TNRC6C Antibody

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Description

Introduction to TNRC6C Antibody

TNRC6C antibodies are polyclonal or monoclonal antibodies developed to target the TNRC6C protein, which facilitates miRNA-mediated gene silencing by bridging Argonaute proteins and deadenylase complexes . These antibodies are widely used in immunohistochemistry (IHC), Western blotting, and immunofluorescence to study TNRC6C expression patterns in cancer biology, particularly in thyroid malignancies.

Applications in Research

TNRC6C antibodies have enabled critical discoveries through the following methodologies:

  • Immunohistochemistry (IHC): Detects TNRC6C protein localization and expression levels in PTC tissues versus adjacent normal tissues .

  • Western Blotting: Validates TNRC6C knockdown or overexpression in cell lines like BCPAP and TPC1 .

  • Functional Assays: Correlates TNRC6C expression with cellular behaviors (proliferation, apoptosis, migration) in vitro .

Table 1: Clinicopathological Associations of TNRC6C in PTC (n=502)

FeatureLow TNRC6C Expression (%)High TNRC6C Expression (%)P Value
Tumor Size (T3/T4)57.337.90.002
Lymph Node Metastasis67.152.50.014
Advanced Stage (III/IV)51.234.30.005
Aggressive Histology13.01.6<0.001
Data derived from TCGA analysis .

3.1. Tumor-Suppressive Role in PTC

  • TNRC6C is downregulated in PTC tissues compared to normal thyroid tissues (IHC validation) .

  • Overexpression of TNRC6C inhibits PTC cell proliferation by 30–40%, reduces migration/invasion by 50–60%, and increases apoptosis by 6-fold .

  • Low TNRC6C correlates with larger tumors, lymph node metastasis, and advanced clinical stages (Table 1) .

3.2. Regulatory Targets of TNRC6C

RNA sequencing and TCGA data identified 12 oncogenic genes (e.g., CTHRC1, MMP14, COL1A1) negatively regulated by TNRC6C. These genes are overexpressed in PTC and linked to aggressive phenotypes .

Table 2: Key TNRC6C Target Genes in PTC

GeneFunction in CancerRegulation by TNRC6C
CTHRC1Promotes metastasisDownregulated
MMP14Enhances extracellular remodelingDownregulated
COL1A1Facilitates tumor stroma formationDownregulated

Clinical Implications

  • Prognostic Marker: Low TNRC6C expression predicts poor outcomes in PTC patients .

  • Therapeutic Target: Restoring TNRC6C expression suppresses tumor growth in vitro, suggesting potential for gene therapy .

  • Antisense RNA Interaction: TNRC6C-AS1, a lncRNA, inversely regulates TNRC6C and could be targeted to modulate its activity .

Future Directions

  • Develop monoclonal TNRC6C antibodies for high-specificity therapeutic applications.

  • Investigate TNRC6C’s role in miRNA-independent pathways, given its nuclear localization in PTC .

  • Explore combinatorial therapies targeting TNRC6C-AS1 to enhance TNRC6C expression .

Product Specs

Buffer
**Preservative:** 0.03% Proclin 300
**Constituents:** 50% Glycerol, 0.01M PBS, pH 7.4
Form
Liquid
Lead Time
Typically, we can ship the products within 1-3 business days after receiving your order. The delivery timeframe may vary depending on the purchase method and location. For specific delivery time estimates, please consult your local distributor.
Synonyms
KIAA1582 antibody; TNR6C_HUMAN antibody; TNRC 6C antibody; Tnrc6c antibody; Trinucleotide repeat containing 6C antibody; Trinucleotide repeat containing gene 6C protein antibody; Trinucleotide repeat-containing gene 6C protein antibody
Target Names
TNRC6C
Uniprot No.

