SEMA3C Human

What is the molecular structure and functional domains of human SEMA3C?

Human SEMA3C is a secreted protein with a highly modular structure. It contains a 20 amino acid signal sequence followed by an approximately 500 amino acid N-terminal Sema domain that forms a beta-propeller structure similar to integrins. The protein also features a cysteine knot, a furin-type cleavage site, an Ig-like domain, and a C-terminal basic domain . The functional activity of SEMA3C, like other class 3 semaphorins, requires both covalent dimerization and cleavage at the C-terminus . Human SEMA3C shares at least 95% amino acid identity with mouse, rat, bovine, and canine SEMA3C, highlighting its evolutionary conservation .

What are the primary receptors and signaling mechanisms of SEMA3C?

SEMA3C transduces signals through transmembrane plexins, either directly or by binding to associated neuropilin receptors . Specifically, SEMA3C signaling is primarily transduced by Plexin-D1 indirectly via Neuropilin 1 (NRP1) or Neuropilin 2 . Recent research has also identified integrin beta-1 (ITGB1) as another functional receptor for SEMA3C . Upon receptor binding, SEMA3C activates multiple downstream signaling pathways, including AKT/Gli1/c-Myc in cancer cells and NF-κB signaling in stromal cells . This receptor diversity allows SEMA3C to function as both a chemorepellent and chemoattractant depending on the cellular context .

What are the normal physiological functions of SEMA3C in human development?

SEMA3C plays crucial roles in several developmental processes. It is expressed in all somitic motor neurons, lung buds, and cardiac neural crest cells during development . In the cardiovascular system, SEMA3C is required for normal cardiovascular development during embryogenesis, guiding neural crest endothelial cell migration to the cardiac outflow tract by providing an attractive force that opposes Sema6A and Sema6B . Defects in aortic arch formation occur when SEMA3C gene expression or its interactions with neuropilins are disrupted . In the nervous system, SEMA3C functions as an axon attractant for migrating cortical axons and promotes axon growth in human dopaminergic neurons . This dual functionality as both attractant and repellent allows SEMA3C to create complex guidance patterns during development .

What recombinant SEMA3C protein variants are available for in vitro studies, and how should they be handled?

Researchers can utilize recombinant human Semaphorin 3C Fc chimera proteins for in vitro studies. These are available in carrier-free formulations, lyophilized from a 0.2 μm filtered solution in PBS . The carrier-free version lacks bovine serum albumin (BSA), making it suitable for applications where BSA might interfere with experimental outcomes . For reconstitution, these proteins should be prepared at 500 μg/mL in sterile PBS . Regarding stability and storage, it's recommended to use a manual defrost freezer and avoid repeated freeze-thaw cycles . When using recombinant SEMA3C in functional assays, research has shown that immobilizing the protein at 10 μg/mL, combined with 2 ng/mL rhFN-1, can induce approximately 50-75% adhesion in appropriate cell types .

What are the optimal methods for detecting and quantifying SEMA3C expression in biological samples?

For detecting and quantifying SEMA3C in human samples, sandwich ELISA provides a robust methodological approach. Commercial ELISA kits offer detection ranges of 0.78-50 ng/mL with sensitivity around 0.36 ng/mL . These assays are validated for various sample types including serum, plasma, tissue homogenates, and cell culture supernatants . For tissue analysis, immunohistochemistry using validated antibodies on tissue microarrays (TMAs) has been successfully employed to evaluate SEMA3C expression across different disease states, including treatment-resistant cancers . When constructing TMAs, researchers typically use triplicate cores of 0.6 mm for each sample to account for tissue heterogeneity . For gene expression analysis, antisense oligonucleotides (ASOs) have been effectively used to achieve in vivo SEMA3C gene silencing in experimental models .

What experimental models are most appropriate for studying SEMA3C functions in vivo?

Xenograft models have proven valuable for studying SEMA3C functions in cancer contexts. For instance, LNCaP xenografts in male athymic nude mice have been used to study SEMA3C's role in castration-resistant prostate cancer progression . In these models, serum prostate-specific antigen (PSA) levels of approximately 75 ng/ml have been used as thresholds for initiating treatment interventions . For SEMA3C inhibition studies, antisense oligonucleotides administered at 12.5 mg/kg every other day by intraperitoneal injection have demonstrated efficacy in modulating SEMA3C expression and function . To evaluate treatment effects, researchers typically monitor multiple parameters including tumor volume, serum biomarkers, and immunohistochemical analysis of tissues for markers such as SEMA3C expression, CD31 (angiogenesis), Ki67 (proliferation), and TUNEL (apoptosis) . When designing such experiments, randomization of treatment groups and matching of baseline parameters (tumor volume, biomarker levels) are essential for valid comparisons .

