Correlation between ferroptosis and adriamycin resistance in breast cancer regulated by transferrin receptor and its molecular mechanism.

Breast cancer is the most prevalent malignant tumor in women. Adriamycin (ADR) is a primary chemotherapy drug, but resistance limits its effectiveness. Ferroptosis, a newly identified cell death mechanism, involves the transferrin receptor (TFRC), closely linked with tumor cells. This study aimed to explore TFRC and ferroptosis's role in breast cancer drug resistance. Bioinformatics analysis showed that TFRC was significantly downregulated in drug-resistant cell lines, and patients with low TFRC expression might demonstrate a poor chemotherapeutic response to standard treatment. High expression of TFRC was positively correlated with most of the ferroptosis-related driver genes. The research findings indicate that ferroptosis markers were higher in breast cancer tissues than in normal ones. In chemotherapy-sensitive cases, Ferrous ion (Fe2+ ) and malondialdehyde (MDA) levels were higher than in resistant cases (all p < .05). TFRC expression was higher in breast cancer than in normal tissue, especially in the sensitive group (all p < .05). Cytological experiments showed increased hydrogen peroxide (H2 O2 ) after ADR treatment in both sensitive and resistant cells, with varying MDA changes (all p < .05). Elevating TFRC increased Fe2+ and MDA in ADR-resistant cells, enhancing their sensitivity to ADR. However, TFRC upregulation combined with ADR increased proliferation and invasiveness in resistant cell lines (all p < .05). In conclusion, ADR resistance to breast cancer is related to the regulation of iron ion-mediated ferroptosis by TFRC. Upregulation of TFRC in ADR-resistant breast cancer cells activates ferroptosis and reverses ADR chemotherapy resistance of breast cancer.

Knockdown of circSlc8a1 inhibited the ferroptosis in the angiotensin II treated H9c2 cells via miR-673-5p/TFRC axis.

BACKGROUND: This study aimed to investigate the role of circSlc8a1 in cardiac hypertrophy (CH), a pathological change in various cardiovascular diseases. METHODS: An in vitro CH model was established using angiotensin II (AngII) treated H9c2 cells, followed by western blotting and RT-qPCR for detecting relative expressions. Cell viability and proliferation were analyzed using CCK-8 and EdU assays, while lactate dehydrogenase (LDH), reactive oxygen species (ROS), glutathione (GSH), and iron levels were determined using corresponding kits. Moreover, dual-luciferase reporter and RNA pull-down assays were performed to demonstrate whether miR-673-5p is bound to circSlc8a1 or transferrin receptor (TFRC). RESULTS: The results indicated that the expressions of circSlc8a1 and TFRC were increased, while miR-673-5p was decreased in the AngII treated H9c2 cells. The ferroptosis inhibitor treatment decreased the atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), and ß-major histocompatibility complex (ß-MHC) protein expressions, and circSlc8a1 expressions. Knocking down of circSlc8a1 inhibited promoted the cell viability and proliferation, increased the GSH content, glutathione peroxidase 4, and solute carrier family 7 member 11 protein expressions, and decreased the LDH, ROS, iron levels, and RAS protein expressions. The MiR-673-5p inhibitor antagonized the role of si-circSlc8a1, and the over-expressed TFRC reversed the miR-673-5p mimicking effects in AngII treated H9c2 cells. CONCLUSION: CircSlc8a1 promoted the ferroptosis in CH via regulating the miR-673-5p/TFRC axis.

LASS2 suppresses metastasis in multiple cancers by regulating the ferroptosis signalling pathway through interaction with TFRC.

