NAIF1 Human

What is NAIF1 and what is its genomic classification?

NAIF1 (Nuclear Apoptosis-Inducing Factor 1) is classified as a "Neogene" that originated through molecular domestication of DNA transposons (second class of transposable elements). These neogenes can play significant roles in human genetic instability, diseases, and cancer development. NAIF1 specifically functions as an apoptosis-inducing factor in various human cancers .

Where is NAIF1 located genetically and within the cell?

The human gene encoding NAIF1 is located on chromosome 9q34.11. At the cellular level, NAIF1 is primarily localized in the nuclei of cells, as confirmed through fluorescent confocal microscopy using NAIF1-GFP fusion protein visualization techniques .

What is the structural composition of the NAIF1 protein?

NAIF1 encodes a protein with a Myb-like domain at its N-terminal region. This structural feature is significant as Myb domains are typically involved in DNA binding and transcriptional regulation, which may be related to NAIF1's role in cell cycle control and apoptosis induction .

How does NAIF1 differ from other apoptosis-inducing factors?

Unlike other apoptosis-inducing factors that may work through extrinsic or intrinsic apoptotic pathways, NAIF1 appears to function through cell cycle regulation, specifically by inducing cell cycle arrest at the G1/S phase. This mechanism involves the alteration of key cell cycle regulatory proteins including cyclinD1, cdc2, and p21, distinguishing NAIF1's apoptotic mechanism from other factors .

What techniques are effective for detecting NAIF1 expression in tissue samples?

Western blot analysis is the primary method used for detecting NAIF1 protein expression in tissues and cell lines. For visualization of cellular localization, fluorescent confocal microscopy using NAIF1-GFP fusion protein has proven effective. Additionally, RNA expression levels can be assessed using quantitative PCR methods to complement protein expression data .

How does NAIF1 expression vary across normal versus cancerous tissues?

NAIF1 expression shows significant differential patterns between normal and cancerous tissues. Research indicates that NAIF1 is significantly expressed in normal gastric tissue, while its expression is down-regulated or completely lost in gastric cancer tissues (P<0.001). Additionally, NAIF1 expression is higher in well-differentiated gastric cancer compared to moderately or poorly differentiated cases (P=0.004) .

What is the expression profile of NAIF1 across different cancer cell lines?

Studies have demonstrated that NAIF1 is minimally expressed across tested gastric cancer cell lines including MKN45, BGC823, AGS, and SGC7901. In leukemia research, NAIF1 protein expression was found to vary across different leukemia cell lines (HL60, NB4, KG1, KG1a, ML2, THP1, and U937), with generally lower expression compared to healthy blood tissue controls .

What reference genes should be used when normalizing NAIF1 expression data?

While the search results don't specifically mention reference genes for NAIF1 normalization, standard housekeeping genes used in cancer research such as GAPDH, β-actin, or 18S rRNA would be appropriate choices. The selection should be validated for the specific tissue or cell type under investigation to ensure stable expression across experimental conditions.

What is the proposed tumor suppressor mechanism of NAIF1?

NAIF1 appears to function as a tumor suppressor through multiple mechanisms:

• Induction of cell cycle arrest at G1/S phase by altering the expression of cell cycle proteins including cyclinD1, cdc2, and p21

• Regulation of cellular apoptosis pathways

• Alteration of the proteomic profile affecting cellular programs involved in cell cycle, apoptosis, and signal transduction regulation

How does NAIF1 overexpression affect the proteomic profile of cancer cells?

Comparative proteomic analysis of gastric cancer cell lines with and without NAIF1 overexpression identified several differentially expressed proteins:

These proteins are involved in various cellular processes including cell cycle regulation, apoptosis, and signal transduction, suggesting multiple pathways through which NAIF1 may exert its tumor suppressive effects .

Is NAIF1 expression level correlated with cancer differentiation status?

Yes, studies have found that NAIF1 expression correlates with cancer differentiation status. In gastric cancer, NAIF1 expression was significantly higher in well-differentiated tumors compared to moderately or poorly differentiated cases (P=0.004). This suggests that loss of NAIF1 expression may be associated with cancer progression and dedifferentiation .

What is the relationship between NAIF1 and leukemia progression?

NAIF1 protein expression shows variable degrees of expression across different leukemia cell lines, with generally reduced expression compared to healthy blood tissue. This suggests that NAIF1 may play a role in inhibiting proliferation, migration, and invasion of leukemia cells by inducing apoptosis, though further research is needed to fully characterize this relationship .

What cell models are appropriate for studying NAIF1 function?

Based on existing research, several cell models have proven effective for studying NAIF1:

• Gastric cancer cell lines (MKN45, BGC823, AGS, SGC7901)

• Leukemia cell lines (HL60, NB4, KG1, KG1a, ML2, THP1, U937)

• Cervical cancer cells (HeLa)

These cell lines provide useful models for investigating NAIF1's role in different cancer types and cellular contexts .

What transfection methods are effective for NAIF1 overexpression studies?

