TROVE2 Human

What is TROVE2 and what are its alternative nomenclature in scientific literature?

TROVE2 (TROVE domain family member 2) is a protein-coding gene in humans that belongs to the RNA-binding protein (RBP) family. The gene is also commonly referred to as RO60, RORNP, and SSA2 in scientific literature . This protein is classified as an RNA binding protein (RBP) that participates in post-transcriptional regulation of gene expression . TROVE2 contains specific structural domains that facilitate RNA recognition and binding, enabling it to perform its cellular functions. As with many proteins with multiple names, researchers should be aware of all alternative nomenclatures when conducting literature searches to ensure comprehensive coverage of existing research. When publishing findings related to this protein, it is recommended to include all common names in keywords and abstracts to improve discoverability of research papers. The official gene ID in the Entrez Gene database is 6738, which should be referenced in genetic studies involving this protein .

How can researchers effectively detect TROVE2 expression in experimental systems?

Detection of TROVE2 expression can be accomplished through multiple complementary techniques. For protein-level detection, western blotting using specific anti-TROVE2 antibodies such as rabbit polyclonal antibodies is a widely adopted approach . For immunocytochemistry and immunofluorescence applications, researchers should select antibodies validated for these specific applications to avoid non-specific binding . At the transcriptional level, researchers can employ RT-PCR or qPCR to quantify TROVE2 mRNA expression, with appropriate normalization to housekeeping genes. For high-throughput analysis, researchers have successfully employed SAGE (Serial Analysis of Gene Expression) and MPSS (Massively Parallel Signature Sequencing) to verify TROVE2 expression levels in normal tissues . When analyzing TROVE2 expression, it is crucial to ensure tag sequences for SAGE or MPSS analyses are specific and unambiguous, as some genes lack unique tags which can compromise accurate quantification . For comprehensive expression profiling, combining multiple detection methods is recommended to overcome the limitations inherent to each individual technique.

What is the normal expression pattern of TROVE2 across human tissues?

TROVE2 expression has been systematically analyzed across multiple normal human tissues through comprehensive transcriptomic approaches. Studies utilizing SAGE and MPSS have established baseline expression profiles that reveal tissue-specific patterns . In liver tissue, TROVE2 shows moderate expression in normal hepatocytes, with LO2 cells (normal liver cell line) exhibiting lower expression levels compared to some hepatocellular carcinoma cell lines . When analyzing TROVE2 expression across neural tissues, research has documented expression in astrocytes, cerebellum, and cortical regions, providing important baseline data for comparative oncology studies . Expression analysis must account for potential technical variations between methodologies, as discrepancies have been observed between microarray and SAGE/MPSS data for some RNA binding proteins . Researchers investigating TROVE2 should consider utilizing multiple expression datasets and technologies to obtain a comprehensive understanding of its tissue distribution. For reliable quantification in experimental systems, appropriate reference tissues or cell lines with validated expression levels should be included as controls.

What are the methodological considerations when selecting antibodies for TROVE2 research?

When selecting antibodies for TROVE2 research, investigators should prioritize reagents validated for specific applications relevant to their experimental design. Commercial antibodies such as rabbit polyclonal anti-TROVE2/SS-A antibodies have been validated for western blotting and immunocytochemistry/immunofluorescence applications in human samples . Researchers should carefully evaluate the immunogen used for antibody production, as antibodies raised against different epitopes may yield varying results in different applications or experimental conditions. For immunoprecipitation studies, it is essential to verify that the antibody can recognize the native conformation of TROVE2 rather than just denatured protein. When conducting immunohistochemistry on tissue sections, appropriate antigen retrieval methods must be optimized, as TROVE2 detection may require specific retrieval conditions depending on tissue fixation protocols. Cross-reactivity testing is particularly important when studying TROVE2 in conjunction with related proteins or in comparative studies across species. Finally, researchers should maintain consistent antibody lots throughout a study to minimize technical variability, as lot-to-lot variations can significantly impact quantitative analyses.

How does anti-TROVE2 antibody serve as a predictor of therapeutic response in rheumatoid arthritis?

