ZFAND3 Human

What is ZFAND3 and what are its primary functions in human cells?

ZFAND3 is an AN1/A20 zinc finger domain-containing protein that functions as a transcriptional regulator in human cells. It acts within a nuclear protein complex to activate gene transcription and regulates promoters of various genes . Research has demonstrated that ZFAND3 requires nuclear localization and intact zinc-finger domains to perform its biological functions . This protein has been particularly implicated in cellular invasion mechanisms, especially in the context of aggressive cancers like glioblastoma, where it regulates invasion-related genes such as COL6A2, FN1, and NRCAM .

How is ZFAND3 structurally organized and what domains are critical for its function?

ZFAND3 contains AN1/A20 zinc finger domains that are essential for its transcriptional regulatory activity. The integrity of these zinc-finger domains is crucial for ZFAND3 function, as demonstrated in experimental models where alterations to these domains impaired the protein's activity . Additionally, nuclear localization is required for ZFAND3 functionality, indicating that its primary mechanistic actions occur within the nucleus rather than in the cytoplasm . The zinc finger domains likely facilitate DNA binding, particularly to GC-rich regions in the promoters of target genes, enabling ZFAND3 to modulate their transcription .

In which human tissues is ZFAND3 typically expressed?

According to protein atlas data, ZFAND3 expression patterns vary across human tissues, with notable presence in brain tissues . This tissue-specific expression pattern may explain its involvement in processes like glioblastoma progression. The Human Protein Atlas contains comprehensive information on ZFAND3 expression across 44 normal human tissue types at both mRNA and protein levels, with data derived from antibody-based profiling using immunohistochemistry . While the search results don't provide the complete tissue expression profile, researchers can access this information through resources like The Human Protein Atlas to understand the physiological distribution of ZFAND3.

What are the most effective techniques for studying ZFAND3 function in human cell models?

Effective techniques for studying ZFAND3 function include:

• Genome-wide interference screens: These have been successfully used to identify ZFAND3 as an invasion-essential gene in glioblastoma .

• Patient-derived cellular models: These provide physiologically relevant contexts for studying ZFAND3's role in disease processes like cancer invasion .

• Gene knockdown techniques: Loss-of-function studies using knockdown approaches have demonstrated that loss of ZFAND3 hampers invasive capacity in GBM cells .

• Overexpression studies: Complementary to knockdown approaches, ZFAND3 overexpression in non-invasive cells has been shown to increase their motility .

• Nuclear localization assays: Given the importance of nuclear localization for ZFAND3 function, techniques that assess subcellular distribution are valuable .

• Protein complex analysis: Mass spectrometry approaches (as indicated by the Q Exactive Plus instrument mentioned in result 5) can help identify ZFAND3's interaction partners within nuclear protein complexes .

What methods are optimal for analyzing ZFAND3 binding to target gene promoters?

To analyze ZFAND3 binding to target gene promoters, researchers should consider:

• Chromatin Immunoprecipitation (ChIP): This technique can identify direct binding of ZFAND3 to promoter regions of putative target genes such as COL6A2, FN1, and NRCAM .

• Promoter analysis of GC-rich regions: Since ZFAND3 has been shown to bind to GC-rich regions, computational analysis to identify such motifs in potential target genes is valuable .

• Reporter gene assays: These can be used to functionally validate the effect of ZFAND3 binding on transcriptional activation of target promoters.

• DNA binding assays: Electrophoretic mobility shift assays (EMSA) or microscale thermophoresis could be used to study the direct interaction between purified ZFAND3 and DNA sequences from target promoters.

• Genome-wide binding profiling: ChIP-seq approaches would provide a comprehensive view of ZFAND3 binding sites across the genome and help identify consensus binding motifs.

How can researchers effectively manipulate ZFAND3 expression in experimental settings?

Effective manipulation of ZFAND3 expression can be achieved through:

• RNA interference (RNAi): siRNA or shRNA approaches have been successfully used to knockdown ZFAND3 in cellular models .

• CRISPR-Cas9 gene editing: For complete knockout or targeted mutations within ZFAND3 domains.

• Overexpression vectors: Plasmid-based overexpression of wild-type or mutant ZFAND3 can be used to study gain-of-function effects .

• Domain-specific mutations: Creating constructs with mutations in zinc-finger domains to study structure-function relationships .

