Phospho-ERBB4 (Y1284) Antibody

What is ERBB4/HER4 and what role does Y1284 phosphorylation play in its function?

ERBB4/HER4 is a 180kDa (observed) receptor tyrosine kinase belonging to the epidermal growth factor receptor subfamily. It is a single-pass type I membrane protein containing multiple functional domains: cysteine-rich domains, a transmembrane domain, a tyrosine kinase domain, a phosphatidylinositol-3 kinase binding site, and a PDZ domain binding motif .

The phosphorylation at tyrosine 1284 (Y1284) represents a critical regulatory site that occurs after ligand binding triggers receptor dimerization and autophosphorylation. This specific phosphorylation event serves as a docking site for downstream signaling molecules, particularly in pathways related to cell proliferation, survival, and differentiation . Y1284 phosphorylation is especially significant in activating signaling cascades that regulate cellular responses to neuregulins and other EGF family members .

How does ERBB4 signaling differ from other ERBB family members, and why is Y1284 phosphorylation specifically important?

ERBB4 signaling is distinguished from other ERBB family members (EGFR/ERBB1, ERBB2/HER2, ERBB3/HER3) by its unique ability to undergo proteolytic processing, releasing both cytoplasmic and extracellular fragments that can function independently . Unlike other family members, ERBB4 also exists in multiple splice variants that affect ligand specificity and downstream signaling pathways.

The Y1284 phosphorylation site is particularly important because it serves as a specific docking point for signaling proteins involved in the MAPK pathway activation. While other ERBB family members share some phosphorylation-dependent signaling mechanisms, the Y1284 site in ERBB4 creates unique signaling capabilities, particularly in neuronal development, cardiac function, and mammary gland differentiation contexts . This phosphorylation site helps determine which downstream pathways are activated after receptor stimulation.

What are the known activators and inhibitors of ERBB4 Y1284 phosphorylation?

Activators:

• Neuregulins (NRG1, NRG2, NRG3, NRG4): Primary physiological ligands that induce receptor dimerization and autophosphorylation

• EGF family members: Betacellulin (BTC), Epiregulin (EREG), and Heparin-binding EGF-like growth factor (HBEGF)

• IGF: As indicated by positive controls using MCF7+IGF cell lines in antibody validation studies

Inhibitors:

• Tyrosine kinase inhibitors targeting the ERBB family

• Phosphatase enzymes that dephosphorylate specific tyrosine residues

• Receptor antagonists that prevent ligand binding and subsequent activation

The detection of these phosphorylation states often requires specific experimental conditions, such as stimulation with growth factors like IGF prior to analysis .

What are the optimal protocols for detecting ERBB4 Y1284 phosphorylation using Western blotting?

For optimal Western blot detection of phosphorylated ERBB4 Y1284:

• Sample Preparation:

  • Stimulate cells with appropriate ligands (neuregulins or IGF) to increase phosphorylation levels

  • Lyse cells in buffer containing phosphatase inhibitors to preserve phosphorylation

  • Use MCF7+IGF as a positive control sample

• Protocol Parameters:

  • Recommended antibody dilutions: 1:500-1:2000 (optimize for your specific application)

  • Protein loading: 20 μg per lane (standard recommendation)

  • Blocking: 3% nonfat dry milk in TBST

  • Secondary antibody: HRP-conjugated anti-rabbit IgG at 1:10000 dilution

  • Detection: ECL-based systems with exposure time of approximately 90 seconds

• Controls and Validation:

  • Include both phosphorylated (stimulated) and non-phosphorylated (unstimulated) samples

  • Consider using 293T cells transfected with ERBB4 as a positive control system

  • Expected molecular weight: 180kDa (observed) versus 147kDa (calculated)

This methodology ensures specific detection of the phosphorylated Y1284 site while minimizing background and non-specific signals.

How can I optimize immunofluorescence experiments using Phospho-ERBB4 (Y1284) antibodies?