Anti-TROVE2 (anti-Ro60) antibody has emerged as a significant predictive biomarker for both the development of anti-drug antibodies (ADAbs) and therapeutic response in rheumatoid arthritis patients treated with adalimumab. Research utilizing immune-related protein microarray technology has identified anti-TROVE2 antibody as having the highest individual discriminating ability for predicting ADAb development, both at baseline (pretreatment) and week 24 of adalimumab therapy . Multivariate logistic regression analysis has convincingly demonstrated that anti-TROVE2 antibody is an independent biomarker associated with ADAb development, with an impressive odds ratio of 70.27 (95% CI 8.17-604.38, p < 0.001) after adjusting for multiple confounding factors including age, sex, disease duration, radiographic stage, baseline DAS28, and the positivity of RF or ACPA . The predictive value extends to therapeutic outcomes, with anti-TROVE2 antibody identified as an independent biomarker for predicting poor EULAR response assessed at week 24 (OR 55.1, 95% CI 1.9-1596.3, p < 0.05) . Mechanistically, evidence suggests that anti-Ro60 (anti-TROVE2) autoantibody directly binds to the fully human monoclonal antibody adalimumab, which could potentially serve as a template for the ensuing development of anti-drug antibodies with high-affinity adaptive response . For clinical implementation, immunofluorescent-based ELISA has been validated as an effective method to discriminate ADAb-positive from ADAb-negative patients, making this approach feasible for routine clinical testing.

What is the optimal methodology for detecting anti-TROVE2 antibodies in patient samples?

The detection of anti-TROVE2 antibodies in patient samples requires careful methodological consideration to ensure accurate and reproducible results. Immunofluorescence-based ELISA has been validated as a particularly effective method for discriminating between patients with and without anti-drug antibodies (ADAbs), showing strong correlation with clinical outcomes . When implementing this methodology, researchers should establish standardized protocols with appropriate positive and negative controls to ensure consistent results across different batches of samples. High-density protein microarray technology, specifically the IMMUNOME Array platform, has demonstrated utility in detecting anti-TROVE2 antibodies with high sensitivity and specificity (77% and 81%, respectively) . This platform offers the advantage of displaying both linear and discontinuous epitopes across thousands of human proteins, enabling comprehensive autoantibody profiling. For quantitative assessment, plasma anti-TROVE2 antibody levels can be measured and have been shown to correlate positively with ADAb titers and negatively with drug levels in adalimumab-treated patients . When interpreting results, researchers should be aware of potential confounding factors such as other autoantibodies that may cross-react or interfere with the assay. For longitudinal studies, consistency in sample collection, processing, and storage conditions is crucial to minimize pre-analytical variables that could affect antibody detection.

What molecular mechanisms underlie the interaction between anti-TROVE2 antibodies and biologic drugs?

The interaction between anti-TROVE2 antibodies and biologic drugs, particularly adalimumab, involves complex molecular mechanisms that require further elucidation. Current evidence suggests that anti-Ro60 (anti-TROVE2) autoantibody directly binds to adalimumab, a fully human monoclonal antibody used in treating rheumatoid arthritis . This initial binding, which may be relatively weak, could potentially serve as a template or scaffold for the subsequent development of high-affinity anti-drug antibodies (ADAbs) through an adaptive immune response. The correlation coefficient between anti-Ro60 titers and ADAb titers (approximately 0.79) supports this mechanistic relationship, though the incomplete correlation suggests other factors may contribute to ADAb development . Mechanistically, this interaction might involve structural epitopes on adalimumab that share similarities with the TROVE2 protein, leading to cross-reactivity of pre-existing anti-TROVE2 antibodies. For experimental investigation of these interactions, researchers should consider employing techniques such as surface plasmon resonance to determine binding kinetics, or hydrogen-deuterium exchange mass spectrometry to map the specific binding interfaces. Co-immunoprecipitation followed by mass spectrometry could also help identify other proteins that might be involved in forming complexes with anti-TROVE2 antibodies and adalimumab. Understanding the three-dimensional structure of these complexes using techniques like cryo-electron microscopy could provide valuable insights into the molecular basis of these interactions.

How does TROVE2 contribute to hepatocellular carcinoma progression and metastasis?

TROVE2 has been identified as a significant contributor to hepatocellular carcinoma (HCC) progression through its effects on cellular invasion and migration. Research has demonstrated that TROVE2 is upregulated in HCC samples compared to normal liver tissue, and this elevated expression correlates with poor prognosis in liver cancer patients . Experimental evidence from cellular models has established a direct functional role, as overexpression of TROVE2 in HCC-LM3 cells enhanced invasion and migration capabilities, while knockdown of TROVE2 in HepG2 cells suppressed these processes . At the molecular level, TROVE2 has been shown to promote the epithelial-mesenchymal transition (EMT), a critical process in cancer metastasis, by modulating the expression of EMT-related proteins. Specifically, overexpression of TROVE2 decreases β-catenin and epithelial marker E-cadherin levels while increasing mesenchymal markers such as Snail, Slug, N-cadherin, and vimentin . Mechanistically, TROVE2 appears to regulate these changes through the HPSE/GSK-3β/Snail pathway, promoting the expression of phosphorylated GSK-3β, which inhibits GSK-3β activity . Since GSK-3β normally promotes Snail degradation, TROVE2-mediated inhibition of GSK-3β leads to increased Snail stability, thereby enhancing the EMT process and facilitating cancer cell invasion and metastasis. These findings collectively suggest that TROVE2 could serve as a novel therapeutic target for developing strategies to prevent or treat liver cancer.