• Inducible expression systems: For temporal control of ZFAND3 expression, which may be particularly valuable when studying dynamic processes like cell invasion.

• Chemical modulation: Certain chemicals have been shown to affect ZFAND3 expression, including 2,3,7,8-tetrachlorodibenzodioxine (increases expression) and 4,4'-diaminodiphenylmethane (decreases expression) .

What is the evidence for ZFAND3's role in glioblastoma progression?

The evidence for ZFAND3's role in glioblastoma (GBM) progression is substantial:

• Genome-wide screening: ZFAND3 was identified through genome-wide interference screening as a crucial driver of GBM invasion .

• Loss-of-function studies: Loss of ZFAND3 has been demonstrated to significantly hamper the invasive capacity of GBM in patient-derived cellular models .

• Gain-of-function studies: Complementarily, ZFAND3 overexpression increases motility in cells that were initially not invasive, supporting its role as a driver of invasion rather than just a marker .

• Mechanistic insights: ZFAND3 regulates the promoters of invasion-related genes such as COL6A2 (collagen type VI alpha 2), FN1 (fibronectin 1), and NRCAM (neuronal cell adhesion molecule), all of which are implicated in cancer cell invasion and migration .

• Nuclear activity: The requirement for nuclear localization and intact zinc-finger domains suggests that ZFAND3 directly regulates transcriptional programs that drive invasive behavior .

How does ZFAND3 regulate genes involved in cancer cell invasion?

ZFAND3 regulates genes involved in cancer cell invasion through several mechanisms:

• Direct promoter binding: ZFAND3 has been shown to bind directly to GC-rich regions in the promoters of invasion-related genes .

• Transcriptional activation: It functions within a nuclear protein complex to activate gene transcription of target genes .

• Regulation of extracellular matrix components: ZFAND3 controls the expression of COL6A2 (collagen) and FN1 (fibronectin), which are key components of the extracellular matrix that facilitate cancer cell migration and invasion .

• Cell adhesion modulation: By regulating NRCAM (neuronal cell adhesion molecule), ZFAND3 may influence cell-cell and cell-matrix interactions that are crucial for invasive behavior .

• Transcriptional complex formation: ZFAND3 likely operates as part of a larger nuclear protein complex, suggesting cooperative interactions with other transcription factors or chromatin modifiers .

What therapeutic implications arise from ZFAND3's role in cancer progression?

The identification of ZFAND3 as a driver of cancer invasion suggests several therapeutic implications:

• Novel therapeutic target: ZFAND3 represents a potential target for developing therapies aimed at reducing cancer invasion and metastasis, particularly in GBM .

• Domain-specific inhibitors: Given the importance of zinc-finger domains for ZFAND3 function, small molecule inhibitors targeting these domains could be developed .

• Nuclear localization disruption: Since nuclear localization is required for ZFAND3 activity, strategies to alter its subcellular distribution might be therapeutically valuable .

• Combination therapies: ZFAND3 inhibition could potentially sensitize tumors to conventional treatments by reducing invasive escape from the tumor core .

• Biomarker potential: ZFAND3 expression or activity could serve as a biomarker for tumor invasiveness and potentially guide treatment decisions .

• Downstream target engagement: Alternatively, targeting the downstream effectors regulated by ZFAND3 (such as COL6A2, FN1, and NRCAM) might provide multiple intervention points .

How do epigenetic modifications affect ZFAND3 expression and function?

Epigenetic regulation of ZFAND3 appears significant based on available data:

• DNA methylation: Several chemicals have been shown to affect ZFAND3 methylation. For example, aflatoxin B1 decreases methylation of the ZFAND3 gene, while arsenic compounds and atrazine affect its methylation patterns .

• Methodological approaches: Researchers investigating epigenetic regulation of ZFAND3 should consider:

  • Bisulfite sequencing to assess CpG methylation in the ZFAND3 promoter

  • Chromatin immunoprecipitation (ChIP) to identify histone modifications at the ZFAND3 locus

  • Assessing the effects of DNA methyltransferase inhibitors or histone deacetylase inhibitors on ZFAND3 expression

• Chemical influences: Various chemicals have been documented to influence ZFAND3 expression or methylation, suggesting environmental or pharmacological factors may modulate its activity through epigenetic mechanisms .

