Recombinant Human Transmembrane protein 87A (TMEM87A)
Description
Introduction to TMEM87A
TMEM87A belongs to the TMEM87 family of eukaryotic transmembrane proteins, which includes two members (TMEM87A and TMEM87B) in humans . The protein has garnered increasing scientific interest due to its diverse proposed functions including roles in protein transport to and from the Golgi apparatus, potential function as an ion channel, and involvement in developmental signaling pathways . This multifaceted protein has become the subject of intensive investigation across various research domains.
The TMEM87A gene encodes a 63-kDa protein characterized by an N-terminal Golgi signal sequence and seven transmembrane (TM) domains . In humans, three distinct isoforms have been identified: isoform 1 (full-length with Golgi signal sequence and transmembrane domains), isoform 2 (lacking transmembrane domains), and isoform 3 (lacking the predicted Golgi signal sequence) . These structural variations suggest potentially diverse functions within cellular systems.
Research interest in TMEM87A has intensified following discoveries linking it to various physiological and pathological processes. Recent studies have suggested its reclassification as GolpHCat (Golgi pH-sensitive Cation channel) to better reflect its functional properties in maintaining Golgi pH homeostasis . Additionally, disruptions in TMEM87A expression or function have been associated with cancers and developmental disorders, highlighting its clinical significance .
Cellular Localization and Expression Patterns
TMEM87A demonstrates specific cellular localization patterns that align with its proposed functions. The protein is predominantly localized to the Golgi apparatus, consistent with the presence of an N-terminal Golgi signal sequence . This localization is particularly significant given the protein's role in maintaining Golgi homeostasis, especially with regard to pH regulation .
Expression analysis reveals that TMEM87A is widely distributed across various cell types. According to brain RNA-seq database findings, TMEM87A is highly expressed in neurons and astrocytes within the brain . This neural expression pattern suggests potential roles in brain function and development, which is further supported by studies showing that TMEM87A knockout mice exhibit impaired spatial memory with significantly reduced long-term potentiation in the hippocampus .
When heterologously overexpressed, EGFP-tagged TMEM87A has been observed not only in the Golgi but also in the plasma membrane . This expanded distribution pattern under overexpression conditions provides experimental advantages for functional studies but should be interpreted with caution when considering the protein's native physiological roles.
Table 2: TMEM87A Expression and Localization
Functional Mechanisms of TMEM87A
The functional characteristics of TMEM87A have been the subject of some scientific debate, with different studies suggesting various potential mechanisms. A significant body of evidence supports TMEM87A's role as a cation channel, particularly within the Golgi apparatus .
Electrophysiological studies using heterologous expression systems have demonstrated that TMEM87A mediates voltage-dependent membrane currents with distinctive properties. The protein displays a non-linear current-voltage relationship with a reversal potential near -7.7 mV and pronounced inward rectification near -150 mV . The average rectification index value from +100 mV to -150 mV has been measured at 2.7 ± 0.3, indicating significant inward rectification .
Further characterization through voltage-step pulse experiments revealed that TMEM87A-mediated current exhibits voltage-dependent inward rectification without time- and voltage-dependent inactivation . These properties are consistent with those of a specialized ion channel involved in maintaining electrochemical gradients across the Golgi membrane.
Single-channel recordings of purified TMEM87A reconstituted in liposomes have provided additional insights into its functional properties. The channel displays stochastic single channel openings at both positive and negative holding potentials but remains closed at 0 mV . The channel's open probability (Po) increases non-linearly at both negative and positive potentials, reaching maximum values of approximately 0.6 at +90 mV and 0.3 at -150 mV .
Beyond its potential channel function, TMEM87A is implicated in maintaining Golgi pH homeostasis, as genetic ablation of the protein results in impaired resting pH in the Golgi . This function may be crucial for proper protein glycosylation and trafficking, which are essential processes for cellular function.
Pathological Implications of TMEM87A
TMEM87A has been implicated in various pathological conditions, highlighting its clinical significance. Of particular interest is its potential role in cancer development and progression. A notable case involves the identification of a TMEM87A-RASGRF1 fusion in a non-small cell lung cancer (NSCLC) patient who demonstrated an exceptional response to sunitinib treatment .