For successful immunofluorescence experiments with Phospho-ERBB4 (Y1284) antibodies:

• Sample Preparation:

  • For cell lines: Use transfected cells (e.g., 293T cells expressing ERBB4) for optimal signal

  • For tissue sections: Fresh-frozen sections generally yield better phospho-specific signals than FFPE

  • Pre-stimulate cells with appropriate ligands to enhance phosphorylation

• Protocol Optimization:

  • Recommended antibody dilutions: 1:50-1:200 for immunofluorescence applications

  • Fixation: 4% paraformaldehyde (10 minutes) preserves phospho-epitopes better than methanol

  • Permeabilization: 0.1% Triton X-100 (5-10 minutes)

  • Blocking: 5% normal serum (matching secondary antibody species) with 1% BSA

  • Secondary detection: Use high-quality fluorophore-conjugated secondary antibodies (e.g., Cy3 Goat Anti-Rabbit IgG)

  • Include phosphatase inhibitors in all buffers

• Controls and Validation:

  • Positive control: 293T cells transfected with ERBB4

  • Negative controls: Untransfected cells and primary antibody omission

  • Phosphatase treatment control: Treat duplicate samples with lambda phosphatase to confirm phospho-specificity

This approach maximizes signal-to-noise ratio while ensuring the specificity of phospho-ERBB4 Y1284 detection in cellular contexts.

What cross-reactivity concerns should researchers be aware of when using Phospho-ERBB4 (Y1284) antibodies?

When working with Phospho-ERBB4 (Y1284) antibodies, researchers should consider several cross-reactivity issues:

To address these concerns:

• Always include appropriate positive and negative controls

• Validate antibody specificity using phosphatase treatment

• Consider using blocking peptides (available from some manufacturers) to confirm specificity

• For critical experiments, validate results with a second antibody from a different supplier

Why might I observe differences between the calculated (147kDa) and observed (180kDa) molecular weights of ERBB4?

The discrepancy between the calculated (147kDa) and observed (180kDa) molecular weight of ERBB4 is a common phenomenon with several possible explanations:

• Post-translational Modifications:

  • Glycosylation: ERBB4 contains multiple glycosylation sites in its extracellular domain

  • Phosphorylation: Multiple tyrosine, serine, and threonine phosphorylation sites

  • Ubiquitination: Can add significant molecular weight

• Structural Features:

  • The extended conformation of ERBB4 may result in slower migration during SDS-PAGE

  • The high number of cysteine-rich domains affects protein folding and SDS binding

• Technical Considerations:

  • Gel percentage and running conditions can affect apparent molecular weight

  • Protein standards used for calibration might not reflect actual migration patterns of heavily modified proteins

This discrepancy is well-established and documented across multiple antibody datasheets , confirming that the 180kDa band represents the authentic ERBB4 protein. When troubleshooting, researchers should expect to observe ERBB4 at approximately 180kDa rather than its calculated weight of 147kDa.

What are common reasons for weak or no signal when detecting phosphorylated ERBB4 Y1284?

When experiencing weak or absent phospho-ERBB4 Y1284 signals, consider these common issues and solutions:

• Insufficient Phosphorylation:

  • Problem: Basal phosphorylation levels may be too low for detection

  • Solution: Stimulate cells with appropriate ligands (neuregulins, IGF) before lysis

  • Validation: Use MCF7+IGF as a positive control system

• Phosphatase Activity:

  • Problem: Phosphorylation sites are rapidly dephosphorylated during sample preparation

  • Solution: Include phosphatase inhibitors in all buffers (lysis, washing, etc.)

