Recombinant Cricetulus griseus NADH dehydrogenase [ubiquinone] 1 beta subcomplex subunit 11, mitochondrial (NDUFB11)

How is NDUFB11 expressed in normal versus disease states?

Gene expression analysis reveals significant differences in NDUFB11 expression between normal and disease states. In studies focused on atherosclerosis and chronic stress, NDUFB11 shows distinctive expression patterns:

• NDUFB11 is consistently downregulated in atherosclerosis and venous thrombosis samples

• NDUFB11 is upregulated in normal vascular tissue samples

• Gene expression heatmaps demonstrate that NDUFB11 is lowly expressed in samples with atherosclerosis accompanied by chronic stress and highly expressed in normal samples

This differential expression pattern has important implications for disease mechanisms and potential therapeutic approaches. The downregulation of NDUFB11 in pathological conditions suggests that loss of normal NDUFB11 function may contribute to disease development, particularly in vascular disorders .

What model systems are commonly used to study NDUFB11?

Several model systems have proven effective for studying NDUFB11:

• Cell Culture Systems:

  • HeLa cells have been used for shRNA-mediated knockdown studies to assess NDUFB11's role in complex I assembly and cell survival

  • Endothelial cells for studying NDUFB11's involvement in atherosclerosis and vascular conditions

• Animal Models:

  • Chinese hamster (Cricetulus griseus) systems, as evidenced by recombinant protein production

• Disease Models:

  • Atherosclerosis and venous thrombosis models to study NDUFB11 dysregulation

  • Microphthalmia with linear skin defects (MLS) syndrome models to investigate the impact of NDUFB11 mutations

• Expression Systems:

  • E. coli for recombinant protein production, as indicated by the commercial recombinant NDUFB11 protein

Each model system offers unique advantages for investigating different aspects of NDUFB11 biology, from molecular mechanisms to disease relevance.

What are the standard methods for detecting NDUFB11 expression?

Standard methods for detecting NDUFB11 expression include:

• Western Blotting: Used to measure protein levels, as mentioned in multiple studies validating NDUFB11 expression changes in disease states

• RT-PCR followed by Sanger sequencing: Used to analyze mRNA expression and detect mutations

• Bioinformatic Analysis of Gene Expression Datasets: Including differential gene expression analysis (DEGs) and weighted gene co-expression network analysis (WGCNA)

• Gene Expression Heatmaps: Used to visualize expression differences between disease and normal samples

These methods have been crucial in establishing NDUFB11's role in various diseases and determining its potential as a biomarker or therapeutic target.

How does NDUFB11 contribute to mitochondrial complex I assembly and function?

NDUFB11 plays a critical role in mitochondrial complex I assembly and function, as demonstrated through knockdown experiments. When NDUFB11 is depleted using shRNA in HeLa cells, researchers observed:

• Disrupted Complex I Assembly: NDUFB11 knockdown prevents the proper formation of complete complex I structures

• Reduced Complex I Activity: Functional assays show decreased NADH:ubiquinone oxidoreductase activity

• Impaired Cellular Growth and Survival: Cells with reduced NDUFB11 show compromised viability, indicating its essential nature

While the exact molecular mechanisms remain under investigation, NDUFB11 likely functions in the early or intermediate stages of complex I assembly. As a supernumerary subunit, it may serve as a scaffold during assembly, stabilize intermediate complexes, or facilitate the incorporation of other subunits . Importantly, mutations in NDUFB11 demonstrate that its proper function is essential not only for mitochondrial energy production but also for normal development, as evidenced by its association with MLS syndrome .

What is the role of NDUFB11 in atherosclerosis and chronic stress?

Recent research has revealed significant associations between NDUFB11 and both atherosclerosis and chronic stress:

• Expression Patterns:

  • NDUFB11 is consistently downregulated in atherosclerosis and venous thrombosis samples

  • Upregulated in normal vascular tissue samples

  • Shows altered expression patterns in chronic stress conditions

• Disease Associations:

  • Comparative Toxicogenomics Database (CTD) analysis identified NDUFB11 as associated with:

    • Arterial diseases

    • Atherosclerosis

    • Arteritis

    • Venous thrombosis formation

    • Venous thromboembolism

    • Pain (potentially related to chronic stress pathways)

• Functional Implications:

  • Downregulation of NDUFB11 may contribute to mitochondrial dysfunction in vascular cells

  • Impaired energy metabolism in vascular smooth muscle and endothelial cells

  • Potential link to inflammation and oxidative stress in vascular pathology

• Prognostic Value:

  • Lower NDUFB11 expression associates with poorer prognosis in vascular diseases

  • Potential biomarker for atherosclerosis progression and venous thrombotic events

These findings suggest that NDUFB11 downregulation may be a key mechanism in atherosclerosis pathogenesis, potentially affecting mitochondrial function in vascular tissues and contributing to disease progression .