What protein complexes does ZFAND3 form in the nucleus and how do they contribute to transcriptional regulation?

Understanding ZFAND3's nuclear protein interactions is critical:

• Current knowledge: Research indicates that ZFAND3 acts within a nuclear protein complex to activate gene transcription , though the specific components of this complex aren't fully detailed in the provided search results.

• Research approaches: To characterize these complexes, researchers should consider:

  • Immunoprecipitation followed by mass spectrometry to identify interaction partners

  • Proximity labeling techniques like BioID or APEX to capture transient interactions

  • Super-resolution microscopy to visualize co-localization with other nuclear factors

  • Functional genomics screens to identify synthetic interactions

• Mechanistic implications: Understanding the composition of ZFAND3-containing complexes would provide insights into how it recruits transcriptional machinery to target genes and potentially reveal additional therapeutic targets.

How does ZFAND3 expression vary across different cancer types and what are the clinical correlations?

While the search results focus primarily on glioblastoma, this question addresses broader cancer implications:

• Current understanding: ZFAND3 has been particularly studied in glioblastoma, where it drives invasion , but its roles in other cancer types warrant investigation.

• Research approaches:

  • Analysis of cancer genomics databases (TCGA, ICGC) for ZFAND3 expression across cancer types

  • Correlation of ZFAND3 expression with clinical outcomes including patient survival

  • Assessment of ZFAND3 expression in relation to cancer staging and grading

  • Investigation of ZFAND3 status in metastatic vs. primary tumors

• Methodological considerations: Researchers should employ both transcriptomic analyses and protein-level assessments, as post-transcriptional regulation may affect ZFAND3 function independently of mRNA levels.

What signaling pathways modulate ZFAND3 expression or activity?

Understanding the regulation of ZFAND3 itself is important:

• Chemical modulators: Several chemicals have been shown to affect ZFAND3 expression, including:

  • 2,3,7,8-tetrachlorodibenzodioxine (increases expression)

  • 4,4'-diaminodiphenylmethane (decreases expression)

  • 4-hydroxyphenyl retinamide (decreases expression)

  • All-trans-retinoic acid (increases expression)

  • Amitrole (decreases expression)

• Research approaches: To further understand the signaling contexts:

  • Pathway inhibition studies using small molecule inhibitors

  • Analysis of transcription factor binding sites in the ZFAND3 promoter

  • Reporter assays to assess promoter activity under different signaling conditions

  • Phosphoproteomic analysis to identify post-translational modifications on ZFAND3

• Potential applications: Identifying pathways that regulate ZFAND3 could reveal indirect strategies to modulate its activity in disease contexts.

How does the structure of ZFAND3's zinc finger domains determine its DNA binding specificity?

Understanding the structure-function relationship is critical:

• Current knowledge: ZFAND3 contains AN1/A20 zinc finger domains that are critical for its function and binds to GC-rich regions in target gene promoters .

• Research approaches:

  • Structural biology techniques (X-ray crystallography, cryo-EM) to determine the three-dimensional structure of ZFAND3 bound to DNA

  • Mutational analysis of specific residues within zinc finger domains

  • DNA binding assays with various sequence motifs to establish binding preferences

  • Computational modeling of ZFAND3-DNA interactions

• Technical considerations: Purification of functional ZFAND3 protein for in vitro studies may require careful optimization of expression systems and buffer conditions to maintain zinc coordination.

What is the evolutionary conservation of ZFAND3 and how does this inform our understanding of its functional domains?

Evolutionary insights can provide functional clues:

• Cross-species comparisons: The search results indicate that ZFAND3 has been studied in multiple species including humans, mice, and rats , suggesting evolutionary conservation.

• Research approaches:

  • Phylogenetic analysis of ZFAND3 across species

  • Identification of highly conserved residues or domains

  • Functional complementation studies across species

  • Analysis of selection pressure on different protein domains

• Applications: Highly conserved regions are likely to be functionally critical and may represent the most promising targets for therapeutic intervention or further mechanistic studies.

How might single-cell technologies advance our understanding of ZFAND3's role in tumor heterogeneity?

Single-cell approaches offer new perspectives:

• Current technologies: The Human Protein Atlas includes single-cell resource data that presents RNA expression profiles based on single-cell and deconvolution of bulk transcriptomics .