  • Technique: Keep samples cold and process quickly

• Antibody-Related Issues:

  • Problem: Improper antibody dilution or incubation conditions

  • Solution: Optimize antibody concentration (test range from 1:500 to 1:2000)

  • Consideration: Extend primary antibody incubation to overnight at 4°C

• Sample Preparation:

  • Problem: Inefficient protein extraction or denaturation

  • Solution: Ensure complete lysis with appropriate buffers containing detergents

  • Technique: Heat samples appropriately (95°C for 5 minutes) in reducing sample buffer

• Detection System Limitations:

  • Problem: Insufficient sensitivity of detection method

  • Solution: Use enhanced chemiluminescence (ECL) or more sensitive detection systems

  • Optimization: Increase exposure time (up to 90 seconds or more)

Implementing these troubleshooting approaches systematically will help identify and resolve the specific cause of weak phospho-ERBB4 Y1284 signals in your experimental system.

How can I distinguish between specific and non-specific bands when analyzing Phospho-ERBB4 (Y1284) Western blots?

Distinguishing between specific and non-specific bands is crucial for accurate interpretation of phospho-ERBB4 Y1284 Western blots:

• Molecular Weight Verification:

  • Specific band: Expect phospho-ERBB4 at approximately 180kDa (observed weight)

  • Non-specific bands: May appear at different molecular weights

  • Strategy: Use molecular weight markers and positive controls to identify the correct band

• Validation Controls:

  • Phosphatase treatment: Treat duplicate samples with lambda phosphatase—specific phospho-bands should disappear

  • Peptide competition: Pre-incubation with phospho-peptide immunogen should eliminate specific bands

  • Knockout/knockdown validation: Test samples with ERBB4 knockdown or knockout—specific bands should be reduced or absent

  • Transfection controls: Compare wild-type ERBB4 with Y1284F mutant transfected cells

• Stimulation Response:

  • Specific phospho-bands should increase in intensity after appropriate stimulation

  • Compare unstimulated vs. stimulated (e.g., with neuregulins or IGF) samples

  • Time-course experiments can confirm the dynamic nature of specific phosphorylation events

• Antibody Specificity Testing:

  • Use multiple antibodies recognizing different epitopes or phosphorylation sites

  • Compare monoclonal (more specific) vs. polyclonal (potentially less specific) antibodies

  • Different antibody clones may have different non-specific binding profiles

By systematically implementing these approaches, researchers can confidently identify specific phospho-ERBB4 Y1284 signals and avoid misinterpretation of non-specific bands.

How does Y1284 phosphorylation of ERBB4 contribute to cancer progression and therapeutic resistance?

Y1284 phosphorylation of ERBB4 plays complex roles in cancer biology through multiple mechanisms:

• Signaling Pathway Activation:

  • The phosphorylated Y1284 site serves as a docking platform for signaling adapters that activate MAPK pathways, promoting cancer cell proliferation

  • Different ERBB4 isoforms with phosphorylated Y1284 can activate distinct downstream pathways, including PI3K/AKT signaling in specific splice variants (JM-A CYT-1 and JM-B CYT-1)

  • These pathways contribute to cancer hallmarks including sustained proliferation, resistance to cell death, and metastatic potential

• Therapeutic Resistance Mechanisms:

  • Y1284 phosphorylation can activate compensatory signaling when other ERBB family members (e.g., EGFR, HER2) are inhibited by targeted therapies

  • The persistence of Y1284 phosphorylation during therapy may indicate pathway reactivation and emerging resistance

  • Cross-talk between ERBB4 Y1284-mediated signaling and other oncogenic pathways can circumvent single-agent targeted therapies

• Tissue-Specific Effects:

  • In breast cancer: ERBB4 Y1284 phosphorylation can have context-dependent effects (tumor-promoting in some subtypes, tumor-suppressive in others)

  • In lung cancer: May contribute to resistance to EGFR-targeted therapies

  • In neurological malignancies: Activates survival pathways due to ERBB4's role in neural development

Monitoring Y1284 phosphorylation status may serve as a biomarker for treatment response and resistance development, potentially guiding combination therapy approaches that target multiple ERBB family members or downstream pathways simultaneously.