How do mutations in NDUFB11 impact cellular bioenergetics?

Mutations in NDUFB11 significantly impact cellular bioenergetics through multiple mechanisms:

• Complex I Assembly Defects:

  • Truncating mutations (p.Arg88* and p.Arg134Serfs*3) lead to loss of functional NDUFB11 protein

  • Prevents proper assembly of complex I intermediate structures

  • Results in reduced levels of fully assembled complex I

• Oxidative Phosphorylation Impairment:

  • Reduced complex I activity compromises the first entry point of electrons into the respiratory chain

  • Decreases proton pumping across the inner mitochondrial membrane

  • Lowers the proton gradient necessary for ATP synthase function

• Developmental Consequences:

  • Severe bioenergetic defects in embryonic development due to NDUFB11 mutations associate with:

    • Microphthalmia with linear skin defects (MLS) syndrome

    • Neurological and cardiac abnormalities

    • Strong negative selection against cells expressing mutant NDUFB11 alleles, resulting in extremely skewed X-chromosome inactivation patterns in female carriers

These impacts highlight the critical role of NDUFB11 in maintaining proper mitochondrial function and cellular energy homeostasis, with significant consequences for development and disease when disrupted .

What techniques are most effective for studying NDUFB11 protein-protein interactions?

Several complementary techniques have proven effective for studying NDUFB11 interactions with other proteins:

• Protein-Protein Interaction (PPI) Network Analysis:

  • STRING database and Cytoscape software for computational prediction of interaction networks

  • Identification of functional protein clusters containing NDUFB11 (as demonstrated in atherosclerosis studies)

  • Core gene clusters and central genes can be recognized using various algorithms (maximum clique centrality, maximum neighborhood component, density of maximum neighborhood component, edge percolated component)

• Complex I Subunit Interaction Studies:

  • Analysis of NDUFB11's interactions with other complex I components

  • Identification of assembly factors that interact with NDUFB11 during complex I biogenesis

  • Investigation of how supernumerary subunits like NDUFB11 interact with core subunits

• Western Blot Experiments:

  • Verification of protein expression and interactions at the cellular level

  • Confirmation of changes in interaction patterns in disease states

These approaches have been instrumental in identifying NDUFB11 as a core gene in disease processes and understanding its role within the mitochondrial respiratory complex I.

What are the optimal conditions for expressing recombinant NDUFB11?

Based on available data for recombinant Cricetulus griseus NDUFB11 expression, the following conditions are recommended:

• Expression System:

  • E. coli has been successfully used as an expression host for NDUFB11

• Storage and Stability Parameters:

  • Liquid form: Store at -20°C/-80°C with a shelf life of approximately 6 months

  • Lyophilized form: Store at -20°C/-80°C with a shelf life of approximately 12 months

  • Avoid repeated freezing and thawing

  • Working aliquots can be stored at 4°C for up to one week

• Reconstitution Protocol:

  • Briefly centrifuge vial before opening to bring contents to the bottom

  • Reconstitute in deionized sterile water to a concentration of 0.1-1.0 mg/mL

  • Add 5-50% glycerol (final concentration) for long-term storage

  • A final glycerol concentration of 50% is commonly used

• Purification Considerations:

  • Recombinant NDUFB11 with >85% purity can be achieved using SDS-PAGE

  • Tag type will be determined during the manufacturing process

These conditions are based on established protocols for recombinant Cricetulus griseus NDUFB11 and may require optimization for specific experimental applications.

How can researchers effectively design knockdown experiments for NDUFB11?