BACKGROUND: As a key enzyme in ceramide synthesis, longevity assurance homologue 2 (LASS2) has been indicated to act as a tumour suppressor in a variety of cancers. Ferroptosis is involved in a variety of tumour processes; however, the role of LASS2 in regulating ferroptosis has yet to be explored. This article explores the potential underlying mechanisms involved. METHODS: Bioinformatics tools and immunohistochemical staining were used to evaluate LASS2 expression, and the results were analysed in relation to overall survival and clinical association in multiple cancers. Coimmunoprecipitation-coupled liquid chromatography-mass spectrometry (co-IP LC-MS) was performed to identify potential LASS2-interacting proteins in thyroid, breast, and liver cancer cell lines. Transcriptomics, proteomics and metabolomics analyses of multiple cancer cell types were performed using MS or LC-MS to further explore the underlying mechanisms involved. Among these tumour cells, the common LASS2 interaction partner transferrin receptor (TFRC) was analysed by protein-protein docking and validated by coimmunoprecipitation western blot, immunofluorescence, and proximity ligation assays. Then, we performed experiments in which tumour cells were treated with Fer-1 or erastin or left untreated, with or without inducing LASS2 overexpression, and assessed the molecular biological and cellular functions by corresponding analyses. RESULTS: Low LASS2 expression is correlated with adverse clinical characteristic and poor prognosis in patients with thyroid cancer, breast cancer or HCC. Multiomics analyses revealed significant changes in the ferroptosis signalling pathway, iron ion transport and iron homeostasis. Our in vitro experiments revealed that LASS2 overexpression regulated ferroptosis status in these tumour cells by affecting iron homeostasis, which in turn inhibited tumour migration, invasion and EMT. In addition, LASS2 overexpression reversed the changes in tumour cell metastasis induced by either Fer-1 or erastin. Mechanistically, LASS2 interacts directly with TFRC to regulate iron homeostasis in these tumour cells. CONCLUSIONS: In summary, our study reveals for the first time that LASS2 can inhibit tumour cell metastasis by interacting with TFRC to regulate iron metabolism and influence ferroptosis status in thyroid, breast, and liver cancer cells, these results suggest potential universal therapeutic targets for the treatment of these cancers.

circ-TFRC downregulation suppresses ovarian cancer progression via miR-615-3p/IGF2 axis regulation.

BACKGROUND: Ovarian cancer (OC) is a malignancy among female globally. Circular RNAs (circRNAs) are a family of circular endogenous RNAs generated from selective splicing, which take part in many traits. Former investigation suggested that circ-TFRC was abnormally expressed in breast cancer (BC). Further, the role of circ-TFRC to the progress of OC remains unclear. So, the aim of this study was to reveal the regulatory mechanism of circ-TFRC. METHODS: Our team made the luciferase reporter assay to validate circ-TFRC downstream target. Transwell migration assay, 5-ethynyl-20-deoxyuridine, and cell counting kit-8 were applied to investigate both proliferation and migration. In vivo tumorigenesis and metastasis assays were performed to investigate the circ-TFRC role in OC. RESULTS: The outputs elucidated that circ-TFRC expression incremented in OC cells and tissues. circ-TFRC downregulation inhibited OC cell proliferation as well as migration in in vivo and in vitro experiments. The luciferase results validated that miR-615-3p and IGF2 were circ-TFRC downstream targets. IGF2 overexpression or miR-615-3p inhibition reversed OC cell migration after circ-TFRC silencing. Also, IGF2 overexpression reversed OC cell migration and proliferation post miR-615-3p upregulation. CONCLUSION: Results demonstrate that circ-TFRC downregulation inhibits OC progression and metastasis via IGF2 expression regulation and miR-615-3psponging.

Elevated expression of receptors for EGF, PDGF, transferrin and folate within murine and human lupus nephritis kidneys.

