Recombinant Human Transmembrane protein 218 (TMEM218)
Description
Gene Structure and Expression
The TMEM218 gene has been characterized across multiple species including humans, rats, mice, and other model organisms. In rats, the gene (Tmem218) was previously annotated as "similar to RIKEN cDNA 1810021J13" before being properly identified and renamed to reflect its human and mouse nomenclature . This conservation across species reinforces the importance of this gene in fundamental biological processes.
Expression analyses show that TMEM218 is expressed in multiple tissues. In Xenopus, expression has been detected in the brain, egg, endomesoderm, fat body, tail, testis, and throughout the whole organism . The broad expression pattern suggests widespread functions in various organ systems, consistent with the known role of cilia in multiple tissues.
Production and Properties of Recombinant Human TMEM218
Recombinant human TMEM218 protein is typically produced in expression systems to facilitate research applications and functional studies. The most common approach involves expressing the protein in bacterial systems like E. coli with affinity tags to enable purification.
Expression Systems and Production Methods
A commercially available recombinant full-length human TMEM218 protein is produced using E. coli expression systems. The protein is expressed as a fusion construct containing the complete human TMEM218 sequence (amino acids 1-115) with an N-terminal His-tag for purification purposes . This expression system allows for high-yield production of the protein in a form suitable for various research applications.
The recombinant protein is typically isolated through affinity chromatography leveraging the His-tag, followed by quality control assessments including SDS-PAGE to confirm purity and size. According to product specifications, the purity typically exceeds 90% as determined by SDS-PAGE analysis .
Physical and Chemical Properties
The recombinant human TMEM218 protein is generally supplied as a lyophilized powder to ensure stability during shipping and storage . The protein is formulated in a Tris/PBS-based buffer containing 6% trehalose with a pH of 8.0, which helps maintain protein stability .
For research applications, the lyophilized protein is typically reconstituted in deionized sterile water to a concentration of 0.1-1.0 mg/mL. For long-term storage, the addition of glycerol (typically 5-50% final concentration) is recommended to prevent protein degradation during freeze-thaw cycles .
The storage recommendations include keeping the protein at -20°C to -80°C for long-term storage, with working aliquots maintained at 4°C for up to one week to minimize degradation from repeated freeze-thaw cycles .
Biological Functions of TMEM218
Research using knockout models and protein interaction studies has revealed several important biological functions of TMEM218, particularly in relation to ciliary structure and function.
Role in Ciliary Structure and Function
TMEM218 appears to be a critical component in the maintenance of ciliary structure and function. It is required for proper ciliary biogenesis and maintenance, including roles in regulating cilia length and ensuring the appropriate number of cilia through modulation of centrosome duplication . These functions are essential for various developmental processes and organ functionality.
The protein is localized at the transition zone of primary cilia, where it acts as part of a complex that forms a barrier preventing the diffusion of transmembrane proteins between the ciliary and plasma membranes . This compartmentalization is crucial for proper ciliary signaling and function.
Developmental Roles
Studies in knockout mouse models (Tmem218-/-) have revealed significant developmental roles for TMEM218. These mice develop progressive cystic kidney disease and retinal degeneration, indicating crucial functions in both renal and ocular development and homeostasis .
The renal phenotype in these knockout mice is characterized by diffuse renal cyst development with tubulointerstitial nephropathy and disruption of tubular basement membranes, although the kidneys remain essentially normal-sized . This pathology closely resembles nephronophthisis (NPHP) in humans, suggesting a potential role for TMEM218 in this disease.
The retinal phenotype is characterized by slow-onset loss of photoreceptors, resulting in reduced electroretinogram responses . This resembles retinitis pigmentosa in humans, particularly when it occurs in conjunction with nephronophthisis as part of Senior-Løken syndrome.
TMEM218 in Disease Models and Pathological Conditions
The phenotypes observed in Tmem218-deficient mice have significant implications for understanding human diseases, particularly ciliopathies that affect the kidneys and eyes.
Nephronophthisis and Kidney Function
The renal cystic disease observed in Tmem218-/- mice closely resembles nephronophthisis in humans . Nephronophthisis is an autosomal recessive cystic kidney disease that leads to end-stage renal failure. It is characterized by tubulointerstitial fibrosis, tubular basement membrane disruption, and cyst formation.
Since known nephronophthisis genes collectively account for only about 30% of human NPHP cases, TMEM218 represents a potential candidate for involvement in the remaining unexplained cases . The mouse model provides a valuable tool for further investigating the pathogenesis of this condition and potential therapeutic interventions.
Retinal Degeneration and Visual Function
The retinal degeneration phenotype in Tmem218-/- mice, characterized by photoreceptor loss and reduced electroretinogram responses, mirrors aspects of retinitis pigmentosa in humans . This condition is often comorbid with nephronophthisis in a condition known as Senior-Løken syndrome.