When designing knockdown experiments for NDUFB11, researchers should consider:

• Cell Type Selection:

  • HeLa cells have been successfully used for NDUFB11 knockdown studies

  • Cell types relevant to disease models (e.g., endothelial cells for atherosclerosis studies)

  • Consider mitochondrial content and dependence on oxidative phosphorylation

• Knockdown Approach:

  • shRNA has been demonstrated effective for NDUFB11 knockdown

  • Design experiments to assess:

    • Complex I assembly

    • Cell growth and survival

    • Mitochondrial function

    • Disease-specific phenotypes

• Important Considerations:

  • Complete knockout may be lethal based on evidence that NDUFB11 is essential for cell growth and survival

  • Include appropriate controls (scrambled shRNA)

  • Validate knockdown efficiency at both mRNA and protein levels

  • Consider X-linked nature of NDUFB11 in mammals

• Phenotypic Assessment:

  • Monitor cellular growth rates and viability

  • Assess mitochondrial function (membrane potential, respiration, ATP production)

  • Evaluate complex I assembly and activity

  • Measure specific effects on vascular cells when studying atherosclerosis models

These design considerations ensure robust and interpretable results when investigating NDUFB11 function through knockdown approaches.

What bioinformatic approaches are most valuable for NDUFB11 research?

Several bioinformatic approaches have proven particularly valuable for NDUFB11 research:

• Differential Expression Analysis:

  • Identifying NDUFB11 as differentially expressed in disease states like atherosclerosis and venous thrombosis

  • Quantifying expression changes between normal and pathological conditions

• Network Analysis:

  • Weighted Gene Co-expression Network Analysis (WGCNA) to identify modules of co-expressed genes including NDUFB11

  • Protein-Protein Interaction (PPI) network construction and analysis using STRING database and Cytoscape software

  • Identification of core gene clusters containing NDUFB11

• Functional Enrichment Analysis:

  • Gene Ontology (GO) analysis showing enrichment in catabolic processes, organic acid metabolism processes, carboxylic acid metabolism processes

  • KEGG pathway analysis revealing enrichment in metabolic pathways, fatty acid metabolism, pentose phosphate pathway, glycolysis/gluconeogenesis, fructose and mannose metabolism

  • Gene Set Enrichment Analysis (GSEA)

• Disease Association Analysis:

  • Comparative Toxicogenomics Database (CTD) to identify diseases most related to NDUFB11, including arterial diseases, atherosclerosis, arteritis, venous thrombosis, and venous thromboembolism

  • Linking NDUFB11 expression patterns to specific pathological conditions

• Visualization Techniques:

  • Gene expression heatmaps to display NDUFB11 expression across multiple samples

  • Venn diagrams to identify core genes across multiple analyses

These bioinformatic approaches have been instrumental in establishing NDUFB11's role in disease processes and identifying it as a potential biomarker and therapeutic target.

How should researchers approach validating NDUFB11 findings in disease models?

For robust validation of NDUFB11 findings in disease models, researchers should implement a multi-faceted approach:

• Multi-level Validation Strategy:

  • Genomic: Confirm altered gene expression using multiple techniques (RT-PCR, RNA-seq)

  • Protein: Verify expression changes at the protein level using Western blot

  • Functional: Assess the impact on mitochondrial complex I assembly and activity

  • Cellular: Evaluate consequences for cell growth, survival, and metabolism

  • Tissue: Examine expression in relevant disease tissues (e.g., atherosclerotic plaques)

• Cross-platform Validation:

  • Combine data from multiple gene expression datasets (e.g., GSE48000, GSE57691)

  • Compare results across different experimental platforms

  • Integrate findings from diverse methodological approaches

• Disease-specific Approaches:

  • For atherosclerosis: Validate in vascular cells and atherosclerotic tissue samples

  • For MLS syndrome: Confirm in appropriate developmental models

  • For chronic stress: Validate in stress-exposure models

• Functional Validation:

  • Perform knockdown/overexpression experiments to confirm causality

  • Assess rescue effects when reintroducing wild-type NDUFB11

  • Evaluate downstream consequences on known pathways

This comprehensive validation approach ensures that findings regarding NDUFB11's role in disease processes are robust and reproducible across multiple experimental systems.

How can researchers analyze NDUFB11 expression data in disease states?