What is the relationship between ERBB4 Y1284 phosphorylation and alternative splicing of ERBB4?

The relationship between ERBB4 Y1284 phosphorylation and alternative splicing represents a sophisticated regulatory mechanism:

• ERBB4 Splice Variants and Y1284 Accessibility:

  • ERBB4 has multiple splice variants affecting both the juxtamembrane domain (JM-a/JM-b) and the cytoplasmic domain (CYT-1/CYT-2)

  • The Y1284 phosphorylation site is present in all major ERBB4 isoforms, but its accessibility and signaling outcomes may differ between variants

  • JM-a variants undergo proteolytic processing more readily than JM-b variants, potentially affecting the cellular localization and signaling capacity of the phosphorylated Y1284 site

• Differential Signaling Outcomes:

  • CYT-1 isoforms contain a PI3K binding site absent in CYT-2 variants

  • Y1284 phosphorylation in CYT-1 variants can cooperate with PI3K binding to enhance AKT activation and anti-apoptotic signaling

  • Y1284 phosphorylation in CYT-2 variants primarily activates MAPK pathways without the PI3K/AKT component

• Tissue-Specific Expression Patterns:

  • Different tissues preferentially express specific ERBB4 splice variants

  • This creates tissue-specific consequences of Y1284 phosphorylation

  • For example, mammary tissue predominantly expresses JM-a CYT-1 variants where Y1284 phosphorylation contributes to differentiation signaling

This complex interplay between alternative splicing and Y1284 phosphorylation allows for fine-tuning of ERBB4 signaling in different cellular contexts and helps explain the diverse biological roles of ERBB4 across different tissues and developmental stages.

How can Phospho-ERBB4 (Y1284) antibodies be used in multiplexed phosphoproteomic analyses?

Incorporating Phospho-ERBB4 (Y1284) antibodies into multiplexed phosphoproteomic analyses enables comprehensive signaling pathway characterization:

• Multiplex Immunoassay Approaches:

  • Antibody microarrays: Include Phospho-ERBB4 (Y1284) antibodies alongside other phospho-specific antibodies

  • Multiplex Western blotting: Use differently labeled secondary antibodies to detect multiple phosphoproteins simultaneously

  • Bead-based assays: Couple Phospho-ERBB4 (Y1284) antibodies to spectrally distinct beads for flow cytometry-based detection

• Mass Spectrometry Integration:

  • Immunoprecipitation with Phospho-ERBB4 (Y1284) antibodies followed by mass spectrometry to identify interacting partners

  • Targeted mass spectrometry (MRM/PRM) using heavy-labeled phosphopeptide standards corresponding to the Y1284 site

  • SILAC or TMT-based quantification to measure changes in Y1284 phosphorylation across different conditions

• Single-Cell Phosphoproteomic Applications:

  • Multiplex immunofluorescence using Phospho-ERBB4 (Y1284) antibodies with other signaling markers

  • Mass cytometry (CyTOF) integration using metal-labeled Phospho-ERBB4 (Y1284) antibodies

  • Spatial proteomics approaches to map Y1284 phosphorylation in tissue contexts

• Technical Considerations:

  • Validate antibody compatibility with fixation protocols used in multiplexed approaches

  • Ensure minimal cross-reactivity with other phospho-epitopes

  • Optimize antibody concentrations to achieve balanced signal across all analytes

  • Include appropriate phosphorylation controls for each target

These approaches allow researchers to position ERBB4 Y1284 phosphorylation within broader signaling networks and to understand how this specific phosphorylation event coordinates with other phosphorylation-dependent signaling processes in normal and disease states.

What are the methodological approaches for studying ERBB4 Y1284 phosphorylation dynamics in live cells?