The concurrent development of both renal and retinal pathologies in the knockout mouse model suggests that TMEM218 could be involved in Senior-Løken syndrome or other ciliopathies affecting both organs . This makes the Tmem218-/- mouse a potentially valuable model for studying these combined pathologies and developing therapeutic approaches.
Protein Interactions and Molecular Pathways
TMEM218 functions within a network of protein interactions that collectively contribute to ciliary formation and function. Understanding these interactions provides insight into the molecular mechanisms underlying TMEM218's biological roles.
The Tectonic-like Complex
TMEM218 appears to interact with components of the tectonic-like complex, a protein assembly localized at the transition zone of primary cilia. This complex acts as a barrier preventing the diffusion of transmembrane proteins between ciliary and plasma membranes .
Key interaction partners within this complex include:
-
TMEM17 - Transmembrane protein 17, a component of the tectonic-like complex required for ciliogenesis and Sonic Hedgehog (SHH) signaling
-
TMEM231 - Transmembrane protein 231, another component of the tectonic-like complex essential for ciliogenesis and SHH signaling
-
B9D1 - B9 domain-containing protein 1, required for ciliogenesis and SHH signaling
-
B9D2 - B9 domain-containing protein 2, part of the tectonic-like complex at the ciliary transition zone
-
MKS1 - Meckel syndrome type 1 protein, involved in centrosome migration during early ciliogenesis
These interactions suggest that TMEM218 functions as part of this larger protein complex to maintain the specialized compartmentalization of the cilium, which is essential for its sensory and developmental signaling functions.
Functional Biochemical Properties
At the biochemical level, TMEM218 has been shown to possess protein binding capabilities . This function likely facilitates its interactions with other components of the tectonic-like complex and additional proteins involved in ciliary formation and maintenance.
The protein binding capacity of TMEM218 places it within a functional network that includes other proteins with similar binding properties, such as TMEM51, ANTXR1, CLDN8, and others . These collective interactions form the basis for the assembly and maintenance of the ciliary transition zone and its barrier function.
Research Applications of Recombinant TMEM218
Recombinant human TMEM218 protein serves as a valuable research tool for investigating various aspects of ciliary biology and associated pathological conditions.
Functional Studies and Protein-Protein Interactions
The availability of purified recombinant TMEM218 enables detailed investigations of its interactions with other proteins. These studies can employ techniques such as pull-down assays, co-immunoprecipitation, and surface plasmon resonance to characterize binding partners and elucidate the molecular mechanisms underlying TMEM218's functions.
Such studies are particularly valuable for understanding how TMEM218 contributes to the assembly and function of the tectonic-like complex at the ciliary transition zone, and how disruptions in these interactions may lead to ciliopathies.
Antibody Production and Detection Methods
Recombinant TMEM218 protein is useful for generating specific antibodies that can be employed in various detection methods, including western blotting, immunohistochemistry, and immunofluorescence. These tools facilitate the study of TMEM218 expression, localization, and function in various tissues and experimental models.
The primary application of the commercially available recombinant TMEM218 protein is for SDS-PAGE analysis , which can be used to validate antibody specificity and for protein interaction studies.
Therapeutic Potential and Drug Development
The involvement of TMEM218 in ciliopathies suggests potential therapeutic applications. Recombinant TMEM218 can serve as a tool in high-throughput screening assays to identify compounds that might modulate its function or interactions, potentially leading to therapeutic interventions for conditions like nephronophthisis and retinal degeneration.
The Tmem218-/- mouse model, originally generated as part of efforts to identify and validate pharmaceutically tractable targets for drug development , represents a valuable resource for testing such interventions and understanding their effects on disease progression.
Future Research Directions
Despite the significant progress in understanding TMEM218, several aspects of its biology and potential clinical relevance remain to be fully elucidated.
Human Disease Associations
While the mouse model suggests potential involvement in nephronophthisis and retinal degeneration, direct evidence linking TMEM218 mutations to human ciliopathies is still limited. Future research should focus on screening patients with unexplained NPHP, retinitis pigmentosa, or Senior-Løken syndrome for mutations in TMEM218 to establish its clinical relevance.
The observation that known NPHP genes account for only about 30% of cases suggests that TMEM218 and other ciliary proteins could be involved in the remaining unexplained cases. Comprehensive genetic analyses of these patients could reveal new disease-causing mutations.
Product Specs
Note: While we prioritize shipping the format currently in stock, please specify your format preference during order placement for customized preparation.
Note: All proteins are shipped with standard blue ice packs. Dry ice shipping requires prior arrangement and incurs additional charges.
The tag type is determined during the production process. If you require a specific tag, please inform us, and we will prioritize its development.
Target Background
Q&A
What is the biological role of TMEM218 in human cells?