OBJECTIVE: Systemic lupus erythematosus (SLE) is a chronic autoimmune disease where the body's immune system targets cells and tissue in numerous organs, including the kidneys. Lupus nephritis (LN) is a highly heterogeneous disease, and diagnosis is difficult because clinical manifestations vary widely among patients. Comprehensive proteomic studies reported recently in LN have identified several urinary proteins which are also cell-surface receptors. If indeed these receptor proteins are also hyper-expressed within the kidneys, ligands to these receptors may be useful for drug targeting. METHODS: scRNA sequence data analysis and immunohistochemistry were performed on LN kidneys for expression of four implicated receptors, EGFR, FOL2R2, PDGF-RB, and TFRC. RESULTS: In reported scRNA sequencing studies from 21 LN patients and 3 healthy control renal biopsies or renal-infiltrating immune cells from 24 LN biopsies, EGFR, FOLR2, PDGF-Rb, and TFRC were all hyper expressed within LN kidneys in comparison to healthy kidneys, either within resident renal cells or infiltrating leukocytes. Immunohistochemistry staining of murine lupus renal biopsies from lupus mice revealed EGFR, FOLR2, TFRC and PDGF-RB were elevated in LN kidneys. Immunohistochemistry staining of human Class II, Class III, and Class IV kidney tissue sections revealed EGFR, TFRC, and PDGF-RB were significantly elevated in proliferative LN kidneys. CONCLUSION: These findings underscore the potential of EGFR, TFRC, FOLR2, and PDGF-RB as promising receptors for potential drug-targeting in LN.

Knockdown of TFRC suppressed the progression of nasopharyngeal carcinoma by downregulating the PI3K/Akt/mTOR pathway.

BACKGROUND: The transferrin receptor (TfR) encoded by TFRC gene is the main cellular iron importer. TfR is highly expressed in many cancers and is expected to be a promising new target for cancer therapy; however, its role in nasopharyngeal carcinoma (NPC) remains unknown. METHODS: The TfR levels were investigated in NPC tissues and cell lines using immunohistochemistry and reverse transcription-quantitative polymerase chain reaction. Knockdown of TFRC using two siRNA to investigate the effects on intracellular iron level and biological functions, including proliferation by CKK-8 assay, colony formation, cell apoptosis and cell cycle by flow cytometry, migration and invasion, and tumor growth in vivo by nude mouse xenografts. RNA sequencing was performed to find possible mechanism after TFRC knockdown on NPC cells and further verified by western blotting. RESULTS: TfR was overexpressed in NPC cell lines and tissues. Knockdown of TFRC inhibited cell proliferation concomitant with increased apoptosis and cell cycle arrest, and it decreased intracellular iron, colony formation, migration, invasion, and epithelial-mesenchymal transition in HK1-EBV cells. Western blotting showed that TFRC knockdown suppressed the levels of the iron storage protein FTH1, anti-apoptotic marker BCL-xL, and epithelial-mesenchymal transition markers. We confirmed in vivo that TFRC knockdown also inhibited NPC tumor growth and decreased Ki67 expression in tumor tissues of nude mouse xenografts. RNA sequencing and western blotting revealed that TFRC silencing inhibited the PI3K/Akt/mTOR signaling pathway. CONCLUSIONS: These results indicated that TfR was overexpressed in NPC, and TFRC knockdown inhibited NPC progression by suppressing the PI3K/Akt/mTOR signaling pathway. Thus, TfR may serve as a novel biomarker and therapeutic target for NPC.

METTL14 promotes doxorubicin-induced cardiomyocyte ferroptosis by regulating the KCNQ1OT1-miR-7-5p-TFRC axis.

Doxorubicin (DOX) has toxic effects on the heart, causing cardiomyopathy and heart injury, but the underlying mechanisms of these effects require further investigation. This study investigated the role of DOX in promoting ferroptosis to induce myocardial injury. AC16 cardiomyocyte and neonatal rat ventricle cardiomyocytes were used as an in vitro model to study the molecules involved in myocardial injury using gene silencing, ectopic expression, and RNA immunoprecipitation. Messenger RNA and protein level analyses showed that DOX treatment resulted in the upregulation of methyltransferase-like 14 (METTL14), which catalyzes the m6A modification of the long non-coding RNA KCNQ1OT1, a miR-7-5p sponge. The RNA-binding protein IGF2BP1 is associated with KCNQ1OT1 to increase its stability and robustly inhibit miR-7-5p activity. Furthermore, a lack of miR-7-5p expression led to increased levels of transferrin receptor, promoting the uptake of iron and production of lipid reactive oxygen species and demonstrating that DOX-induced ferroptosis occurs in AC16 cells. Additionally, we found that miR-7-5p targets METTL14 in AC16 cells. Meanwhile, the role of METTL14/KCNQ1OT1/miR-7-5p axis in regulating ferroptosis in neonatal rat ventricle cardiomyocytes was also confirmed. Our results indicate that selectively inhibiting ferroptosis mediated by a METTL14/KCNQ1OT1/miR-7-5p positive feedback loop in cardiomyocytes could provide a new therapeutic approach to control DOX-induced cardiac injury.