Investigating ERBB4 Y1284 phosphorylation dynamics in live cells requires specialized techniques:

• Genetically Encoded Biosensors:

  • FRET-based biosensors incorporating the Y1284 region between fluorescent proteins

  • Phosphorylation-dependent conformational changes alter FRET efficiency

  • Design considerations: Optimize linker length and fluorophore pairs for maximum dynamic range

  • Analysis methods: Ratiometric imaging with appropriate controls for photobleaching and spectral bleed-through

• Phospho-Specific Cellular Probes:

  • Membrane-permeable peptide probes with phosphorylation-sensitive fluorophores

  • Cell-penetrating antibody fragments (e.g., ScFvs) derived from Phospho-ERBB4 (Y1284) antibodies

  • Optimization requirements: Balance probe concentration, incubation time, and signal-to-background ratio

• Engineered ERBB4 Constructs:

  • ERBB4-FP fusion proteins for tracking localization concurrent with biochemical phosphorylation assays

  • Split luciferase complementation systems triggered by phosphorylation-dependent protein interactions

  • CRISPR-modified endogenous ERBB4 with minimal tags for physiological relevance

• Temporal Analysis Approaches:

  • High-speed confocal microscopy to capture rapid phosphorylation events

  • Optogenetic control of ERBB4 activation for precise temporal stimulation

  • Microfluidic systems for controlled ligand delivery and temporal resolution of phosphorylation

• Quantification Methods:

  • Single-cell analysis to account for cell-to-cell variability

  • Computational approaches for tracking phosphorylation kinetics across subcellular compartments

  • Correlative light and electron microscopy to link phosphorylation events to ultrastructural features

These methodologies provide insights into the spatiotemporal dynamics of ERBB4 Y1284 phosphorylation that are not accessible through traditional fixed-cell or biochemical approaches, revealing how this phosphorylation event coordinates signaling in real time.

How can Phospho-ERBB4 (Y1284) antibodies be used to investigate neuregulin-dependent signaling in cardiac development?

Phospho-ERBB4 (Y1284) antibodies offer powerful tools for investigating neuregulin-dependent signaling in cardiac development:

• Developmental Stage Analysis:

  • Immunohistochemistry using Phospho-ERBB4 (Y1284) antibodies on cardiac tissue sections at different embryonic stages

  • Western blot analysis of heart lysates from various developmental timepoints

  • Correlation of phosphorylation patterns with cardiomyocyte differentiation markers

• Cellular Response Characterization:

  • Primary cardiomyocyte cultures treated with neuregulins to assess Y1284 phosphorylation kinetics

  • Comparison between proliferating and differentiating cardiomyocytes

  • Co-localization studies with other cardiac signaling components using multiplexed immunofluorescence

• Functional Correlation Studies:

  • Inhibition of Y1284 phosphorylation (via ERBB4 inhibitors or Y1284F mutants) and assessment of cardiac developmental outcomes

  • Correlation between phosphorylation levels and functional parameters (contractility, calcium handling)

  • Integration with transcriptomic data to link Y1284 phosphorylation to cardiac gene expression programs

• Disease Model Applications:

  • Comparison of Y1284 phosphorylation in normal versus congenital heart disease models

  • Assessment in cardiac stress conditions (hypoxia, pressure overload)

  • Evaluation in regenerative medicine applications involving cardiomyocyte proliferation

This methodological approach enables researchers to establish causal relationships between ERBB4 Y1284 phosphorylation and specific aspects of cardiac development, potentially identifying therapeutic targets for congenital heart diseases or cardiac regeneration strategies.

What are the key considerations when using Phospho-ERBB4 (Y1284) antibodies in patient-derived samples for personalized medicine?

