Recombinant Human Transmembrane protein 208 (TMEM208)

What is TMEM208 and where is it localized in cells?

TMEM208 (Transmembrane Protein 208) is an endoplasmic reticulum (ER) protein involved in the signal-independent pathway that facilitates the translocation of nascent proteins into the ER. It is part of the machinery that handles proteins destined for the cell membrane and secretory pathway . The protein has been identified as broadly expressed across multiple tissue types, with particularly notable expression in various cancers including Head and Neck Squamous Cell Carcinoma (HNSCC) . Subcellular localization studies demonstrate that TMEM208 is primarily found in the endoplasmic reticulum membrane, consistent with its role in protein translocation and processing .

What are the known biological functions of TMEM208?

TMEM208 has several documented functions that highlight its importance in cellular processes:

• Protein translocation: It participates in the transport of nascent polypeptides into the endoplasmic reticulum via the signal recognition particle (SRP) pathway .

• Autophagy regulation: TMEM208 has been reported to be associated with cellular autophagy, which is strongly tied to the onset and development of numerous illnesses, including cancer .

• Developmental processes: Knockout studies in Drosophila have revealed that TMEM208 is essential for proper development, as its loss causes lethality and developmental defects related to cell polarity .

• Cell polarity maintenance: TMEM208 physically interacts with Frizzled (Fz), a planar cell polarity (PCP) receptor, and is required to maintain proper levels of Fz protein .

• ER homeostasis: Loss of TMEM208 has been associated with mild ER stress in both fruit fly models and human patient fibroblasts .

How does TMEM208 expression vary across different tissue types?

Analysis of TCGA data demonstrates that TMEM208 expression varies significantly across tissue types. The protein shows particularly high expression in several malignant tumor tissues, with HNSCC being one of the tumor types where TMEM208 is substantially expressed . Immunohistochemical staining has confirmed deepened cytoplasmic staining in cancer tissues compared to normal counterparts, and in vitro cell line analysis shows high expression in squamous carcinoma cell lines . Gender-based analysis has indicated that TMEM208 is significantly more highly expressed in male HNSCC patients than female patients, and expression levels also differ by HPV status, with significantly higher expression in HPV-negative patients .

What molecular mechanisms underlie TMEM208's role in cancer progression?

TMEM208 appears to promote cancer progression through multiple mechanisms:

How does TMEM208 influence immune cell infiltration in the tumor microenvironment?

Analysis of the relationship between TMEM208 expression and immune cell infiltration has revealed several important patterns:

• Reduction of immune cell infiltration: In samples with high TMEM208 expression, CIBERSORTx analysis showed significantly reduced infiltration of B cell naive, T cell CD4 memory resting, NK cells resting, NK cells activated, and neutrophils compared to samples with low expression .

• Negative correlation with anti-tumor immune cells: TIMER and TISIDB database analyses demonstrated that TMEM208 expression was negatively correlated with the infiltration of numerous immune cells such as B cells, CD8+T, CD4+T, neutrophils, dendritic cells, T follicular helper cells, NK cells, NKT cells, and mast cells .

• Association with immune checkpoints: Correlation analysis showed that TMEM208 expression was positively correlated with CD24, CD276, LAG3, and HVEM immune checkpoints . This is significant because:

  • CD276 and LAG-3 can inhibit T cell proliferation and activation

  • CD24 inhibits macrophage phagocytosis of tumors

  • HVEM interacts with T cells, B cells, and NK cells, producing both immunosuppressive and immune-activating signals

These findings suggest that TMEM208 may play a critical role in immune escape mechanisms of HNSCC and potentially other cancers, making it relevant for immunotherapy approaches .

What is the significance of TMEM208 in developmental processes and disorders?

The developmental importance of TMEM208 has been highlighted through several lines of evidence:

• Lethality in model organisms: CRISPR-induced null allele studies in Drosophila showed that loss of Tmem208 causes lethality, with only a few short-lived flies eclosing .

• Developmental defects: The surviving Tmem208-null flies exhibited wing and eye developmental defects consistent with impaired cell polarity .

• Human developmental disorders: A case study identified a child with compound heterozygous variants in TMEM208 who presented with developmental delay, skeletal abnormalities, multiple hair whorls, cardiac and neurological issues - symptoms associated with planar cell polarity (PCP) defects in mice and humans .

• Functional validation: Expression of reference human TMEM208 in flies fully rescued the loss of Tmem208, while the two proband-specific variants failed to rescue, suggesting they are loss-of-function alleles .

These findings collectively demonstrate that TMEM208 plays an essential role in proper development, likely through its effects on ER homeostasis and cell polarity .

What experimental approaches are most effective for studying TMEM208 function?

Based on published research, several methodological approaches have proven valuable for investigating TMEM208:

• Bioinformatic analysis using cancer databases:

  • UALCAN, HPA, CVCDAP, DAVID, TIMER, CIBERSORTx, TISIDB, and cBioPortal online databases have been successfully used to analyze TMEM208 expression, prognostic significance, and immune infiltration correlations .

  • These approaches allow for comprehensive analysis of gene expression patterns, survival correlations, and functional associations across large patient cohorts.

• In vitro cellular studies:

  • Cell culture models using squamous carcinoma cell lines have confirmed high TMEM208 expression .

  • These models can be used to evaluate the effects of TMEM208 manipulation on cellular processes.

• Gene editing in model organisms:

  • CRISPR-induced null alleles in Drosophila, such as replacing the gene with Kozak-GAL4 sequence, have provided insights into developmental roles .

  • Rescue experiments with human TMEM208 variants in flies offer an efficient system for functional validation .

• Patient-derived cell studies:

  • Fibroblasts from patients with TMEM208 mutations have been used to assess ER stress and other cellular phenotypes .