The TMEM87A-RASGRF1 fusion was discovered through RNA sequencing in a never-smoker patient who exhibited a partial response to sunitinib lasting 33 months . This fusion represents a novel oncogenic driver, as RASGRF1 encodes a guanine exchange factor that activates RAS by facilitating the conversion from GDP-RAS to GTP-RAS . The fusion protein lacks the N-terminal regulatory PH1 domain of RASGRF1 but retains the motif of the ERK-inducing PH2 domain, potentially leading to constitutive activation of the RAS signaling pathway .
Validation studies using CRISPR-Cas9-edited cell models demonstrated the oncogenic potential of this fusion. NIH/3T3 cells expressing the TMEM87A-RASGRF1 fusion formed foci with marked pile-up, whereas parental cells were inhibited from growing when they became confluent . Additionally, PC9 cells edited to express this fusion showed activation of MAPK, though sensitivity to MAPK inhibition was observed without apparent sensitivity to sunitinib .
Beyond cancer, TMEM87A has been linked to neurological functions and potentially to neurodevelopmental disorders. Studies in TMEM87A-knockout mice revealed dilated Golgi morphology, altered glycosylation and protein trafficking in the hippocampus, leading to impaired spatial memory with significantly reduced long-term potentiation . These findings suggest that TMEM87A dysfunction may contribute to cognitive impairments and possibly neurodevelopmental conditions.
Recombinant TMEM87A: Production and Applications
Recombinant TMEM87A proteins have become valuable tools for investigating the structure, function, and potential therapeutic applications of this protein. The production of full-length human TMEM87A typically involves expression systems such as E. coli, with the addition of tags (such as His-tag) to facilitate purification and detection .
Commercially available recombinant full-length human TMEM87A protein typically encompasses amino acids 22-555 of the mature protein, fused to an N-terminal His tag . These proteins are often supplied in lyophilized powder form for stability and ease of storage .
Table 3: Specifications of Recombinant Human TMEM87A
The applications of recombinant TMEM87A are diverse and include:
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Structural studies: Purified recombinant TMEM87A has been instrumental in determining the protein's three-dimensional structure through techniques such as cryo-EM .
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Functional characterization: Reconstitution of purified TMEM87A in liposomes has allowed for detailed electrophysiological studies to characterize its channel properties .
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Antibody production: Recombinant proteins serve as antigens for generating specific antibodies against TMEM87A, which are essential tools for detection and localization studies.
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Drug screening: Recombinant TMEM87A can be used in high-throughput screening assays to identify potential modulators of its activity.
Research on TMEM87A continues to evolve, with several key areas of focus emerging in recent years. One significant area involves resolving the apparent contradictions in the literature regarding TMEM87A's function as an ion channel. While substantial electrophysiological evidence supports this role , structural analyses have raised questions about this function .
Another active area of research involves exploring the therapeutic implications of TMEM87A, particularly in cancer treatment. The identification of the TMEM87A-RASGRF1 fusion as an oncogenic driver in NSCLC suggests that targeting this fusion or its downstream pathways could represent a novel therapeutic strategy . The exceptional response to sunitinib observed in a patient harboring this fusion warrants further investigation into potential targeted therapies .
Neurological functions of TMEM87A also represent an emerging area of interest. Given the observed cognitive deficits in TMEM87A-knockout mice and the protein's high expression in brain cells, further investigation into its roles in neural development, synaptic function, and cognition is warranted . This research direction could potentially yield insights into neurodevelopmental and neurodegenerative conditions.
Technological advances continue to enhance our ability to study TMEM87A. The development of improved recombinant protein production methods, more sophisticated structural analysis techniques, and refined genetic manipulation tools will likely accelerate progress in understanding this protein's complex biology.
Product Specs
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Target Background
Q&A
What is the cellular localization of TMEM87A?
TMEM87A predominantly localizes to the Golgi apparatus, as confirmed through multiple immunofluorescence studies. While native TMEM87A is mainly concentrated in the Golgi of human astrocytes, heterologously overexpressed EGFP-tagged TMEM87A can be detected both in the Golgi and at the plasma membrane . This dual localization in overexpression systems has facilitated electrophysiological characterization via patch-clamp recordings. For accurate localization studies, researchers should employ co-staining with established Golgi markers and subcellular fractionation techniques to quantify distribution across compartments.
What is the molecular structure of TMEM87A?