How does SEMA3C contribute to cancer stem cell maintenance and tumor progression?

SEMA3C has emerged as a significant regulator of cancer stemness properties and tumor progression across multiple cancer types. In hepatocellular carcinoma (HCC), SEMA3C is significantly upregulated in fibrotic liver and HCC tissues, as well as in peripheral blood of patients . This upregulation correlates with the acquisition of stemness properties in HCC cells . Mechanistically, SEMA3C activates downstream AKT/Gli1/c-Myc signaling pathways through NRP1 and ITGB1 receptors, which bolsters self-renewal capacity and tumor initiation potential . Beyond its direct effects on cancer cells, SEMA3C facilitates extracellular matrix remodeling through promoting contraction and collagen deposition . This dual action on both cancer cells and the surrounding stroma creates a microenvironment favorable for tumor progression. Additionally, research has demonstrated that SEMA3C is upregulated in sorafenib-resistant tissues and cells, suggesting its involvement in drug resistance mechanisms .

What is the role of SEMA3C in modulating the tumor microenvironment, particularly regarding cancer-associated fibroblasts?

SEMA3C plays a crucial role in orchestrating the tumor microenvironment through complex interactions with stromal cells, particularly cancer-associated fibroblasts (CAFs). In HCC, SEMA3C secreted by cancer cells promotes the proliferation and activation of hepatic stellate cells (HSCs), which are precursors to CAFs . Mechanistically, SEMA3C interacts with NRP1 and ITGB1 receptors on HSCs, activating downstream NF-κB signaling . This activation stimulates the release of IL-6 and upregulates HMGCR expression, which enhances cholesterol synthesis in HSCs . Importantly, this interaction creates a positive feedback loop, as CAF-secreted TGF-β1 activates AP1 signaling in cancer cells, which further augments SEMA3C expression . This reciprocal communication between cancer cells and stromal components accelerates tumor progression by creating a microenvironment conducive to cancer growth and invasion. This mechanism is particularly significant in HCC, where more than 90% of cases develop in the presence of fibrosis or cirrhosis, making the tumor microenvironment distinctive due to the intricate interplay between CAFs and cancer stem cells .

How is SEMA3C expression correlated with treatment resistance in prostate cancer?

SEMA3C expression has been significantly associated with treatment resistance in prostate cancer. Tissue microarray analysis of 280 prostate cancer specimens has revealed important correlations between SEMA3C expression and treatment response . Expression patterns differ markedly between untreated cases, cases treated with neoadjuvant hormone therapy (NHT), and cases treated with both NHT and docetaxel . These findings suggest that SEMA3C levels may serve as a biomarker for predicting treatment response and potentially indicate the development of resistance mechanisms. In experimental models, targeting SEMA3C with antisense oligonucleotides has demonstrated significant inhibition of castration-resistant prostate cancer (CRPC) tumor growth . In LNCaP xenograft models, SEMA3C inhibition post-castration resulted in sustained suppression of tumor volume and serum PSA levels compared to controls that exhibited standard kinetics of CRPC progression . Histological analysis of these tumors revealed decreased CD31 and Ki67 staining, with increased TUNEL staining, indicating reduced angiogenesis and proliferation with enhanced apoptosis . These findings collectively position SEMA3C as both a biomarker for treatment resistance and a potential therapeutic target for advanced prostate cancer.

How do the multiple receptor interactions of SEMA3C result in context-dependent cellular responses?

SEMA3C exhibits remarkable context-dependent cellular responses through its interactions with multiple receptor systems. This protein can function as both a chemorepellent and chemoattractant depending on the cellular context and receptor expression patterns . The molecular basis for this functional duality lies in SEMA3C's ability to engage different receptor complexes: it can signal through Plexin-D1 indirectly via either Neuropilin 1 or Neuropilin 2, and it can also activate integrin-mediated signaling . In certain cellular contexts, SEMA3C activates integrins, thereby functioning as a chemoattractant rather than a repellent . This chemoattraction appears to complement Sema3A repulsion in adjacent cell layers during neural development . The mechanistic complexities extend to downstream signaling pathways, where SEMA3C can activate distinct cascades including AKT/Gli1/c-Myc and NF-κB pathways depending on the cellular context and receptor engagement pattern . This signaling diversity allows SEMA3C to orchestrate complex biological processes such as axon guidance, where precise spatial control of attraction and repulsion is essential, and in tumor microenvironments, where it can simultaneously affect both cancer cells and stromal components .