Target Background

Function
TNRC6C (GW182) plays a crucial role in RNA-mediated gene silencing through micro-RNAs (miRNAs). It is essential for miRNA-dependent translational repression of complementary mRNAs by argonaute family proteins. Acting as a scaffold protein, TNRC6C associates with argonaute proteins bound to partially complementary mRNAs, simultaneously recruiting CCR4-NOT and PAN deadenylase complexes.
Gene References Into Functions
  1. These findings demonstrate that despite species-specific differences in the relative strength of PABPC1-binding sites, the interaction between GW182 proteins and PABPC1 is crucial for miRNA-mediated silencing in animal cells. PMID: 21063388
  2. The authors reveal that a conserved motif within the human GW182 paralog TNRC6C interacts with the C-terminal domain of polyadenylate binding protein 1 (PABC) and present the crystal structure of this complex. PMID: 20098421
  3. Through deletion and mutagenesis, the study identified the C-terminal portion of TNRC6C encompassing the RRM RNA-binding motif as a key effector domain mediating protein synthesis repression by TNRC6C. PMID: 19304925
  4. Our findings indicate that TNRC6C is recruited to miRNA targets through an interaction between its N-terminal domain and an Argonaute protein. PMID: 19383768
Database Links

HGNC: 29318

OMIM: 610741

KEGG: hsa:57690

STRING: 9606.ENSP00000336783

UniGene: Hs.406810

Protein Families
GW182 family

Q&A

What is TNRC6C and why is it important in research?

TNRC6C (Trinucleotide Repeat Containing 6C) functions as an important component within miRNA-induced silencing complex . It has been identified as a potential tumor suppressor in papillary thyroid cancer (PTC), with studies showing that it inhibits cell proliferation, migration, and invasion while promoting apoptosis in PTC cells . The protein's role in post-transcriptional gene regulation makes it a valuable target for cancer research, particularly in understanding gene expression control mechanisms that contribute to tumorigenesis.

What applications are TNRC6C antibodies validated for?

TNRC6C antibodies are validated for multiple experimental applications including ELISA, Western Blot (WB), Immunohistochemistry (IHC), and Immunofluorescence (IF) . Different commercially available antibodies show varying application profiles - for example, Sigma-Aldrich's HPA062051 antibody is specifically validated for immunofluorescence at concentrations of 0.25-2 μg/mL . When selecting an antibody, researchers should verify validation data for their specific application to ensure reliable results.

How should TNRC6C antibodies be stored and handled to maintain efficacy?

TNRC6C antibodies are typically shipped on wet ice and should be stored at -20°C for optimal preservation . Most commercial TNRC6C antibodies are provided in buffered aqueous glycerol solutions to maintain stability . Researchers should avoid repeated freeze-thaw cycles by aliquoting the antibody upon receipt. For short-term use (1-2 weeks), storage at 4°C is acceptable, but long-term storage requires -20°C conditions to prevent degradation and maintain binding efficacy across multiple experiments.

What are the common positive and negative controls for TNRC6C antibody experiments?

When validating TNRC6C antibodies, researchers should use:

Positive Controls:

  • PTC cell lines with known TNRC6C expression such as BCPAP or TPC1 cells

  • Normal thyroid tissue samples where TNRC6C is expressed

  • HEK293 cells transfected with TNRC6C expression vectors

Negative Controls:

  • TNRC6C siRNA-transfected cells showing knockdown (validation by qPCR)

  • Secondary antibody-only controls to assess non-specific binding

  • Isotype controls with irrelevant antibodies of the same class/species

Validation should include multiple controls to ensure specificity before undertaking extensive experimental work.

How can researchers optimize immunohistochemical detection of TNRC6C in thyroid tissue samples?

Optimizing IHC for TNRC6C in thyroid tissues requires attention to several parameters:

Recommended Protocol:

  • Fixation: Use 10% neutral buffered formalin for 24-48 hours

  • Antigen retrieval: Heat-induced epitope retrieval in citrate buffer (pH 6.0) for 20 minutes

  • Blocking: 5% normal goat serum in PBS for 1 hour at room temperature

  • Primary antibody incubation: Anti-TNRC6C at 1-2 μg/mL overnight at 4°C

  • Detection system: HRP-polymer based with DAB chromogen

  • Counterstaining: Hematoxylin for 30 seconds

Critical Considerations:

  • Nuclear localization of TNRC6C requires careful optimization of permeabilization steps

  • Comparison between tumor and adjacent normal tissue on the same slide helps control for technical variations

  • Quantification should include both intensity and percentage of positive cells using standardized scoring systems

What are the best experimental approaches to study TNRC6C's function in miRNA-induced silencing complex?