Researchers can employ several approaches to analyze NDUFB11 expression data in disease states:

• Differential Expression Analysis:

  • Compare NDUFB11 expression between disease and control samples

  • Apply proper normalization techniques for specific platforms

  • Consider fold change and statistical significance thresholds

• Co-expression Network Analysis:

  • Implement Weighted Gene Co-expression Network Analysis (WGCNA) to identify modules of co-expressed genes

  • Determine which modules contain NDUFB11 and their correlation with disease traits

  • Identify gene clusters functionally related to NDUFB11

• Expression Pattern Visualization:

  • Generate gene expression heatmaps to display NDUFB11 expression across multiple samples

  • Cluster samples based on expression patterns

  • Integrate with clinical data when available

• Pathway Analysis:

  • Perform GO and KEGG pathway analysis for gene sets containing NDUFB11

  • Use GSEA to identify pathways enriched in disease states

  • Focus on mitochondrial, metabolic, and disease-specific pathways

• Immune Infiltration Analysis:

  • Apply tools like CIBERSORT for calculating immune cell infiltration in disease tissues

  • Correlate immune parameters with NDUFB11 expression

These approaches have successfully identified NDUFB11 as a key gene in conditions like atherosclerosis and venous thrombosis, highlighting its potential as a biomarker and therapeutic target .

What disease associations have been identified for NDUFB11?

NDUFB11 has been associated with several distinct disease conditions:

• Vascular Diseases:

  • Atherosclerosis: NDUFB11 is downregulated in atherosclerotic tissues

  • Venous thrombosis: Decreased expression in thrombotic conditions

  • Arteritis: Identified through Comparative Toxicogenomics Database (CTD) analysis

  • Venous thromboembolism: Associated with NDUFB11 expression changes

• Developmental Disorders:

  • Microphthalmia with linear skin defects (MLS) syndrome: Caused by heterozygous mutations in NDUFB11

  • Also known as MIDAS (microphthalmia, dermal aplasia, and sclerocornea)

  • Associated with additional neurological and cardiac abnormalities

• Stress-Related Conditions:

  • Chronic stress: Altered NDUFB11 expression in stress conditions

  • Pain: Identified as associated through CTD analysis

• Prognostic Significance:

  • Lower NDUFB11 expression associates with poorer prognosis in vascular diseases

  • Potential biomarker for disease progression and therapeutic responses

These disease associations span multiple physiological systems, reflecting NDUFB11's fundamental role in mitochondrial function and cellular energy metabolism across various tissues .

How do mutations in NDUFB11 relate to developmental disorders?

Mutations in NDUFB11 have been specifically linked to developmental disorders through several mechanisms:

• MLS Syndrome Characteristics:

  • Microphthalmia with linear skin defects (MLS) syndrome is an X-linked male-lethal disorder

  • Also known as MIDAS (microphthalmia, dermal aplasia, and sclerocornea)

  • Features include eye abnormalities, skin defects, and neurological and cardiac manifestations

• Specific Mutations:

  • Truncating mutations like c.262C>T (p.Arg88*) and c.402delG (p.Arg134Serfs*3) in NDUFB11 have been identified in MLS syndrome patients

  • These mutations lead to loss of functional NDUFB11 protein

• X-linked Inheritance Pattern:

  • NDUFB11 is located on the X chromosome

  • Mutations are lethal in males (hemizygous)

  • Female carriers show extremely skewed X-chromosome inactivation patterns, indicating strong selection against cells expressing mutant NDUFB11

• Developmental Impact:

  • NDUFB11 mutations disrupt mitochondrial function during critical developmental periods

  • Essential for proper energy metabolism in developing tissues

  • Part of a group of mitochondrial diseases with neurodevelopmental and skin defects as major clinical features

This connection between NDUFB11 mutations and developmental disorders reveals an unexpected role of complex I dysfunction in developmental phenotypes, further underscoring the existence of a group of mitochondrial diseases associated with neurocutaneous manifestations .

What regulatory mechanisms control NDUFB11 expression?

Several regulatory mechanisms have been identified that may control NDUFB11 expression:

• Transcriptional Regulation:

  • As part of mitochondrial biogenesis pathways, NDUFB11 expression is likely coordinated with other mitochondrial genes

  • May be regulated by transcription factors involved in mitochondrial function and energy metabolism

• Post-transcriptional Regulation:

  • miRNAs may play a role in regulating NDUFB11 expression

  • TargetScan has been used to screen for miRNAs regulating NDUFB11 as a central DEG

• X-chromosome Inactivation:

  • As an X-linked gene, NDUFB11 expression in females is subject to X-chromosome inactivation

  • Extremely skewed X-inactivation patterns are observed in females with heterozygous NDUFB11 mutations, suggesting strong selection against cells expressing mutant alleles

• Disease-specific Regulation:

  • Downregulation in atherosclerosis and venous thrombosis suggests disease-specific regulatory mechanisms

  • Altered expression in chronic stress conditions indicates stress-responsive regulatory pathways

Understanding these regulatory mechanisms could provide insights into how NDUFB11 expression is dysregulated in disease states and potentially offer targets for therapeutic intervention.