For NAIF1 overexpression studies, X-tremeGENE HP DNA transfection reagent has been successfully employed. This method can be used with expression plasmids such as pEGFP-N1-NAIF1 (expressing NAIF1-GFP fusion protein) for both functional studies and visualization of cellular localization. Standard transfection protocols using 37°C incubation in a humidified 5% CO₂ atmosphere are appropriate for these experiments .

How can cell cycle effects of NAIF1 be accurately measured?

To measure NAIF1's effects on cell cycle:

• Transfect cells with NAIF1 expression vectors and appropriate controls

• Harvest cells at specific time points (24h and 48h post-transfection)

• Wash cells with PBS and fix with 4% paraformaldehyde

• Stain with propidium iodide solution (0.05 mg/ml PI, 0.2 mg/ml RNase, and 0.1% Triton X-100)

• Perform flow cytometry analysis for both GFP (transfection marker) and PI (DNA content)

• Analyze cell cycle distribution using appropriate flow cytometry software

What approaches can be used to study the relationship between NAIF1 and apoptosis?

To investigate NAIF1's role in apoptosis:

• Overexpress NAIF1 in appropriate cell lines

• Assess apoptotic markers using techniques such as:

  • Annexin V/PI double staining followed by flow cytometry

  • TUNEL assay for DNA fragmentation

  • Western blot analysis of apoptotic proteins (caspases, PARP cleavage)

  • Microscopic examination of nuclear morphology

• Compare results with appropriate control groups

• Validate findings using multiple cell lines and complementary techniques

How do the Myb-like domains in NAIF1 contribute to its molecular function?

While the search results mention that NAIF1 contains a Myb-like domain at its N-terminal region, detailed functional analysis of this domain is not provided. Myb domains typically function as DNA-binding motifs involved in transcriptional regulation. Advanced research should investigate whether NAIF1's Myb-like domain directly interacts with specific DNA sequences to regulate gene expression, potentially controlling cell cycle or apoptosis-related genes. Methodological approaches could include:

• Chromatin immunoprecipitation (ChIP) to identify DNA binding sites

• Mutagenesis of key residues within the Myb domain to assess functional impact

• Structural studies using X-ray crystallography or NMR to determine binding specificity

What are the molecular interactions between NAIF1 and the cell cycle machinery?

NAIF1 has been shown to induce cell cycle arrest at G1/S phase by altering expression of cyclinD1, cdc2, and p21. Advanced research should explore:

• Direct vs. indirect interactions between NAIF1 and these cell cycle regulators

• Phosphorylation status of key cell cycle proteins in response to NAIF1 expression

• Temporal dynamics of these interactions using synchronization techniques

• Upstream regulators and downstream effectors in the NAIF1-mediated cell cycle control pathway

What is the evolutionary history of NAIF1 as a domesticated transposon?

NAIF1 is classified as a "Neogene" resulting from molecular domestication of DNA transposons. Advanced evolutionary research should investigate:

• The specific transposon family from which NAIF1 originated

• When this domestication event occurred in evolutionary history

• Whether NAIF1 homologs exist in other species and their functional conservation

• How the original transposon sequence was modified to generate NAIF1's current function

• Comparative analysis with other domesticated transposons involved in apoptosis regulation

What epigenetic mechanisms regulate NAIF1 expression in different cancer types?

Given that NAIF1 shows differential expression between normal and cancerous tissues, investigation of epigenetic regulation is warranted:

• DNA methylation analysis of the NAIF1 promoter in different cancer types

• Histone modification patterns in the NAIF1 gene region

• Effect of epigenetic modifying drugs (HDAC inhibitors, DNA methyltransferase inhibitors) on NAIF1 expression

• Role of microRNAs in post-transcriptional regulation of NAIF1

• Long non-coding RNAs that might interact with NAIF1 genomic regions

How might NAIF1 expression serve as a prognostic biomarker?

Based on the correlation between NAIF1 expression and cancer differentiation status, NAIF1 could potentially serve as a prognostic biomarker. Research approaches should include:

• Large-scale clinical studies correlating NAIF1 expression with patient outcomes

• Multivariate analysis to determine if NAIF1 provides independent prognostic value

• Development of standardized immunohistochemistry protocols for clinical use

• Evaluation of NAIF1 expression in combination with other established biomarkers

What are the barriers to targeting NAIF1 pathways therapeutically?

While NAIF1 shows potential as a tumor suppressor, several challenges must be addressed for therapeutic application:

• Developing methods to restore or increase NAIF1 expression in cancer cells

• Identifying the critical downstream effectors that could be targeted pharmaceutically

• Understanding potential off-target effects of NAIF1 pathway modulation

• Determining cancer types most likely to respond to NAIF1-based therapies

• Establishing appropriate delivery methods for NAIF1-targeted therapeutics

How does NAIF1 interact with established cancer treatment modalities?

Research should investigate potential synergistic effects between NAIF1 expression and:

• Traditional chemotherapeutic agents

• Radiation therapy

• Targeted molecular therapies

• Immunotherapeutic approaches

Experimental designs should include combination treatment protocols in both in vitro and in vivo models, with assessment of cell cycle effects, apoptosis induction, and long-term survival outcomes .