TMEM87A is a GOLD-domain seven-transmembrane helix protein with structural features similar to ion-conducting opsins . The protein contains a critical GYG sequence (positions 318-320) that appears essential for its ion channel function, as mutation of this sequence to AAA abolishes current in electrophysiological recordings . Cryo-EM structural analysis has resolved the lipid-bound form of TMEM87A at approximately 4.7 Å resolution . When investigating structure-function relationships, researchers should consider targeted mutagenesis of key domains, particularly the GYG sequence and other potential pore-forming regions, followed by functional assays.
What expression systems are appropriate for producing recombinant TMEM87A?
For research applications, E. coli-derived human TMEM87A recombinant protein (specifically positions D38-E555) has been successfully used for antibody production and validation . For functional studies, CHO-K1 cells have proven effective for heterologous expression of TMEM87A, allowing both localization studies and electrophysiological recordings . When expressing TMEM87A in heterologous systems, researchers should verify correct folding and trafficking through surface biotinylation assays, which have confirmed the presence of both wild-type TMEM87A and mutant variants at the cell surface .
How do you reconcile contradictory findings about TMEM87A's channel activity?
This apparent contradiction can be resolved through methodological considerations:
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Recording conditions: Earlier studies focused specifically on mechanosensitive properties, while later work examined voltage-dependent characteristics.
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Experimental approach: Whole-cell patch-clamp recordings from cells expressing TMEM87A revealed channel activity that may not have been detectable in the liposome reconstitution system.
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Mutation studies: The critical role of the GYG sequence suggests specific structural requirements for channel function that may have been compromised in earlier reconstitution attempts.
To address these contradictions, researchers should employ multiple complementary approaches, including both cellular and reconstituted systems, and carefully control for protein orientation, post-translational modifications, and interaction partners.
What are the biophysical properties of TMEM87A-mediated currents?
TMEM87A mediates voltage-dependent inwardly rectifying membrane currents with distinct properties:
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Current-voltage relationship: Non-linear with a reversal potential near -7.7 mV and pronounced inward rectification near -150 mV .
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Rectification index: The average rectification index from +100 mV to -150 mV is 2.7 ± 0.3 .
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Inactivation properties: TMEM87A currents show no time- or voltage-dependent inactivation .
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pH sensitivity: The channel is sensitive to pH, functioning as a pH-sensitive cation channel .
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Ion selectivity: TMEM87A is a non-selective cation channel .
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Inhibition profile: The channel is potently inhibited by gluconate .
For rigorous biophysical characterization, researchers should employ a combination of whole-cell and single-channel recordings across a range of voltages, pH conditions, and ionic compositions to fully delineate channel properties.
How should researchers approach TMEM87A knockout studies?
Genetic ablation of TMEM87A in mice has revealed significant phenotypes affecting Golgi morphology, protein trafficking, and cognitive function . When designing knockout studies, researchers should consider:
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Cell-specific vs. global knockouts: Different phenotypes may emerge based on the cell types affected.
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Temporal control: Inducible knockout systems may help distinguish developmental from acute effects.
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Compensation: Closely related proteins (e.g., TMEM87B) may compensate for TMEM87A loss.
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Readouts: Multiple assays should be employed to assess:
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Golgi morphology and pH homeostasis
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Protein glycosylation and trafficking
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Cellular function (particularly in neurons and astrocytes)
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Behavioral phenotypes in animal models
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The observation that TMEM87A knockout mice exhibit dilated Golgi morphology, altered glycosylation and protein trafficking, and impaired spatial memory with reduced long-term potentiation suggests that comprehensive phenotyping across cellular, tissue, and behavioral levels is essential.
What methodologies are most effective for studying TMEM87A's ion channel function?
To rigorously characterize TMEM87A's channel properties, researchers should consider:
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Heterologous expression systems: CHO-K1 cells have proven effective for electrophysiological studies of TMEM87A .
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Voltage protocols:
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Ramp protocols from -150 mV to +100 mV reveal the non-linear I-V relationship
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Step protocols from +100 mV to -150 mV demonstrate the lack of time- and voltage-dependent inactivation
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Mutagenesis: The GYG sequence (positions 318-320) is critical for channel function and should be a focal point for structure-function studies .
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Reconstitution: Purified TMEM87A has been successfully reconstituted in liposomes, enabling single-channel recordings .
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Pharmacological manipulation: Gluconate has been identified as a potent inhibitor and can serve as a pharmacological tool .
The combination of cellular electrophysiology and reconstitution approaches provides complementary insights into channel function in different contexts.