  • These provide direct evidence of pathophysiological mechanisms in human cells.

How can researchers accurately measure and manipulate TMEM208 expression?

For accurate measurement and manipulation of TMEM208, several approaches have been validated:

• Expression analysis:

  • mRNA expression: RNA-seq and qPCR analysis from databases like TCGA provide reliable quantification of TMEM208 at the transcript level .

  • Protein expression: Immunohistochemical staining has successfully detected TMEM208 in tissue samples, showing deepened cytoplasmic staining in cancer tissues .

• Experimental manipulation:

  • Gene knockout: Complete replacement of the gene with reporter constructs (e.g., Kozak-GAL4) has been effective in Drosophila .

  • Expression of variant forms: Transgenic expression of human reference or variant TMEM208 in model organisms allows for functional complementation testing .

• Interaction studies:

  • Co-immunoprecipitation has identified physical interactions between TMEM208 and other proteins, such as the Frizzled receptor .

  • This approach can uncover potential mechanisms and pathways involving TMEM208.

What analytical techniques help determine TMEM208's role in disease pathogenesis?

Several analytical approaches have proven valuable in understanding TMEM208's role in disease:

• Survival and prognostic analysis:

  • Kaplan-Meier survival curves comparing high versus low TMEM208 expression groups can reveal prognostic significance .

  • Cox regression analysis helps determine if TMEM208 expression is an independent risk factor, controlling for other clinical variables .

• Immune infiltration analysis:

  • CIBERSORTx analysis can evaluate the relationship between TMEM208 expression and the abundance of 22 immune cell types .

  • TIMER and TISIDB databases help investigate correlations between TMEM208 and specific immune cell populations .

• Functional enrichment analysis:

  • Gene set variation analysis (GSVA) can identify biological processes and pathways associated with TMEM208 expression .

  • This approach has revealed TMEM208's relationships with ribosomal/mitochondrial functions and immune cell differentiation .

• Developmental phenotyping:

  • Careful characterization of model organism phenotypes (wing/eye defects in Drosophila) can provide insights into developmental roles .

  • Correlation with human patient symptoms enables translational understanding of pathogenic mechanisms .

What is the prognostic value of TMEM208 in different cancer types?

TMEM208 has demonstrated significant prognostic value, particularly in HNSCC:

These findings suggest that TMEM208 expression level could serve as a valuable biomarker for risk stratification and treatment planning in HNSCC patients .

How might TMEM208 be targeted in cancer therapy approaches?

Based on its biological functions and correlations, several potential therapeutic approaches targeting TMEM208 can be considered:

• Direct targeting strategies:

  • Inhibition of TMEM208 expression or function could potentially reduce cancer cell survival and proliferation, given its association with ribosomal and mitochondrial functions that support tumor growth .

  • Since TMEM208 appears to be involved in ER protein translocation, targeting this function might disrupt cancer cell protein homeostasis .

• Immunotherapy implications:

  • The negative correlation between TMEM208 and immune cell infiltration suggests that inhibiting TMEM208 might enhance anti-tumor immune responses .

  • TMEM208's positive correlation with immune checkpoints (CD24, CD276, LAG3, HVEM) indicates that combination therapies targeting both TMEM208 and these checkpoints could potentially enhance immunotherapy efficacy .

• Personalized medicine approach:

  • Given the variation in TMEM208 expression across different patient populations (e.g., higher in males and HPV-negative patients), expression levels could guide patient selection for TMEM208-targeted therapies .

  • TMEM208 expression might serve as a biomarker to predict response to immune checkpoint inhibitor (ICI) treatment in HNSCC .

What developmental disorders are associated with TMEM208 dysfunction?

Research has identified a connection between TMEM208 variants and developmental abnormalities:

• Human case study:

  • A child with compound heterozygous variants in TMEM208 presented with developmental delay, dysmorphism, multiple hair whorls, seizures, and other developmental abnormalities involving multiple organs .

  • These symptoms are consistent with planar cell polarity (PCP) defects observed in mice and humans .

• Functional validation:

  • The patient-specific TMEM208 variants failed to rescue the developmental defects in Tmem208-null flies, confirming that these were loss-of-function mutations .

  • This provides strong evidence linking TMEM208 dysfunction to the observed developmental abnormalities .

• Cellular pathology:

  • Fibroblasts from the affected individual displayed mild ER stress, suggesting a mechanism by which TMEM208 dysfunction might disrupt normal development .

  • The interaction between TMEM208 and the Frizzled receptor (a key PCP component) further supports the role of TMEM208 in developmental processes .

These findings establish TMEM208 as a potential candidate gene for developmental disorders, particularly those involving cell polarity defects and multiple system abnormalities .

What are the most pressing unanswered questions about TMEM208?

Several critical questions about TMEM208 remain to be addressed:

• Molecular mechanisms:

  • How exactly does TMEM208 regulate the immune microenvironment in cancer?

  • What is the precise mechanism by which TMEM208 affects cell polarity and developmental processes?

  • What are the complete interaction networks of TMEM208 in different cell types?

• Therapeutic applications:

  • Can TMEM208 inhibition sensitize tumors to immune checkpoint inhibitors?

  • What is the efficacy and safety profile of targeting TMEM208 in cancer models?

  • How can TMEM208-targeting approaches be optimized for clinical application?

• Biomarker development:

  • Can TMEM208 expression be reliably measured in liquid biopsies for non-invasive cancer monitoring?

  • Does TMEM208 expression predict response to specific cancer therapies beyond immunotherapy?

  • What are the optimal cutoff values for TMEM208 expression in prognostic applications?

Addressing these questions will require multidisciplinary approaches combining molecular biology, biochemistry, immunology, and clinical research to fully understand and leverage TMEM208's biological significance.