Expression of genes related to iron homeostasis in breast cancer.

BACKGROUND: The dysfunctions in the metabolism of iron have an important role in many pathological conditions, ranging from disease with iron deposition to cancer. Studies on malignant diseases of the breast reported irregular expression in genes associated with iron metabolism. The variations are related to findings that have prognostic significance. This study evaluated the relationship of the expression levels of transferrin receptor 1 (TFRC), iron regulatory protein 1 (IRP1), hepcidin (HAMP), ferroportin 1 (FPN1), hemojuvelin (HFE2), matriptase 2 (TMPRSS6), and miR-122 genes in the normal and malignant tissues of breast cancer patients. METHODS & RESULTS: The normal and malignant tissues from 75 women with breast malignancies were used in this study. The patients did not receive any treatment previously. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was used in figuring the levels of gene expression associated with iron metabolism. When the malignant and normal tissues gene expression levels were analyzed, expression of TFRC increased (1.586-fold); IRP1 (0.594 fold) and miR-122 (0.320 fold) expression decreased; HAMP, FPN1, HFE2, and TMPRSS6 expressions did not change. FPN1 and IRP1 had a positive association, and this association was statistically significant (r = 0.266; p = 0.022). IRP1 and miR-122 had a positive association, and this association had statistical significance (r = 0.231; p = 0.048). CONCLUSIONS: Our study portrayed the important association between genes involved in iron hemostasis and breast malignancy. The results could be used to establish new diagnostic techniques in the management of breast malignancies. The alterations in the metabolism of malignant breast cells with normal breast cells could be utilized to achieve advantages in treatment.

A novel signature constructed by ferroptosis-associated genes (FAGs) for the prediction of prognosis in bladder urothelial carcinoma (BLCA) and associated with immune infiltration.

BACKGROUND: Ferroptosis, a novel form of regulated cell death, has been implicated in the pathogenesis of cancers. Nevertheless, the potential function and prognostic values of ferroptosis in bladder urothelial carcinoma (BLCA) are complex and remain to be clarified. Therefore, we proposed to systematically examine the roles of ferroptosis-associated genes (FAGs) in BLCA. METHODS: According to The Cancer Genome Atlas (TCGA) database, differently expressed FAGs (DEFAGs) and differently expressed transcription factors (DETFs) were identified in BLCA. Next, the network between DEFAGs and DETFs, GO annotations and KEGG pathway analyses were performed. Then, through univariate, LASSO and multivariate regression analyses, a novel signature based on FAGs was constructed. Moreover, survival analysis, PCA analysis, t-SNE analysis, ROC analysis, independent prognostic analysis, clinicopathological and immune correlation analysis, and experimental validation were utilized to evaluate the signature. RESULTS: Twenty-eight DEFAGs were identified, and four FAGs (CRYAB, TFRC, SQLE and G6PD) were finally utilized to establish the FAGs based signature in the TCGA cohort, which was subsequently validated in the GEO database. Moreover, we found that immune cell infiltration, immunotherapy-related biomarkers and immune-related pathways were significantly different between two risk groups. Besides, nine molecule drugs with the potential to treat bladder cancer were identified by the connectivity map database analysis. Finally, the expression levels of crucial FAGs were verified by the experiment, which were consistent with our bioinformatics analysis, and knockdown of TFRC could inhibit cell proliferation and colony formation in BLCA cell lines in vitro. CONCLUSIONS: Our study identified prognostic ferroptosis-associated genes and established a novel FAGs signature, which could accurately predict prognosis in BLCA patients.

Matriptase-2 and Hemojuvelin in Hepcidin Regulation: In Vivo Immunoblot Studies in Mask Mice.