TMEM218 is a transmembrane protein that localizes to the transition zone of the primary cilium. The primary cilium functions as a microtubule-based, antenna-like organelle projecting from the surface of most human cell types, enabling cells to respond to extracellular signals. TMEM218 plays a critical role in maintaining the functional compartmentalization of the cilium, which is partitioned from the cell body by the transition zone. This zone represents a known hotspot for ciliopathy-related proteins, including TMEM218 .
How is TMEM218 structurally organized and what are its critical domains?
While detailed structural information is limited in current research, TMEM218 contains highly conserved amino acid sequences that are essential for its proper function. Position 115 appears particularly significant, as it represents a mutational hotspot found in multiple families with ciliopathy phenotypes . As a transmembrane protein, TMEM218 has membrane-spanning domains that anchor it to the ciliary transition zone where it performs its specialized functions.
What cellular pathways involve TMEM218?
TMEM218 functions primarily in the ciliary transition zone pathway, which regulates molecular traffic in and out of the cilium. Dysfunction of TMEM218 disrupts proper cilium formation and function, leading to disturbances in critical developmental signaling pathways that rely on the primary cilium. The protein's involvement in the ciliary transition zone suggests it plays a role in maintaining the composition of the ciliary compartment, which is essential for proper ciliary signaling functions .
What spectrum of disorders are linked to TMEM218 dysfunction?
TMEM218 dysfunction causes ciliopathies within the Joubert-Meckel syndrome spectrum, a continuum of recessive conditions resulting from primary cilium dysfunction. Specifically, TMEM218 variants have been associated with Joubert syndrome with retinal dystrophy, Meckel syndrome, and Senior-Loken syndrome . These conditions share overlapping features but vary in severity and organ involvement, reflecting the diverse roles of primary cilia across different tissues.
What are the phenotypic presentations in patients with TMEM218-related ciliopathies?
Clinical manifestations of TMEM218-related ciliopathies include the characteristic molar tooth sign on brain imaging (observed in 2 of 7 reported cases), occipital encephalocele (found in 5 fetuses), retinal dystrophy (present in all 4 living individuals studied), polycystic kidneys (2 cases), and polydactyly (2 cases) . Notably, liver involvement was not observed in the reported cases, distinguishing TMEM218-related ciliopathies from some other forms of ciliopathies where hepatic manifestations are common.
How does TMEM218 function correlate with ciliopathy phenotypes?
TMEM218 dysfunction disrupts the ciliary transition zone, compromising the primary cilium's ability to act as a signaling hub. This disruption affects multiple developmental and homeostatic signaling pathways, leading to the observed spectrum of congenital abnormalities . The connection between TMEM218 and ciliary transition zone function explains how mutations in this gene can cause diverse manifestations across different organ systems, all of which depend on proper ciliary function during development and for ongoing cellular homeostasis.
What types of genetic variants in TMEM218 are associated with disease?
Disease-causing TMEM218 variants predominantly consist of biallelic, rare, predicted-deleterious missense variants. A significant finding is that four of six families with ciliopathy phenotypes carry missense variants affecting the same highly conserved amino acid position 115, indicating this residue's critical importance for proper protein function . This mutation hotspot provides valuable insight into structure-function relationships within the TMEM218 protein.
What methodologies are used to identify and classify TMEM218 variants?
Identification of TMEM218 variants typically employs exome and targeted sequencing approaches. In the key study identifying TMEM218 as a ciliopathy gene, researchers sequenced 655 families with Joubert syndrome . Classification of variants involves analysis of evolutionary conservation, prediction of functional impact using bioinformatic tools, segregation analysis in families, and assessment of allele frequency in population databases. Functional studies in cellular or animal models may provide additional evidence for pathogenicity.
What proportion of ciliopathy cases remain genetically undiagnosed, and how might TMEM218 contribute to this gap?
Despite extensive gene discovery efforts, the genetic cause remains unidentified in up to 30% of individuals with Joubert syndrome, depending on the cohort, sequencing method, and criteria used for pathogenic variant classification . The identification of TMEM218 as a ciliopathy gene helps narrow this diagnostic gap. Continuing research into TMEM218 and other ciliary genes will likely reveal additional genetic causes, improving diagnostic rates and enabling better prognostic counseling for affected families.
What expression systems are optimal for producing recombinant human TMEM218?
While specific expression systems for TMEM218 are not detailed in the provided references, insights from studies on recombinant protein expression in CHO cells are applicable. Research shows that recombinant protein yield is influenced by transgene mRNA levels, protein-specific features, and host cell expression signatures . For transmembrane proteins like TMEM218, mammalian expression systems that support proper membrane protein folding and post-translational modifications would be most appropriate, with CHO cells representing a viable option given their established use in producing recombinant human proteins.