Investigating TNRC6C's role in miRNA-induced silencing requires multifaceted approaches:

Recommended Methodologies:

  • Co-Immunoprecipitation (Co-IP):

    • Use anti-TNRC6C antibody to pull down the protein complex

    • Analyze interacting partners by mass spectrometry or Western blot

    • Confirm interactions with Argonaute proteins

  • RNA-Immunoprecipitation (RIP):

    • Immunoprecipitate TNRC6C-containing complexes

    • Extract and identify associated miRNAs and mRNAs by sequencing

  • Microscopy techniques:

    • Immunofluorescence co-localization with other miRISC components

    • Live-cell imaging with fluorescently tagged TNRC6C

  • Functional assays:

    • Luciferase reporter assays with miRNA target sequences

    • TNRC6C knockout/knockdown followed by transcriptome analysis

How can researchers accurately distinguish between TNRC6C and its paralogs (TNRC6A/B) in experimental settings?

Distinguishing between TNRC6 family members requires careful antibody selection and validation:

Recommended Approaches:

  • Antibody epitope analysis:

    • Select antibodies targeting unique regions like the immunogen sequence "GGLSVKDPSQSQSRLPQWTHPNSMDNLPSAASPLEQNPSKHGAIPGGLSIGPPGKSSIDDSYGRYDLIQNSESPASPPVAVPHSWSRAKSDSDKISNGSSINWPPEFHPGV"

    • Verify specificity using peptide competition assays

  • Paralog-specific knockdown controls:

    • siRNA targeting each paralog independently

    • Western blot with different antibodies to confirm specificity

  • Recombinant protein standards:

    • Include purified TNRC6A/B/C proteins as controls

    • Assess cross-reactivity quantitatively

TNRC6 Family MemberMolecular WeightKey Distinguishing FeaturesRecommended Validation Approach
TNRC6C175 kDaDistinct C-terminal regionWestern blot with size verification
TNRC6B194 kDaDifferent migration patternParallel detection with Anti-TNRC6B (e.g., AB9913)
TNRC6A183 kDaUnique epitopesSpecific knockdown verification

What is the evidence supporting TNRC6C as a tumor suppressor in thyroid cancer?

Multiple lines of evidence establish TNRC6C's tumor suppressor role:

Experimental Evidence:

  • Expression analysis: TNRC6C is downregulated in PTC compared to normal thyroid tissue

  • Functional studies:

    • Overexpression inhibits proliferation, migration, and invasion in BCPAP and TPC1 cells

    • Knockdown promotes malignant phenotypes in thyroid cancer cell lines

  • Clinical correlations:

    • Lower TNRC6C expression associates with worse clinicopathological features

    • TCGA data analysis confirms downregulation in PTC and correlation with prognosis

  • Mechanistic insights:

    • TNRC6C regulates multiple cancer-associated genes including SCD, CRLF1, APCDD1L, CTHRC1, and others

These findings collectively demonstrate that TNRC6C functions to suppress thyroid cancer progression and may serve as a therapeutic target and prognostic marker for PTC patients.

How does TNRC6C-AS1 differ from TNRC6C, and what are the implications for antibody-based research?

TNRC6C-AS1 is a long non-coding RNA that is distinct from the TNRC6C protein:

Key Differences and Research Implications:

  • Molecular nature:

    • TNRC6C: Protein functioning in miRNA-induced silencing complex

    • TNRC6C-AS1: Long non-coding RNA that acts as a competing endogenous RNA

  • Expression patterns:

    • TNRC6C: Downregulated in thyroid cancer

    • TNRC6C-AS1: Upregulated in thyroid cancer tissues

  • Functional roles:

    • TNRC6C: Tumor suppressor

    • TNRC6C-AS1: Oncogenic, promotes cancer progression

  • Experimental considerations:

    • Antibody-based detection only works for TNRC6C protein, not the AS1 transcript

    • RNA-based methods (qRT-PCR) are needed to study TNRC6C-AS1

    • Researchers must carefully distinguish between these entities in experimental design

  • Mechanistic interaction:

    • TNRC6C-AS1 functions through a TNRC6C-AS1/miR-513c-5p/LPAR5 axis

    • This pathway represents a potential therapeutic target distinct from TNRC6C protein

What methodological approaches can resolve contradictory findings in TNRC6C expression across different cancer studies?

