Recombinant Chicken Amyloid beta A4 precursor protein-binding family B member 1-interacting protein (APBB1IP)

What is APBB1IP and what are its primary functions in immune systems?

APBB1IP was initially identified as a binding partner of amyloid β (A4) precursor protein-binding, family B, member 1 (APBB1) and was subsequently found to interact with the small guanosine triphosphatase (GTPase) Rap1. It belongs to the MRL (Mig-10/RIAM/Lamellipodin) family of adaptor proteins, characterized by a proline-rich region at the C terminus and a highly conserved pattern of 27 amino acids in a predicted coiled-coil region immediately N-terminal to the RA domain .

The primary functions of APBB1IP include:

• Regulation of leukocyte recruitment

• Facilitation of pathogen clearance through complement-mediated phagocytosis

• Serving as an intrinsic element of integrin activation machinery

• Required for Rap1-induced affinity changes in β1 and β2 integrins in T cells

• Involvement in Rap1-mediated activation of αIIbβ3 integrin in platelets

Methodologically, researchers investigating APBB1IP functions should implement knockout studies, protein-protein interaction assays, and immune cell migration assays to comprehensively evaluate its role in immune regulation.

How does APBB1IP expression correlate with immune cell infiltration in different tissues?

APBB1IP expression has been found to significantly correlate with immune cell infiltration in multiple tissue types. Research has demonstrated that APBB1IP expression is negatively correlated with tumor purity in most cancer types, with exceptions in CHOL, DLBC, KIRC, KIRP, MESO, THCA, THYM, UCS, and UVM .

To methodologically assess this correlation, researchers should:

• Utilize bioinformatic tools like TIMER database to analyze tumor-infiltrating immune cells

• Apply statistical deconvolution methods to infer immune cell abundance from gene expression profiles

• Calculate Spearman correlation coefficients between APBB1IP expression and immune cell markers

• Perform multivariate analyses to control for confounding factors

The research indicates that APBB1IP expression in most cancers markedly increased the infiltration of immune cells, especially in BRCA, CESC, HNSC, PRAD, SKCM, TGCT, and UCEC. Particularly strong correlations have been observed with CD8+ T cells, natural killer (NK) cells, and various immune regulators .

What experimental methods are most effective for producing and purifying recombinant APBB1IP protein?

Based on successful recombinant protein production in related research, the following methodological approach is recommended for recombinant APBB1IP production:

• Expression System Selection:

  • Baculovirus expression systems have demonstrated success for producing functional recombinant proteins of complex structure, as evidenced by the effective production of recombinant fiber-1 protein in viral research

  • Alternatively, E. coli or mammalian expression systems may be employed depending on post-translational modification requirements

• Purification Protocol:

  • Implement affinity chromatography using appropriate tags (His-tag, GST-tag)

  • Follow with size-exclusion chromatography to enhance purity

  • Validate protein identity and purity through Western blotting and mass spectrometry

• Quality Control Measures:

  • Assess proper folding through circular dichroism

  • Verify functional activity through binding assays with known interaction partners like Rap1

  • Test immunogenicity in appropriate animal models if applicable

The baculovirus expression system has proven particularly effective, as demonstrated in the successful production of immunogenic recombinant proteins that induced high levels of neutralizing antibodies in chicken models .

How can researchers evaluate the immunogenic potential of recombinant APBB1IP?

Evaluation of immunogenic potential requires systematic testing using the following methodological approach:

• Immunization Protocol Design:

  • Administer purified recombinant APBB1IP with appropriate adjuvants (e.g., incomplete Freund's adjuvant)

  • Implement a prime-boost strategy with intervals of 14 days between immunizations

  • Include appropriate control groups (adjuvant-only and non-immunized)

• Antibody Response Assessment:

  • Collect serum samples at regular intervals (weekly for 10+ weeks post-immunization)

  • Determine neutralizing antibody titers using virus neutralization assays

  • Analyze antibody persistence through longitudinal sampling

• Cellular Immune Response Analysis:

  • Isolate peripheral blood mononuclear cells from immunized subjects

  • Perform ELISpot assays to quantify APBB1IP-specific T cell responses

  • Assess cytokine profiles through flow cytometry

Research with similar recombinant proteins has demonstrated that neutralizing antibody levels may not increase significantly after initial immunization but can increase substantially following booster immunization. In studies with recombinant proteins in chicken models, statistically significant antibody titers were maintained for at least 10 weeks after the second immunization .

What is the prognostic value of APBB1IP expression in different cancer types?

APBB1IP expression has varying prognostic implications across different cancer types, necessitating a cancer-specific analytical approach:

Researchers should be aware that APBB1IP's prognostic significance varies significantly by cancer type and should carefully interpret results in the context of specific tumor microenvironments .

What methods are recommended for studying APBB1IP protein-protein interactions in avian systems?

To effectively study APBB1IP protein interactions in avian systems, researchers should implement the following methodological approaches:

Research has shown that APBB1IP interacts with immune-related proteins including RAP1A/B, TLN1/2, and VCL, forming an interaction network that may be critical for immune function. These interactions should be validated in avian systems to determine conservation across species .

How can cross-species comparative analysis of APBB1IP contribute to understanding its conserved functions?