What experimental approaches are most effective for studying TROVE2 function in cancer models?

The investigation of TROVE2 function in cancer models requires a multi-faceted experimental approach to comprehensively characterize its roles and mechanisms. Cell culture-based studies have proven effective, with multiple cell lines exhibiting differential TROVE2 expression levels that can be leveraged for comparative analyses. HepG2 and QGY-7701 cells show significantly higher expression of TROVE2 protein compared to HCC-LM3 and LO-2 cells, making these systems valuable for studying the effects of TROVE2 in different cellular contexts . Genetic manipulation through overexpression and knockdown approaches has been successfully implemented to modulate TROVE2 levels. Plasmid-based overexpression using vectors like pENTER-TROVE2 shuttle plasmid provides a method for increasing TROVE2 levels in low-expressing cells, while RNA interference techniques can effectively reduce TROVE2 expression in high-expressing cells . Functional assays such as Transwell assays have been particularly informative for assessing the impact of TROVE2 on cancer cell invasion and migration capabilities, directly linking molecular changes to phenotypic outcomes . For mechanistic studies, protein expression analysis of EMT markers and signaling pathway components through western blotting has revealed how TROVE2 influences cellular phenotypes. When designing animal models, researchers should consider orthotopic implantation of TROVE2-manipulated cells to recapitulate the appropriate tumor microenvironment, particularly for studying metastasis in vivo.

How can the HPSE/GSK-3β/Snail pathway regulated by TROVE2 be targeted for therapeutic intervention?

The HPSE/GSK-3β/Snail pathway regulated by TROVE2 represents a promising avenue for therapeutic intervention in hepatocellular carcinoma. Research has established that TROVE2 promotes the expression of phosphorylated GSK-3β (p-GSK-3β), which inhibits GSK-3β activity and consequently prevents Snail degradation, thereby enhancing the epithelial-mesenchymal transition (EMT) process . For direct targeting of TROVE2, RNA interference approaches using siRNAs or shRNAs have demonstrated efficacy in experimental settings, reducing TROVE2 expression and consequently inhibiting cancer cell invasion and migration . Alternatively, researchers could develop small molecule inhibitors that disrupt the interaction between TROVE2 and its downstream effectors in the HPSE/GSK-3β/Snail pathway. Targeting GSK-3β phosphorylation might also prove effective, as preventing the inhibition of GSK-3β could restore its activity in promoting Snail degradation, thereby reversing the EMT process. For experimental validation of pathway targeting, researchers should employ a combination of biochemical assays to measure GSK-3β activity and phosphorylation status, along with assessment of Snail protein stability and transcriptional activity. Patient-derived xenograft models would be particularly valuable for preclinical testing of therapeutic strategies targeting this pathway, as they better recapitulate the heterogeneity and complexity of human tumors. Combination approaches that simultaneously target multiple components of the HPSE/GSK-3β/Snail pathway might yield synergistic effects and reduce the likelihood of resistance development.

What cell models are most appropriate for studying various aspects of TROVE2 function?

Selection of appropriate cell models is crucial for investigating specific aspects of TROVE2 function across different research contexts. For hepatocellular carcinoma research, multiple established cell lines with differential TROVE2 expression have been characterized, providing valuable experimental systems. HepG2 and QGY-7701 cells exhibit high endogenous TROVE2 expression, making them suitable for knockdown studies, while HCC-LM3 cells show lower expression and are appropriate for overexpression experiments . The non-tumorigenic liver cell line LO-2 serves as an important control representing normal hepatocytes . For autoimmune disease research, immune cell models capable of producing anti-TROVE2 antibodies would be most relevant, with B lymphocyte cell lines or primary B cells from patients with autoimmune conditions being particularly valuable. When studying RNA-binding functions of TROVE2, cell models should be selected based on the expression of relevant RNA targets and interacting partners, which may vary across different cell types. For mechanistic studies of TROVE2 in the HPSE/GSK-3β/Snail pathway, cells with well-characterized baseline activity of this pathway should be prioritized to clearly discern TROVE2-specific effects. When investigating tissue-specific functions, researchers should consider using primary cells or organoid models that better recapitulate the native cellular environment compared to immortalized cell lines. Finally, for translational research with clinical implications, patient-derived cell models can provide insights into how TROVE2 function may vary across different genetic backgrounds and disease states.