Using Phospho-ERBB4 (Y1284) antibodies in patient-derived samples requires careful methodological considerations:

• Sample Collection and Preservation:

  • Immediate fixation/freezing is crucial to preserve phosphorylation status

  • Standardized protocols must be established for consistent results across samples

  • Consider using phosphatase inhibitors during collection and processing

  • Document ischemia time as this affects phosphorylation integrity

• Tissue Heterogeneity Management:

  • Microdissection techniques to isolate relevant cell populations

  • Single-cell approaches to account for cellular heterogeneity

  • Spatial mapping of phosphorylation patterns within heterogeneous tissues

  • Correlation with cell-type specific markers

• Quantification and Standardization:

  • Establish quantitative scoring systems (H-score, digital image analysis)

  • Include calibration standards across batches

  • Use phosphorylation-insensitive ERBB4 antibodies to normalize for total ERBB4 expression

  • Implement quality control metrics for phosphorylation preservation

• Clinical Correlation Methods:

  • Correlate Y1284 phosphorylation with treatment response data

  • Longitudinal sampling to track changes during treatment

  • Integration with other molecular biomarkers

  • Standardized reporting formats for clinical decision support

• Technical Validation Requirements:

  • Confirm antibody specificity in patient-derived tissues

  • Establish thresholds for positivity based on clinical outcomes

  • Cross-validate with orthogonal methods (e.g., mass spectrometry)

  • Participate in inter-laboratory standardization initiatives

These methodological considerations are essential for developing Phospho-ERBB4 (Y1284) as a reliable biomarker for personalized treatment decisions, particularly in cancer therapies targeting ERBB family members.

How can mass spectrometry complement antibody-based detection of ERBB4 Y1284 phosphorylation?

Mass spectrometry (MS) provides complementary approaches to antibody-based detection of ERBB4 Y1284 phosphorylation:

• Unbiased Phosphosite Identification:

  • Shotgun phosphoproteomics can confirm Y1284 phosphorylation without antibody bias

  • Enables discovery of novel phosphorylation sites co-regulated with Y1284

  • Characterizes phosphorylation stoichiometry across multiple ERBB4 phosphosites

  • Identifies previously uncharacterized phosphorylation sites near Y1284

• Quantitative Analysis Methods:

  • Targeted MS approaches (MRM/PRM) for absolute quantification of Y1284 phosphopeptides

  • SILAC, TMT, or label-free quantification for relative abundance measurements

  • Heavy-labeled phosphopeptide standards enable direct quantification of phosphorylation stoichiometry

  • Time-course experiments with higher temporal resolution than possible with antibodies

• Integrated Approaches:

  • Phospho-ERBB4 (Y1284) antibody immunoprecipitation followed by MS analysis

  • Identification of proteins that interact specifically with phosphorylated Y1284

  • Comprehensive mapping of the phosphorylated ERBB4 interactome

  • Correlation between Y1284 phosphorylation and other post-translational modifications

• Technical Considerations:

  • Sample preparation optimization for phosphopeptide enrichment

  • MS instrument parameters for improved sensitivity and specificity

  • Bioinformatic workflows for phosphosite assignment and quantification

  • Integration of MS and antibody-based data in systems biology approaches

This complementary approach leverages the strengths of both techniques: antibodies provide high sensitivity and spatial information, while MS offers unbiased detection, multiplexing capability, and absolute quantification potential.

What emerging technologies will enhance Phospho-ERBB4 (Y1284) detection and functional characterization?

Several emerging technologies promise to revolutionize Phospho-ERBB4 (Y1284) research:

• Advanced Imaging Technologies:

  • Super-resolution microscopy: Nanoscale visualization of phosphorylation-dependent protein interactions

  • Expansion microscopy: Physical magnification of specimens for improved phospho-epitope detection

  • Light-sheet microscopy: Rapid 3D imaging of phosphorylation dynamics in organoids or embryos

  • Correlative light and electron microscopy: Linking phosphorylation events to ultrastructural features

• Single-Cell Technologies:

  • Single-cell phosphoproteomics: Cell-specific phosphorylation profiles in heterogeneous populations

  • Spatial transcriptomics integration: Correlating Y1284 phosphorylation with gene expression in tissue contexts

  • CyTOF and imaging mass cytometry: Multiplexed detection of phosphorylation events in single cells