Matriptase-2, a serine protease expressed in hepatocytes, is a negative regulator of hepcidin expression. The purpose of the study was to investigate the interaction of matriptase-2 with hemojuvelin protein in vivo. Mice lacking the matriptase-2 proteolytic activity (mask mice) display decreased content of hemojuvelin protein. Vice versa, the absence of hemojuvelin results in decreased liver content of matriptase-2, indicating that the two proteins interact. To further characterize the role of matriptase-2, we investigated iron metabolism in mask mice fed experimental diets. Administration of iron-enriched diet increased liver iron stores as well as hepcidin expression. Treatment of iron-overloaded mask mice with erythropoietin increased hemoglobin and hematocrit, indicating that the response to erythropoietin is intact in mask mice. Feeding of an iron-deficient diet to mask mice significantly increased spleen weight as well as the splenic content of erythroferrone and transferrin receptor proteins, indicating stress erythropoiesis. Liver hepcidin expression was decreased; expression of Id1 was not changed. Overall, the results suggest a complex interaction between matriptase-2 and hemojuvelin, and demonstrate that hepcidin can to some extent be regulated even in the absence of matriptase-2 proteolytic activity.

Using microarray analysis to identify genes and pathways that regulate fetal hemoglobin levels.

Increased levels of fetal hemoglobin (HbF: α2γ2) can ameliorate the clinical severity of the ß-hemoglobinopathies. Microarray analysis represents a powerful approach to identify novel genetic factors regulating the γ-globin gene. Gene expression profiling was previously performed on 14 individuals with high or normal HbF levels to identify the genetic factors that control γ-globin gene expression. To obtain more accurate and reliable results, our results were combined with public microarray dataset GSE22109 deposited in the Gene Expression Omnibus database. Annotation of case versus control samples was taken directly from the microarray documentation. The differentially expressed genes (DEGs) were obtained and were deeply analyzed by bioinformatics methods. Combined with our own chip expression data, potential genes HBE1, TFRC, and CSF2 were selected out for subsequent qRT-PCR validation. A total of 184 DEGs were identified from GSE22109 and the protein-protein interaction network was constructed. Gene set enrichment analysis showed that the hematopoietic cell lineage pathway overlaps in the two datasets. HBE1, CSF2, and TFRC were confirmed by qRT-PCR. Our results suggest novel candidate genes and pathways associated with the γ-globin gene expression.

Clinical and Immunological Characterization of Combined Immunodeficiency Due to TFRC Mutation in Eight Patients.

PURPOSE: Combined immunodeficiency (CID), due to mutations in TFRC gene that encodes the transferrin receptors (TfR1), is a rare monogenic disorder. In this study, we further characterize the clinical and immunological phenotypes in a cohort of eight patients. METHODS: A retrospective review of clinical and immunological features of patients diagnosed with a TFRC gene mutation between 2015 and 2019 in three tertiary centers. RESULTS: Eight patients from six unrelated families were enrolled. The patients had a median age of 7 years (4-32 years). All patients presented with recurrent sinopulmonary infections, chronic diarrhea, and failure to thrive in early life. Less common features were skin abscesses, conjunctivitis, global developmental delay, optic nerve atrophy, vitiligo, multinodular goiter, and hemophagocytic lymphohistiocytosis-like symptoms. All patients had intermittent neutropenia and 87% of the patients had recurrent thrombocytopenia. Anemia was found in 62%. All patients had hypogammaglobinemia and one had a persistent high IgM level. All patients had impaired function of T cells. The same homozygous missense mutation c.58T>C:p.Y20H, in the TFRC gene, was detected in all patients. Stem cell transplantation from matched donors was successful in two patients. Five patients did not receive stem cell transplantation, and they are on prophylactic treatment. One patient died due to severe sepsis and neurological complications. CONCLUSION: This report provides a large cohort with a long follow up of patients with this disease. Our cohort showed variable disease severity.

Circular RNA cTFRC acts as the sponge of MicroRNA-107 to promote bladder carcinoma progression.