Cross-species analysis of APBB1IP requires sophisticated comparative genomics and functional validation approaches:

• Comparative Sequence Analysis:

  • Perform multiple sequence alignment of APBB1IP across species (human, mouse, chicken, etc.)

  • Identify conserved domains, motifs, and regulatory elements

  • Calculate evolutionary conservation scores for each amino acid position

• Functional Domain Conservation Assessment:

  • Compare protein domain architecture across species

  • Identify species-specific insertions/deletions or domain rearrangements

  • Assess conservation of post-translational modification sites

• Cross-Species Experimental Validation:

  • Generate species-specific APBB1IP constructs for functional testing

  • Perform complementation studies by expressing avian APBB1IP in mammalian systems and vice versa

  • Use CRISPR/Cas9 technology to introduce conserved mutations across species

• Systems-Level Comparative Analysis:

  • Implement systems genetics approaches similar to those used in cross-species studies linking APBB1IP to neuropsychiatric phenotypes

  • Compare APBB1IP-associated gene networks across species

  • Identify conserved pathways that may represent fundamental biological functions

This approach can reveal evolutionarily conserved functions while highlighting species-specific adaptations, potentially uncovering novel insights into APBB1IP's role in immune regulation and disease processes across vertebrates.

What are the methodological considerations for studying APBB1IP's role in avian tumor microenvironments?

Investigating APBB1IP in avian tumor microenvironments requires specialized methodological approaches:

• Avian-Specific Tumor Model Development:

  • Establish appropriate avian tumor models (e.g., avian leukosis virus-induced tumors)

  • Develop methods for consistent tumor induction and monitoring

  • Create APBB1IP knockout or overexpression systems in avian cells

• Tumor Microenvironment Characterization:

  • Implement multiparameter flow cytometry panels optimized for avian immune cells

  • Develop spatial transcriptomics or multiplex immunohistochemistry protocols for avian tissues

  • Assess tumor purity and immune cell infiltration using computational deconvolution methods

• Functional Assessment Protocol:

  • Measure APBB1IP expression correlation with tumor purity across different avian tumor types

  • Analyze relationship between APBB1IP expression and infiltration of specific immune cell subsets

  • Assess impact of APBB1IP modulation on tumor progression and immune infiltration

• Translational Considerations:

  • Compare findings between avian models and human cancer data

  • Identify conserved mechanisms that could inform therapeutic strategies

  • Develop targeted approaches based on APBB1IP's role in immune cell recruitment

Research in human cancers has shown that APBB1IP expression is significantly negatively correlated with tumor purity in most cancer types, suggesting it may influence immune cell infiltration in the tumor microenvironment . Similar mechanisms may exist in avian tumors, providing valuable comparative insights.

How can researchers develop and validate APBB1IP-targeted interventions in avian disease models?

Development of APBB1IP-targeted interventions requires a systematic approach:

• Target Validation Strategy:

  • Confirm APBB1IP expression and function in relevant avian tissues

  • Establish causal relationship between APBB1IP and disease phenotypes through knockdown/knockout studies

  • Identify critical binding partners or downstream effectors as potential co-targets

• Intervention Development Methodology:

  • Recombinant Protein Production: Optimize expression and purification of functional recombinant APBB1IP using baculovirus expression systems

  • Antibody Development: Generate and characterize antibodies targeting specific APBB1IP domains

  • Small Molecule Screen: Identify compounds that modulate APBB1IP-protein interactions

• Efficacy Assessment Protocol:

  • Develop appropriate challenge models (similar to viral challenge models used in vaccine testing)

  • Implement meaningful clinical and laboratory endpoints

  • Evaluate both therapeutic and prophylactic potential

• Safety and Dosing Studies:

  • Determine optimal dosing regimens through dose-escalation studies

  • Monitor for potential immune-related adverse events

  • Assess long-term effects of APBB1IP modulation

Research with recombinant proteins in chickens has demonstrated that proper immunization protocols can induce high levels of protective antibodies that persist for at least 10 weeks, providing a methodological framework for intervention development .

What computational approaches are recommended for predicting the functional impact of APBB1IP genetic variants in avian species?

Predicting functional impacts of APBB1IP variants requires sophisticated computational methodologies:

• Variant Identification Protocol:

  • Implement whole-genome or targeted sequencing of APBB1IP across avian populations

  • Develop appropriate bioinformatic pipelines for variant calling in avian genomes

  • Filter variants based on quality metrics and population frequency

• Functional Impact Prediction Methods:

  • Apply multiple prediction algorithms (SIFT, PolyPhen, CADD, etc.) adapted for avian proteins

  • Implement protein structure modeling to assess variant effects on protein folding and stability

  • Use machine learning approaches trained on known functional variants

• Experimental Validation Design:

  • Prioritize variants based on conservation, predicted impact, and location in functional domains

  • Implement site-directed mutagenesis to introduce variants into expression constructs

  • Assess functional consequences through binding assays, cell migration studies, and signaling pathway analysis

• Population-Level Analysis:

  • Correlate variant frequencies with phenotypic traits or disease susceptibility

  • Implement genome-wide association studies in diverse avian populations

  • Apply systems genetics approaches as used in cross-species studies

This comprehensive approach enables researchers to identify functionally significant APBB1IP variants in avian species and understand their potential role in immune function and disease susceptibility.