  • Microfluidic approaches for high-throughput single-cell signaling analysis

• Artificial Intelligence Applications:

  • Deep learning for phosphorylation site prediction and functional annotation

  • Computer vision algorithms for automated quantification of phosphorylation patterns

  • Integrated systems biology modeling of phosphorylation networks

  • AI-assisted experimental design for phosphorylation studies

• Genome Engineering Approaches:

  • CRISPR-based tagging of endogenous ERBB4 for physiological studies

  • Base editing to introduce specific mutations affecting Y1284 phosphorylation

  • Optogenetic control of ERBB4 signaling pathways

  • Synthetic biology circuits responsive to Y1284 phosphorylation

These technological advances will enable more precise, comprehensive, and physiologically relevant characterization of ERBB4 Y1284 phosphorylation in various biological and disease contexts.

How does phosphorylation at Y1284 coordinate with other post-translational modifications of ERBB4?

The coordination between Y1284 phosphorylation and other post-translational modifications creates a sophisticated regulatory network:

• Phosphorylation Code Integration:

  • Y1284 phosphorylation may influence or be influenced by other phosphorylation events on ERBB4

  • Hierarchical phosphorylation patterns may exist where Y1284 phosphorylation primes for additional modifications

  • Different kinases and phosphatases may regulate specific subsets of ERBB4 phosphorylation sites

• Cross-Modal PTM Interactions:

  • Ubiquitination: Y1284 phosphorylation may regulate receptor ubiquitination and degradation

  • Glycosylation: May affect receptor localization and consequently Y1284 phosphorylation accessibility

  • Proteolytic processing: Y1284 phosphorylation may influence susceptibility to γ-secretase cleavage

  • SUMOylation: May regulate nuclear localization of ERBB4 fragments containing phosphorylated Y1284

• Spatiotemporal Coordination:

  • Different subcellular compartments may feature distinct patterns of Y1284 phosphorylation and other PTMs

  • Temporal sequences of modifications may create signaling waves that regulate different cellular processes

  • PTM patterns may vary with cell cycle phase or differentiation state

• Methodological Approaches for Integrated Analysis:

  • Multi-antibody approaches targeting different modifications simultaneously

  • Mass spectrometry analysis of sequential immunoprecipitations

  • Proximity ligation assays to detect co-occurrence of modifications

  • Computational modeling of PTM interplay based on experimental data

Understanding this complex interplay will provide a more complete picture of ERBB4 regulation and potentially reveal new therapeutic strategies targeting specific PTM combinations.

What are the best practices for integrating Phospho-ERBB4 (Y1284) data into systems biology models of receptor tyrosine kinase signaling?

Integrating Phospho-ERBB4 (Y1284) data into systems biology models requires methodical approaches:

• Data Collection Standardization:

  • Standardized protocols for phosphorylation quantification across experimental systems

  • Time-resolved measurements capturing kinetics of Y1284 phosphorylation

  • Dose-response relationships between ligand concentrations and Y1284 phosphorylation

  • Multi-omics data collection (phosphoproteomics, transcriptomics, metabolomics) in parallel

• Computational Modeling Frameworks:

  • Ordinary differential equation (ODE) models capturing phosphorylation kinetics

  • Bayesian network approaches to infer causal relationships

  • Agent-based models for spatial aspects of receptor activation and phosphorylation

  • Integration of Y1284 phosphorylation nodes into existing RTK signaling models

• Network Analysis Approaches:

  • Identification of Y1284 phosphorylation-dependent protein interactions

  • Pathway enrichment analysis downstream of Y1284 phosphorylation

  • Network perturbation analysis to identify key regulatory nodes

  • Comparison of network structures across different cellular contexts

• Validation and Refinement Strategies:

  • Experimental validation of model predictions using targeted interventions

  • Sensitivity analysis to identify critical parameters in Y1284 phosphorylation regulation

  • Iterative model refinement based on new experimental data

  • Cross-validation across different cell types and experimental conditions