP18) appears to serve as a functional bridge between chromatin remodeling and histone modification activities:

• Role in NURF Complex: As part of the human NURF complex, C17ORF49 likely contributes to nucleosome remodeling activities that relax condensed chromatin, promoting DNA accessibility. The human core NURF complex shares three components with its Drosophila counterpart: BPTF, SNF2L/SMARCA1, and RBAP46/48 .

• MLL1 Complex Integration: C17ORF49's association with the MLL1 complex suggests involvement in H3K4 methylation, particularly the establishment of H3K4me3 marks at transcription start sites of active genes .

• Functional Integration Hypothesis: The dual association suggests C17ORF49 may coordinate the recognition of histone modifications (through BPTF's PHD finger and bromodomain) with the deposition of new H3K4me3 marks by MLL1, creating a potential feed-forward mechanism for maintaining active chromatin states.

• Regulatory Impact: This coordination may be particularly important at promoters in active chromatin domains where multiple looping possibilities exist, potentially regulating entire transcriptional neighborhoods rather than individual genes .

The exact molecular mechanism by which C17ORF49 facilitates communication between these complexes remains an active area of investigation, with particular focus on potential conformational changes or recruitment activities.

What is known about the relationship between C17ORF49 and HMGXB4?

The relationship between C17ORF49 and HMGXB4 represents an emerging area of research:

• Interaction Evidence: Yeast two-hybrid (Y2H) assays using a human HeLa cDNA library have identified a potential interaction between HMGXB4 and C17ORF49 .

• Functional Implications: HMGXB4 is involved in Wnt signaling regulation, suggesting C17ORF49 may influence this pathway through their interaction .

• Research Gaps: While the interaction has been initially identified, further confirmation through co-immunoprecipitation and functional studies is needed to establish the biological relevance and specificity of this association.

• Potential Mechanisms: The interaction may represent:

  • A regulatory mechanism where HMGXB4 modulates C17ORF49's chromatin-associated functions

  • A recruitment pathway where C17ORF49 brings chromatin-modifying activities to HMGXB4 target sites

  • A competitive interaction affecting the availability of either protein for their respective complexes

This relationship warrants further investigation, particularly examining how HMGXB4's HMG-box domain might interact with C17ORF49 and the consequences for transcriptional regulation.

How might C17ORF49 contribute to the regulation of transcription start sites through H3K4me3 modification?

C17ORF49's role in transcription start site (TSS) regulation appears to be multifaceted:

• H3K4me3 Association: H3K4me3 marks are highly enriched at TSSs of active genes and control gene transcription. C17ORF49, through its association with MLL1 (a writer of H3K4me3) and NURF (a reader of histone modifications), may facilitate both the deposition and recognition of these marks .

• Chromatin Accessibility Regulation: As part of the NURF complex, C17ORF49 likely contributes to relaxing chromatin structure around TSSs, making them accessible to transcription machinery .

• Promoter Architecture: The protein appears to play a role at promoters in critical genomic positions, particularly active chromatin domains that offer multiple looping possibilities. This suggests C17ORF49 may influence three-dimensional chromatin organization around TSSs .

• Potential Model of Action:

  • Initial recruitment to TSSs through existing H3K4me3 marks recognized by BPTF

  • Facilitation of MLL1 complex activity to maintain or extend H3K4me3 domains

  • Contribution to chromatin remodeling through NURF complex activity

  • Stabilization of chromatin loops that bring enhancers and promoters into proximity

These activities would collectively promote and maintain active transcriptional states at target genes.

What are the key challenges in studying C17ORF49 function in different cellular contexts?

Researchers face several methodological challenges when investigating C17ORF49:

• Complex Multi-Protein Associations: C17ORF49 functions within large multi-protein complexes (NURF and MLL1), making it difficult to distinguish its specific contribution from the activities of these complexes as a whole.

• Functional Redundancy: Potential redundancy with other chromatin regulators may mask phenotypes in knockdown studies.

• Cell Type Specificity: Expression and function may vary significantly across cell types, necessitating context-specific studies.