What are the structural determinants of SEMA3C that regulate its binding specificity to different receptors?

The structural determinants of SEMA3C that regulate its receptor binding specificity are complex and involve multiple domains. The ~500 amino acid N-terminal Sema domain, which forms a beta-propeller structure similar to integrins, is critical for receptor recognition and binding . Specific mutations within the SEMA3C protein can alter its binding properties and functional outcomes. For instance, the recombinant SEMA3C variant with mutations at Arg551Ala, Arg552Ala, Arg611Ala, and Arg612Ala has been used in experimental studies, suggesting these residues may be important for functional activity . The cysteine knot domain facilitates dimerization, which is essential for the biological activity of class 3 semaphorins including SEMA3C . Additionally, the C-terminal basic domain and the furin-type cleavage site are important post-translational regulatory elements that influence SEMA3C's receptor binding and signaling capabilities . The high evolutionary conservation of SEMA3C across species (95% amino acid identity with mouse, rat, bovine, and canine orthologs) suggests that these structural features are functionally critical . Understanding these structural determinants is essential for developing targeted approaches to modulate SEMA3C functions in diverse biological contexts.

How does the proteolytic processing of SEMA3C influence its biological activities?

Proteolytic processing is a critical regulatory mechanism that influences SEMA3C's biological activities. Class 3 semaphorins, including SEMA3C, require cleavage at the C-terminus for full biological activity . The furin-type cleavage site in SEMA3C provides a specific recognition sequence for proteolytic processing . This processing, combined with covalent dimerization, is essential for SEMA3C to exert its guidance functions in developmental and pathological contexts . The proteolytic regulation creates an additional layer of control over SEMA3C activity in different tissue microenvironments, where protease expression may vary. In cancer contexts, altered protease expression patterns could potentially influence SEMA3C processing and thereby modify its effects on tumor progression and treatment response . The specific proteases involved in SEMA3C processing in different biological contexts, and how this processing affects receptor binding preferences and downstream signaling outcomes, represent important areas for future investigation. Understanding these regulatory mechanisms could potentially reveal new approaches for therapeutic modulation of SEMA3C activity in disease contexts.

What is the potential of SEMA3C as a therapeutic target for advanced cancers?

SEMA3C shows significant promise as a therapeutic target for advanced cancers, particularly in treatment-resistant contexts. In hepatocellular carcinoma, SEMA3C blockade effectively inhibits tumor growth and sensitizes cancer cells to sorafenib in vivo . Similarly, in prostate cancer models, targeting SEMA3C with antisense oligonucleotides (ASOs) demonstrates significant inhibition of castration-resistant tumor growth . When administered at 12.5 mg/kg every other day by intraperitoneal injection, SEMA3C ASO treatment prevented the progression to castration-resistant disease in LNCaP xenograft models over a 6-week treatment period . Importantly, no treatment-related adverse side effects (changes in body weight, gross morphology, or behavior) were observed with SEMA3C ASO treatment, suggesting a potentially favorable safety profile . The mechanism of action appears multifaceted, affecting not only cancer cell proliferation but also angiogenesis and the tumor microenvironment . The upregulation of SEMA3C in treatment-resistant contexts across multiple cancer types suggests that targeting this protein might overcome resistance mechanisms that limit current therapies . These findings collectively position SEMA3C as a promising therapeutic target worthy of further clinical development.

How can SEMA3C expression be used as a biomarker for disease progression or treatment stratification?

SEMA3C expression patterns offer considerable potential as biomarkers for disease progression and treatment stratification. Research has demonstrated significant upregulation of SEMA3C in multiple disease contexts, including fibrotic liver, HCC tissues, and peripheral blood of HCC patients . This upregulation correlates with the acquisition of stemness properties in cancer and is also observed in sorafenib-resistant tissues and cells . In prostate cancer, tissue microarray analysis has revealed associations between SEMA3C expression patterns and treatment responses, with different expression profiles observed in untreated cases versus those treated with various therapeutic regimens . For clinical application, multiple detection methods are validated for assessing SEMA3C levels in different sample types. Sandwich ELISA assays can quantify SEMA3C in serum, plasma, and other biological fluids with a detection range of 0.78-50 ng/mL and sensitivity of 0.36 ng/mL . For tissue analysis, immunohistochemistry protocols using validated antibodies can evaluate SEMA3C expression in biopsy or surgical specimens . These methodological approaches provide practical tools for incorporating SEMA3C assessment into clinical decision-making frameworks for personalized treatment strategies.