Researchers encountering contradictory findings should implement these methodological approaches:

Reconciliation Strategies:

  • Comprehensive tissue analysis:

    • Compare matched tumor-normal pairs from the same patients

    • Analyze different histological subtypes separately

    • Use laser capture microdissection to isolate specific cell populations

  • Multi-level expression analysis:

    • Assess both mRNA (qRT-PCR) and protein (Western blot, IHC) levels

    • Evaluate subcellular localization (nuclear vs. cytoplasmic fractions)

    • Consider post-transcriptional regulation

  • Technical standardization:

    • Use consistent antibody clones and detection protocols

    • Implement quantitative scoring systems with defined thresholds

    • Include multiple positive and negative controls

  • Integrative data analysis:

    • Compare findings with public datasets (TCGA, GEO)

    • Perform meta-analysis of published studies

    • Correlate with clinical parameters to identify context-dependent effects

What are the common technical issues when using TNRC6C antibodies in Western blotting, and how can they be resolved?

Researchers frequently encounter these challenges with TNRC6C Western blotting:

Common Issues and Solutions:

IssuePossible CausesSolution
High molecular weight band detection difficultiesLarge protein size (175 kDa)Use gradient gels (4-12%); Extend transfer time (overnight at low voltage)
Multiple bands/non-specific bindingCross-reactivity with TNRC6A/BUse validated antibodies with confirmed specificity; Include knockout controls
Weak signalLow expression levelsIncrease protein loading (50-80 μg); Use enhanced chemiluminescence detection systems
Inconsistent results between experimentsProtein degradationAdd additional protease inhibitors; Minimize freeze-thaw cycles of samples
Background issuesSecondary antibody cross-reactivityOptimize blocking (5% BSA often better than milk for phospho-antibodies); Increase washing steps

Optimized Western Blot Protocol for TNRC6C:

  • Use fresh lysates with complete protease inhibitor cocktails

  • Load 60-80 μg protein per lane on 4-8% gradient gels

  • Transfer at 30V overnight at 4°C to PVDF membranes

  • Block with 5% BSA for 2 hours at room temperature

  • Incubate with anti-TNRC6C antibody (1:1000) overnight at 4°C

  • Wash extensively (4 × 10 min) with TBST

  • Use HRP-conjugated secondary antibody (1:5000) for 1 hour

  • Develop using enhanced chemiluminescence reagents

How can researchers optimize TNRC6C immunofluorescence staining to accurately detect subcellular localization?

Precise subcellular localization of TNRC6C requires these optimization steps:

Optimized Immunofluorescence Protocol:

  • Cell preparation:

    • Culture cells on poly-L-lysine coated coverslips

    • Fix with 4% paraformaldehyde (10 minutes at room temperature)

    • Permeabilize with 0.2% Triton X-100 (for nuclear proteins)

  • Antibody incubation:

    • Block with 3% BSA + 10% normal serum for 1 hour

    • Apply primary anti-TNRC6C antibody at 0.25-2 μg/mL overnight at 4°C

    • Use fluorophore-conjugated secondary antibodies (1:500-1:1000)

  • Nuclear co-staining:

    • Include DAPI (1 μg/mL) for nuclear visualization

    • Consider co-staining with other miRISC components

  • Image acquisition:

    • Use confocal microscopy for precise localization

    • Z-stacking to confirm true nuclear vs. cytoplasmic distribution

    • Consistent exposure settings between samples

  • Controls:

    • Include TNRC6C knockdown cells

    • Secondary-only controls

    • Known subcellular markers co-staining

The IHC results showed that TNRC6C is primarily localized in the nucleus, suggesting it may regulate gene expression at the transcriptional level in addition to its known role in miRNA-induced silencing complexes .

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