• Technical Challenges:

  • Limited availability of highly specific antibodies for chromatin immunoprecipitation

  • Difficulty in reconstituting functional chromatin complexes in vitro

  • Challenges in distinguishing direct from indirect effects in functional studies

• Recommended Approaches:

  • Employing multiple complementary techniques (genetic, biochemical, genomic)

  • Using acute depletion systems (e.g., auxin-inducible degron tags) to minimize compensation

  • Conducting studies across multiple cell types to identify context-dependent functions

  • Developing in vitro reconstitution systems with defined components

How should researchers interpret conflicting data on C17ORF49 function across different experimental systems?

When confronting conflicting results regarding C17ORF49 function, researchers should:

• Consider System-Specific Factors:

  • Cell type differences in expression of interaction partners

  • Variations in chromatin state and transcriptional programs

  • Differences in post-translational modifications affecting protein function

• Evaluate Methodological Differences:

  • Knockdown efficiency and duration (acute vs. chronic)

  • Specificity of antibodies and tags used

  • Resolution and sensitivity of detection methods

• Reconciliation Strategies:

  • Perform direct comparisons using standardized protocols across systems

  • Identify the specific variables that might explain discrepancies

  • Consider developing an integrated model that accounts for context-specific functions

• Validation Approaches:

  • Rescue experiments with wild-type and mutant constructs

  • Cross-validation using orthogonal techniques

  • Careful controls for off-target effects in genetic approaches

A systematic approach to resolving conflicts can often reveal important insights about context-dependent protein functions and regulatory mechanisms.

What emerging technologies could advance our understanding of C17ORF49 function?

Several cutting-edge technologies hold promise for elucidating C17ORF49's functions:

• CUT&RUN and CUT&Tag Approaches: These techniques offer higher resolution and lower background than traditional ChIP-seq, potentially revealing precise genomic binding sites of C17ORF49.

• Proximity Labeling Methods (BioID, APEX): Can identify transient or weak interaction partners in living cells, providing a more comprehensive view of C17ORF49's protein interaction network.

• Single-Cell Approaches: Single-cell techniques could reveal cell-to-cell variability in C17ORF49 expression and function, potentially identifying subpopulations with distinct regulatory states.

• Cryo-EM and Structural Biology: Advances in structural determination methods could reveal how C17ORF49 integrates into NURF and MLL1 complexes and how these interactions affect complex function.

• CRISPR Screens: Both loss-of-function and gain-of-function screens could identify genetic interactions and pathways involving C17ORF49.

• Proteomics Approaches: Quantitative analysis of post-translational modifications and protein complex composition following perturbation of C17ORF49 could reveal regulatory mechanisms.

What are the unexplored aspects of C17ORF49 in disease contexts?

Several aspects of C17ORF49 function in disease contexts remain underexplored:

• Cancer Biology: Given its role in chromatin regulation and association with MLL complexes (which are frequently dysregulated in leukemias), C17ORF49 may have unrecognized roles in oncogenesis or tumor suppression.

• Developmental Disorders: As a component of chromatin regulatory complexes, C17ORF49 dysfunction could contribute to developmental abnormalities, particularly those involving epigenetic dysregulation.

• Neurological Conditions: Many chromatin regulators are implicated in neurological disorders, suggesting C17ORF49 might play roles in these contexts.

• Potential Research Approaches:

  • Analysis of mutation and expression databases across disease states

  • Functional studies in disease-relevant cellular models

  • Assessment of C17ORF49 as a potential biomarker or therapeutic target

How might targeting C17ORF49 offer new approaches to modulating chromatin states?

The strategic position of C17ORF49 at the intersection of multiple chromatin regulatory complexes suggests several potential therapeutic applications:

• Selective Modulation: Unlike targeting core enzymatic components of chromatin complexes, modulating C17ORF49 might allow more selective effects on specific genomic regions or transcriptional programs.

• Complex Assembly Disruption: Small molecules targeting the interaction interfaces between C17ORF49 and its binding partners could selectively disrupt specific chromatin complex assemblies.

• Context-Specific Intervention: Understanding the tissue-specific functions of C17ORF49 could enable the development of context-specific approaches to modulating chromatin states.

• Research Priorities:

  • Identification of the structural determinants of C17ORF49's protein-protein interactions

  • Development of cell-permeable peptides or small molecules that can disrupt specific interactions

  • Characterization of the genomic and transcriptomic consequences of selective C17ORF49 modulation

These approaches could ultimately lead to novel therapeutic strategies for conditions involving dysregulated chromatin states.