BACKGROUND: Circular RNA (circRNA) represents a broad and diverse endogenous RNAs that can regulate gene expression in cancer. However, the regulation and function of bladder cancer (BC) circRNAs remain largely unknown. METHODS: Here we generated circRNA microarray data from three BC tissues and paired non-cancerous matched tissues, and detected circular RNA-cTFRC up-regulated and correlated with tumor grade and poor survival rate of BC patients. We subsequently performed functional analyses in cell lines and an animal model to support clinical findings. Mechanistically, we demonstrated that cTFRC could directly bind to miR-107 and relieve suppression for target TFRC expression. RESULTS: We detected circular RNA-cTFRC up-regulated and correlated with tumor grade and poor survival rate of BC patients. Knock down of cTFRC inhibited invasion and proliferation of BC cell lines in vitro and tumor growth in vivo. Furthermore, the expression of cTFRC correlated with TFRC and negatively correlated with miR-107 both in BC cell lines and BC clinical samples. In addition, up-regulation of cTFRC promoted TFRC expression and contributed to an epithelial to mesenchymal transition phenotype in BC cells. Finally, we found that cTFRC acts as a competing endogenous RNA (ceRNA) for miR-107 to regulate TFRC expression. CONCLUSIONS: cTFRC may exert regulatory functions in BC and may be a potential marker of BC diagnosis or progression.

Dermatoglyphic assessment in subjects with different dental arch forms: an appraisal.

Successful rehabilitation of edentulous individuals involves selection and arrangement of artificial teeth in accordance with the patient's original arch form. Various criteria exist for harmonious tooth arrangement but none is accepted universally. Finger and palm prints are unique to an individual and once formed in the sixth week of intra-uterine life, remain constant thereafter. Since dental arches are also formed during the same prenatal period, it is believed that the similar genetic factors may be involved in formation of dental arches and dermal patterns. This study was conducted to identify the association if any between type of dental arch forms and type of dermatoglyphic patterns. If specific dermal characteristics exist in individuals with specific dental arch forms, dermatoglyphic assessment of long standing edentulous subjects may help identify the patients preexisting dental arch form and thus aid in proper tooth arrangement. Ninety dentulous subjects were categorized into three groups on the basis of dental arch form (square, tapering or ovoid) and their finger and palm prints were recorded. The type of fingertip patterns, distribution of palmar patterns, Total Finger Ridge Count and angle atd were assessed. Subjects with square arches demonstrated a significantly high frequency of loops and a large atd angle with palmar patterns being most frequent in I3 region. Subjects with tapering arches showed a high frequency of whorls, a small atd angle and greatest distribution of palmar patterns in I4 region. In ovoid arched subjects, loops were the most common and palmar patterns were mostly observed in I4. Since distinctive dermal patterns were observed in subjects with different dental arch forms, it is believed that dermatoglyphics may be used as a reliable tool for identifying original arch form in edentulous patients.

METTL3 attenuates ferroptosis sensitivity in lung cancer via modulating TFRC.

Overexpression of methyltransferase-like 3 (METTL3) is significantly correlated with the malignancy of lung cancer (LC). In the present study, we demonstrated that METTL3 had higher levels in LC tissues relative to normal tissues. METTL3 showed superior sensitivity and specificity for diagnosis and identification of LC functions. In addition, silencing METTL3 resulted in enhanced ferroptosis sensitivity, whereas overexpression of METTL3 exhibited the opposite effect. Inhibition of METTL3 impeded LC growth in cell-derived xenografts. Further exploratory studies found that METTL3 stimulated the low expression of transferrin receptor (TFRC), which was critical for ferroptosis sensitization in LC cells induced by silenced METTL3, as silencing of TFRC caused a decrease in negative regulators of ferroptosis (FTH1 and FTL) in METTL3 knockdown A549 and PC9 cells. Finally, we confirmed that METTL3 attenuation effectively maintained the stability of TFRC mRNA. In conclusion, we reported a novel mechanism of METTL3 desensitization to ferroptosis via regulating TFRC, and an appropriate reduction of METTL3 might sensitize cancer cells to ferroptosis-based therapy.