What are the optimal approaches for TROVE2 knockdown and overexpression in experimental systems?

Effective genetic manipulation of TROVE2 requires careful consideration of experimental design to achieve robust and specific modulation of expression levels. For overexpression studies, plasmid-based approaches using vectors such as pENTER-TROVE2 shuttle plasmid have been successfully employed in research settings . When designing overexpression constructs, researchers should consider incorporating epitope tags (such as FLAG or HA) to facilitate detection and purification, while ensuring these modifications do not interfere with TROVE2 function. For transient overexpression, lipid-based transfection methods have proven effective in various cell types, while stable overexpression can be achieved using viral vectors followed by appropriate selection. For knockdown experiments, RNA interference through siRNA or shRNA targeting TROVE2 has been successfully implemented to reduce protein expression levels . When designing siRNAs, researchers should carefully select target sequences unique to TROVE2 to avoid off-target effects, and validate knockdown efficiency at both mRNA and protein levels. For more complete gene knockout, CRISPR-Cas9 genome editing technology offers an advanced approach to generate TROVE2-null cell lines for studying loss-of-function phenotypes. Regardless of the approach used, experimental designs should include appropriate controls such as scrambled siRNAs or empty vectors to account for non-specific effects of the manipulation methodology. Finally, rescue experiments, where TROVE2 expression is restored in knockdown or knockout cells, provide powerful validation that observed phenotypes are specifically due to TROVE2 modulation rather than off-target effects.

What assays can effectively measure TROVE2-mediated effects on cellular phenotypes?

A comprehensive assessment of TROVE2-mediated effects on cellular phenotypes requires a strategic combination of functional and molecular assays. For investigating cancer-related phenotypes, Transwell assays have proven particularly effective in measuring cell invasion and migration capabilities influenced by TROVE2 expression levels . These assays provide quantifiable data on the ability of cells to traverse a membrane barrier, often coated with extracellular matrix components to simulate in vivo conditions. For assessing epithelial-mesenchymal transition (EMT), both morphological evaluation and molecular profiling of EMT markers should be conducted. Western blotting analysis to measure expression changes in epithelial markers (E-cadherin, β-catenin) and mesenchymal markers (N-cadherin, vimentin, Snail, Slug) provides crucial molecular evidence of EMT progression or inhibition . For signaling pathway analysis, phosphorylation-specific antibodies can detect changes in key signaling molecules like p-GSK-3β, revealing how TROVE2 modulates specific pathways . RNA immunoprecipitation (RIP) assays would be valuable for identifying RNA targets directly bound by TROVE2, providing insights into its function as an RNA-binding protein. For in vivo relevance, metastasis assays in animal models using TROVE2-manipulated cells can determine the impact on tumor dissemination and colonization of distant sites. Finally, correlation studies between TROVE2 expression and clinical outcomes in patient samples provide critical translational relevance to experimental findings, connecting cellular phenotypes to disease progression.

What factors might explain variability in anti-TROVE2 antibody detection and prediction accuracy?

Variability in anti-TROVE2 antibody detection and its predictive accuracy for clinical outcomes can be attributed to multiple technical and biological factors that researchers must consider. Assay methodology significantly impacts detection sensitivity and specificity, with immunofluorescent-based ELISA demonstrating different performance characteristics compared to protein microarray platforms . Epitope specificity of detection antibodies is particularly crucial, as anti-TROVE2 antibodies may target different regions of the protein, potentially leading to variable detection depending on the assay design. Patient heterogeneity introduces substantial biological variability, as genetic background, disease duration, and concurrent medications can all influence anti-TROVE2 antibody production and characteristics. Studies have shown that prediction models incorporating multiple biomarkers, such as a panel of 8 biomarkers including TROVE2, SSB, NDE1, ZHX2, SH3GL1, CARD9, PTPN20, and KLHL12, achieve higher accuracy (79.0%) compared to individual markers . Sample handling and storage conditions can affect antibody stability and detection, with variations in collection, processing, and preservation potentially introducing pre-analytical variability. For analytical considerations, the statistical approaches used for cutoff determination and prediction model development significantly impact the reported accuracy, with machine learning approaches like random forest analysis showing particular utility in identifying stable biomarker panels . When designing studies to address these variability factors, researchers should implement rigorous standardization of sample handling, assay protocols, and data analysis pipelines, while also including appropriate technical and biological replicates to quantify variability at different levels.