• Research opportunities:

  • Single-cell RNA-seq to analyze ZFAND3 expression patterns within heterogeneous tumors

  • Spatial transcriptomics to examine ZFAND3 expression at invasive fronts versus tumor cores

  • Single-cell ATAC-seq to assess chromatin accessibility at ZFAND3-regulated loci

  • Combined single-cell approaches to correlate ZFAND3 expression with cellular phenotypes

• Potential insights: These approaches could reveal whether ZFAND3 defines specific cellular subpopulations with enhanced invasive potential within tumors.

What are the non-cancer functions of ZFAND3 in normal human physiology?

Understanding physiological roles provides context:

• Current knowledge: While the search results focus on ZFAND3's role in cancer, understanding its normal functions is important for predicting potential side effects of therapeutic targeting.

• Research approaches:

  • Analysis of ZFAND3 expression during development

  • Conditional knockout models in specific tissues

  • Identification of ZFAND3-regulated genes in normal cells versus cancer cells

  • Assessment of ZFAND3's role in tissue homeostasis and response to stress

• Methodological considerations: Careful selection of appropriate control tissues and comprehensive phenotyping are essential when investigating normal functions.

How do post-translational modifications affect ZFAND3 function?

Post-translational regulation may be significant:

• Research opportunities:

  • Mass spectrometry to identify phosphorylation, ubiquitination, or other modifications

  • Site-directed mutagenesis of potential modification sites

  • Use of inhibitors targeting specific modifying enzymes

  • Dynamic analysis of modifications during cellular processes like invasion

• Technical approaches: Proximity labeling methods combined with mass spectrometry could identify enzymes responsible for ZFAND3 modifications in their native cellular context.

• Functional implications: Modifications could affect ZFAND3's stability, nuclear localization, DNA binding affinity, or protein-protein interactions, providing additional regulatory layers.

What biomarker potential does ZFAND3 expression or activity hold for cancer prognosis?

Translating mechanistic insights to clinical applications:

• Current evidence: ZFAND3's role in driving GBM invasion suggests potential as a prognostic biomarker , though specific clinical correlations aren't detailed in the search results.

• Research approaches:

  • Retrospective analysis of ZFAND3 expression in tumor samples with known outcomes

  • Development of immunohistochemical protocols for ZFAND3 detection in clinical specimens

  • Assessment of ZFAND3 as part of multi-marker panels for invasive potential

  • Correlation with imaging features associated with tumor invasiveness

• Clinical considerations: Standardization of detection methods and establishment of relevant cutoff values would be necessary for clinical application.

What strategies might be effective for therapeutic targeting of ZFAND3 or its downstream pathways?

Therapeutic development considerations:

• Potential approaches:

  • Small molecule inhibitors targeting zinc finger domains

  • Degrader technologies (PROTACs) to eliminate ZFAND3 protein

  • Antisense oligonucleotides or RNAi-based approaches to reduce expression

  • Disruption of protein-protein interactions within the nuclear complex

  • Targeting downstream effectors (COL6A2, FN1, NRCAM)

• Challenges and considerations:

  • Specificity among zinc finger proteins

  • Delivery to the brain across the blood-brain barrier for GBM applications

  • Potential compensatory mechanisms

  • Safety considerations based on normal tissue expression

• Combination strategies: Given the complexity of invasion mechanisms, combining ZFAND3-targeted approaches with standard therapies may offer the most promise.

How might chemical modulators of ZFAND3 be leveraged for therapeutic development?

Chemical biology approaches:

• Known chemical modulators: Several chemicals affect ZFAND3 expression or methylation , including:

  • Compounds that increase expression: 2,3,7,8-tetrachlorodibenzodioxine, all-trans-retinoic acid

  • Compounds that decrease expression: 4,4'-diaminodiphenylmethane, 4-hydroxyphenyl retinamide, amitrole

  • Compounds affecting methylation: aflatoxin B1, arsenic compounds, atrazine

• Research opportunities:

  • High-throughput screening for novel modulators with improved properties

  • Structure-activity relationship studies based on known modulators

  • Medicinal chemistry optimization of lead compounds

  • In vivo testing in relevant disease models

• Translational pathway: From the identified modulators, structural insights could guide development of more specific and potent compounds for potential clinical development.