What factors influence the successful expression of recombinant TMEM218?
Key factors affecting recombinant protein expression include protein-specific features that impact productivity, cellular responses to ER stress, and secretory pathway activity . For TMEM218 specifically, optimization should consider its transmembrane nature and potential challenges in folding and trafficking. Successful expression strategies might involve monitoring and modulating the unfolded protein response, as cells that successfully produce recombinant protein show better adaptation to ER stress than failed producers, with upregulation of protein folding genes like HYOU1, ERO1A, and PDIA3 .
What purification strategies are most effective for recombinant TMEM218?
Purification of transmembrane proteins like TMEM218 typically requires specialized approaches. While not specifically addressed in the provided references, effective strategies would likely include:
-
Initial solubilization with appropriate detergents
-
Affinity chromatography using tags engineered into the recombinant protein
-
Size exclusion chromatography for final purification
-
Quality control using techniques such as circular dichroism to confirm proper folding
The membrane protein nature of TMEM218 presents unique purification challenges compared to soluble proteins, necessitating careful optimization of detergent types and concentrations to maintain native structure.
How can researchers investigate TMEM218 interactions within the ciliary transition zone complex?
Advanced investigation of TMEM218's protein-protein interactions within the ciliary transition zone would benefit from:
-
Proximity labeling techniques such as BioID or APEX to identify interacting partners in the intact cellular context
-
Co-immunoprecipitation followed by mass spectrometry to identify stable interactors
-
FRET or BRET assays to study dynamic interactions in living cells
-
Cryo-electron microscopy to visualize TMEM218 within the larger transition zone complex
These approaches would help elucidate how TMEM218 contributes to transition zone architecture and function, potentially revealing new therapeutic targets for ciliopathies.
What cellular models are most appropriate for studying TMEM218 function in ciliopathies?
Optimal cellular models for studying TMEM218 function include:
-
Primary ciliated cells from patients with TMEM218 variants
-
CRISPR-engineered cell lines with specific TMEM218 mutations, particularly those affecting position 115
-
iPSC-derived organoids that recapitulate ciliated tissues affected in ciliopathies (retina, kidney, brain)
-
3D spheroid cultures that better maintain ciliary structures compared to 2D cultures
These models allow investigation of how TMEM218 dysfunction affects ciliary formation, maintenance, and signaling functions across different tissue contexts relevant to the ciliopathy disease spectrum.
How might gene editing approaches be applied to correct TMEM218 mutations?
Therapeutic gene editing strategies for TMEM218-related ciliopathies could include:
-
CRISPR-Cas9 correction of specific mutations, particularly the recurrent position 115 variants
-
Base editing technologies for precise correction of missense mutations
-
Prime editing for more complex sequence changes
-
Evaluation of editing efficiency in patient-derived cells and appropriate animal models
Given the recessive nature of TMEM218-related ciliopathies, successful correction of one allele might be sufficient to mitigate disease severity . Proof-of-concept studies in cellular and animal models would be crucial before clinical translation.
What imaging techniques best visualize TMEM218 localization and dynamics?
Optimal imaging approaches for TMEM218 include:
-
Super-resolution microscopy (STED, PALM, STORM) to precisely localize TMEM218 within the ciliary transition zone
-
Live-cell imaging with fluorescently tagged TMEM218 to track its dynamics during ciliogenesis
-
Correlative light and electron microscopy to connect TMEM218 localization with ultrastructural features
-
Expansion microscopy to enhance visualization of the transition zone architecture
What animal models are valuable for studying TMEM218-related ciliopathies?
Several animal models could provide insights into TMEM218 function:
-
Mouse models with Tmem218 mutations or knockout, examining phenotypes relating to retina, kidney, and brain development
-
Rat models, where Tmem218 has been associated with Senior-Loken syndrome
-
Zebrafish models for high-throughput screening of phenotypes and potential therapeutics
-
C. elegans or Drosophila models for rapid genetic interaction studies
Cross-species comparison of TMEM218 function would help identify conserved mechanisms and distinguish them from species-specific roles, guiding translational research efforts.
How can transcriptomic analyses inform our understanding of cellular responses to TMEM218 dysfunction?
Transcriptomic approaches reveal how cells respond to ciliary dysfunction caused by TMEM218 mutations:
-
RNA-Seq analysis of patient cells versus controls to identify dysregulated pathways
-
Single-cell RNA-Seq to capture cell type-specific responses within affected tissues
-
Temporal transcriptomic profiles during development to identify critical windows when TMEM218 function is most essential
-
Integration with proteomic data to connect transcriptional changes with altered protein networks
Research shows that host cell gene expression signatures can account for 75% of the variability in recombinant protein yield , suggesting that transcriptomic profiles could similarly provide insights into how cells respond to and compensate for TMEM218 dysfunction.