What are the most promising therapeutic applications targeting NDUFB11?

Several promising therapeutic applications targeting NDUFB11 warrant further investigation:

• Vascular Disease Applications:

  • Restoration of NDUFB11 expression in atherosclerotic vessels to improve mitochondrial function

  • Targeting upstream regulators of NDUFB11 expression in vascular tissues

  • Combined approaches addressing both NDUFB11 and NDUFS3 dysfunction in atherosclerosis

  • Prevention of venous thrombosis by maintaining normal NDUFB11 function

• Developmental Disorder Approaches:

  • Gene therapy strategies for NDUFB11-deficient conditions like MLS syndrome

  • Early intervention targeting developmental pathways affected by NDUFB11 mutations

  • Management approaches for female carriers with skewed X-inactivation patterns

• Chronic Stress Interventions:

  • Maintaining NDUFB11 function during chronic stress to prevent vascular complications

  • Mitochondrial-targeted interventions to mitigate effects of stress on NDUFB11 expression

• Biomarker Applications:

  • NDUFB11 expression as a prognostic biomarker in vascular diseases

  • Monitoring therapy response through changes in NDUFB11 expression

  • Stratification of patients for targeted interventions

These therapeutic directions highlight the potential clinical significance of NDUFB11 research and suggest avenues for translating basic science findings into clinical applications.

What key questions remain unresolved in NDUFB11 research?

Despite significant advances, several critical questions about NDUFB11 function remain unresolved:

• Precise Molecular Function:

  • What is the exact molecular mechanism by which NDUFB11 contributes to complex I assembly?

  • How does NDUFB11 interact with other complex I subunits during assembly?

  • Does NDUFB11 have functions independent of its role in complex I?

• Disease Mechanisms:

  • What is the mechanistic link between NDUFB11 downregulation and atherosclerosis development?

  • How does chronic stress influence NDUFB11 expression and function?

  • What downstream pathways mediate the developmental defects observed in MLS syndrome?

• Tissue-Specific Effects:

  • Why do NDUFB11 mutations predominantly affect specific tissues (eye, skin, heart) in MLS syndrome despite ubiquitous expression?

  • Are there tissue-specific regulatory mechanisms controlling NDUFB11 expression?

• Therapeutic Potential:

  • Can modulation of NDUFB11 expression or function offer therapeutic benefits in conditions like atherosclerosis?

  • What approaches might bypass NDUFB11 deficiency in mitochondrial diseases?

  • Are there specific vulnerabilities in cells with altered NDUFB11 function that could be therapeutically exploited?

Addressing these questions will require integrated approaches combining structural biology, biochemistry, cellular and molecular biology, genetics, and systems biology.

How might technological advances impact future NDUFB11 research?

Emerging technologies are poised to significantly advance NDUFB11 research:

• Advanced Structural Biology:

  • High-resolution structural analysis of NDUFB11 within intact complex I

  • Visualization of assembly intermediates containing NDUFB11

  • Structural changes under different functional states

• Single-Cell Technologies:

  • Single-cell RNA-seq to resolve cell type-specific NDUFB11 expression patterns in disease tissues

  • Spatial transcriptomics to map NDUFB11 expression within tissue architecture

  • Single-cell metabolomics to connect NDUFB11 function to cellular metabolism

• Advanced Genetic Engineering:

  • Precise gene editing for correction of NDUFB11 mutations

  • Inducible, tissue-specific CRISPR systems for in vivo studies

  • CRISPR screening for genetic interactors of NDUFB11

• Disease Modeling:

  • Patient-derived organoids to study tissue-specific NDUFB11 function

  • Vascular-on-a-chip models for atherosclerosis studies

  • Advanced animal models of NDUFB11 dysfunction

• Multi-omics Integration:

  • Comprehensive integration of genomic, transcriptomic, proteomic, and metabolomic data

  • Systems biology approaches to place NDUFB11 in broader cellular networks

  • Computational modeling of complex I assembly and function

These technological advances will enable deeper insights into NDUFB11 function, its role in disease processes, and its potential as a therapeutic target, particularly in atherosclerosis, venous thrombosis, and developmental disorders.