Transferrin receptor levels and its rare variant are associated with human obesity.

AIM: Iron homeostasis is critical for functional respiratory chain complex of mitochondrial, thus potentially contributing to fat biology and energy homeostasis. Transferrin receptor (Tfrc) binds to transferrin for extracellular iron uptake and is recently reported to be involved in brown fat development and functionality. However, whether TFRC levels and variants are associated with human obesity is unknown. METHODS: To investigate the association of TFRC levels and variants with human obesity, fat biopsies were obtained from surgery. Exon-sequencing and genetic assessments were conducted of a case-control study. For TFRC levels assessment in fat biopsy, 9 overweight and 12 lean subjects were involved. For genetic study, obese (n = 1271) and lean subjects (n = 1455) were involved. TFRC levels were compared in abdominal mesenteric fat of pheochromocytoma patients versus control subjects, and overweight versus lean subjects. For genetic study, whole-exome sequencing of obese and matched control subjects were conducted and analyzed. In addition, the possible disruption in protein stability of TFRC variant was assessed by structural and molecular analysis. RESULTS: TFRC levels are increased in human browning adipose tissue and decreased in fat of overweight patients. Besides, TFRC levels are negatively correlated with body mass index and positively correlated with uncoupling protein 1 levels. Furthermore, a rare heterozygous missense variant p.I337V in TFRC shows a tendency to enrich in obese subjects. Structural and functional study reveals impaired protein stability of the TFRC variant compared to wild-type. CONCLUSIONS: Reduced TFRC levels and its rare variant p.I337V with protein instability are associated with human obesity.

Transferrin receptor modulated by microRNA-497-5p suppresses cervical cancer cell malignant phenotypes.

BACKGROUND: Cervical cancer is prevalent throughout the world, and microRNA-497-5p (miR-497-5p) plays an important role in its development. However, the specific mechanism by which miR-497-5p targets the transferrin receptor (TFRC) during cervical cancer development has not been clarified. OBJECTIVES: The aim of the study was to unravel TFRC expression and its role in cervical cancer cells, as well as the impact of the miR-497-5p/TFRC axis on cervical cancer cells. MATERIAL AND METHODS: The target mRNA was determined through differential analysis, followed by the evaluation of its impact on survival and clinical staging. Then, quantitative real-time polymerase chain reaction (qPCR) was conducted to analyze the TFRC mRNA level in cervical cancer cells and normal cervical epithelial cells. Western blot (WB) was utilized to examine the expression levels of TFRC, cleaved caspase-3, cleaved caspase-9, and epithelial-mesenchymal transition (EMT)-related proteins. The miRNAs upstream of the target mRNA were predicted, and Pearson correlation analysis was performed, followed by the validation through the dual-luciferase reporter assay. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and colony formation assays were performed to analyze cancer cell viability, followed by a transwell assay aimed at measuring cell migratory and invasive abilities. Finally, flow cytometry was conducted to examine cell apoptosis and cell cycle. RESULTS: The transferrin receptor was significantly increased in cervical cancer cells and positively associated with clinical T and N stages. Silencing TFRC could constrain cell proliferative, migratory and invasive abilities, arrest the cell cycle and facilitate cell apoptosis in cervical cancer cells. The bioinformatics analysis showed a significantly negative correlation between miR-497-5p and TFRC in cervical cancer. Moreover, upregulated miR-497-5p hampered cervical cancer progression and decreased TFRC expression. The overexpression of TFRC reversed the suppressive impact of miR-497-5p overexpression on cervical cancer progression. CONCLUSIONS: The modulatory role of the miR-497-5p/TFRC axis was confirmed in cervical cancer cells. This axis may present a new avenue for the diagnosis of cervical cancer and provide a novel target for cervical cancer treatment.

Expression of transferrin receptor/TFRC protein in bladder cancer cell T24 and its role in inducing iron death in bladder cancer.