How does TMEM87A contribute to Golgi function and neurological processes?
TMEM87A (GolpHCat) maintains Golgi membrane potential, regulating ionic and osmotic homeostasis, which in turn affects protein glycosylation and trafficking . In the brain, where TMEM87A is expressed in various cell types including neurons and astrocytes, its function appears critical for proper hippocampal function .
The pathway from molecular function to neurological effects can be studied through:
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Golgi pH measurements in wild-type versus TMEM87A-deficient cells
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Glycosylation analysis of secreted and membrane proteins
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Trafficking assays for Golgi-processed proteins
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Electrophysiological recordings of hippocampal neurons
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Behavioral testing focusing on spatial memory tasks
The observation that TMEM87A knockout mice exhibit impaired spatial memory and reduced long-term potentiation suggests that this protein represents a novel molecular target for investigating cognitive impairment and potentially developing therapeutics for Golgi-related diseases.
What antibody validation is necessary for TMEM87A studies?
Proper antibody validation is critical for reliable TMEM87A research. The anti-TMEM87A antibody has been validated through multiple approaches :
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Western blot: Detecting bands at approximately 70 kDa across various human cell lines (T-47D, MDA-MB-453, PC-3, MCF-7), rat tissues (brain, PC-12 cells), and mouse tissues (brain, RAW264.7 cells) .
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Immunohistochemistry: Successful detection in paraffin-embedded sections of multiple tissues, including human colon adenocarcinoma, liver cancer, esophageal squamous carcinoma, testicular seminoma, lung adenocarcinoma, and normal brain tissues from both rat and mouse .
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Immunofluorescence: Detection in U87 cells with appropriate subcellular localization .
Researchers should perform similar validation in their specific experimental systems, particularly when studying novel tissue types or disease models.
How can researchers distinguish between TMEM87A's roles in ion transport versus protein trafficking?
TMEM87A plays dual roles in ion transport and retrograde transport in the Golgi . To differentiate between these functions:
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Selective mutations: Generate mutations that specifically disrupt channel function (e.g., GYG sequence) versus trafficking domains.
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Acute manipulation: Use rapid inhibition approaches (pharmacological when available) to distinguish immediate ion transport effects from longer-term trafficking consequences.
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Rescue experiments: Attempt rescue of knockout phenotypes with wild-type versus channel-dead mutants to determine which functions underlie specific phenotypes.
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Direct measurement: Simultaneously monitor both ion transport (using fluorescent indicators) and protein trafficking (using tagged cargo proteins) in the same cells under various manipulations.
This multi-faceted approach can help parse the relative contributions of TMEM87A's distinct functions to cellular physiology and disease pathology.
What is the evidence for TMEM87A involvement in disease?
TMEM87A has been implicated in several pathological conditions:
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Cancer: TMEM87A expression has been detected in various cancer tissues, including colon adenocarcinoma, liver cancer, esophageal squamous carcinoma, testicular seminoma, and lung adenocarcinoma .
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Heart disease: TMEM87A has been implicated in cardiac pathology, though the mechanisms remain to be fully elucidated .
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Neurological function: TMEM87A knockout mice display impaired spatial memory and reduced long-term potentiation in the hippocampus, suggesting potential roles in cognitive disorders .
Research approaches to explore disease connections should include:
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Expression analysis in patient samples versus controls
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Genetic association studies
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Functional studies in disease-relevant cell types
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Animal models of specific diseases with TMEM87A manipulation
The emerging role of TMEM87A in Golgi homeostasis suggests it may be particularly relevant to diseases involving protein trafficking and glycosylation abnormalities.
How should researchers approach drug discovery targeting TMEM87A?
For researchers considering TMEM87A as a therapeutic target:
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Assay development: Establish high-throughput screening assays based on:
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Ion channel activity using fluorescent ion indicators or automated electrophysiology
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Golgi pH homeostasis using pH-sensitive fluorescent proteins targeted to the Golgi
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Protein trafficking efficiency using reporter proteins
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Structure-based design: Utilize the available structural information on TMEM87A to design compounds that might modulate its function.
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Validation strategy:
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Primary screens in heterologous expression systems
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Secondary validation in disease-relevant cell types
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Tertiary validation in animal models of relevant diseases
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Potential therapeutic applications:
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Cognitive enhancement for memory disorders
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Targeting cancer cells that may depend on altered Golgi function
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Addressing specific glycosylation disorders
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