Bladder cancer (BC) is a very common malignant tumor in the urinary system. However, the incidence rate, recurrence rate, progression rate and metastasis rate of bladder cancer are still very high, leading to poor long-term prognosis of patients. This study was to investigate the expression of transferrin receptor/TFRC protein in bladder cancer tissue and its role in inducing iron death of T24 human bladder cancer cells. Based on the intersection of 259 FerrDb genes in the iron death database with GSE13507 and GSE13167 data sets, 54 genes related to iron death in bladder cancer were obtained. Analyzing 54 genes, KEGG enrichment analysis showed that the pathways involved were mainly focused on iron death, autophagy, and tumor center carbon metabolism. GO analysis found that the molecular functions mainly gather in ubiquitin like protein ligase binding, ubiquitin protein ligase binding, and antioxidant activity. In the cellular components, it is mainly distributed in pigment granules, melanosomes, and the basal lateral plasma membrane. In biological processes, it is enriched in nutrient level responses, responses to extracellular stimuli, and cellular redox homeostasis. Screen out the top 10 core genes. The 10 core genes are SLC2A1, TFRC, EGFR, KRAS, CAV1, HSPA5, NFE2L2, VEGFA, PIK3CA, and HRAS. Finally, TFRC was selected as the research object. TCGA analysis showed that the expression level in bladder cancer tissue was higher than that in normal tissue, and the difference was statistically significant (P < 0.001). Conclusion (1) TFRC is highly expressed in many kinds of tumors, and it is more highly expressed in bladder cancer than in normal bladder tissue. (2) TFRC has certain diagnostic and prognostic value in bladder cancer. (3) Erastin, an iron death inducer, induced the iron death of T24 human bladder cancer cells, knocked down the expression of TFRC in T24 human bladder cancer cells, and preliminarily verified that silencing TFRC could inhibit the iron death of T24 human bladder cancer cells.

TRIM33 enhances the ubiquitination of TFRC to enhance the susceptibility of liver cancer cells to ferroptosis.

BACKGROUND: Hepatocellular carcinoma (HCC) is a common malignancy, and ferroptosis is a novel form of cell death driven by excessive lipid peroxidation. In recent years, ferroptosis has been widely utilized in cancer treatment, and the ubiquitination modification system has been recognized to play a crucial role in tumorigenesis and metastasis. Increasing evidence suggests that ubiquitin regulates ferroptosis-related substrates involved in this process. However, the precise mechanism of utilizing ubiquitination modification to regulate ferroptosis for HCC treatment remains unclear. METHODS: In this study, we detected the expression of TRIM33 in HCC using immunohistochemistry and western blotting techniques. The functional role of TRIM33 was verified through both in vitro and in vivo experiments. To evaluate the level of ferroptosis, mitochondrial superoxide levels, MDA levels, Fe2+ levels, and cell viability were assessed. Downstream substrates of TRIM33 were screened and confirmed via immunoprecipitation, immunofluorescence staining, and ubiquitination modification experiments. RESULTS: Our findings demonstrate that TRIM33 inhibits the growth and metastasis of HCC cells both in vitro and in vivo while promoting their susceptibility to ferroptosis. Mechanistically speaking, TRIM33 induces cellular ferroptosis through E3 ligase-dependent degradation of TFRC-a known inhibitor of this process-thus elucidating the specific type and site at which TFRC undergoes modification by TRIM33. CONCLUSION: In summary, our study reveals an important role for TRIM33 in HCC treatment while providing mechanistic support for its function. Additionally highlighted is the significance of ubiquitination modification leading to TFRC degradation-an insight that may prove valuable for future targeted therapies.

O-GlcNAcylation regulates the stability of transferrin receptor (TFRC) to control the ferroptosis in hepatocellular carcinoma cells.

Ferroptosis is an iron-dependent programmed cell death (PCD) enforced by lipid peroxidation accumulation. Transferrin receptor (TFRC), one of the signature proteins of ferroptosis, is abundantly expressed in hepatocellular carcinoma (HCC). However, post-translational modification (PTM) of TFRC and the underlying mechanisms for ferroptosis regulation remain less understood. In this study, we found that TFRC undergoes O-GlcNAcylation, influencing Erastin-induced ferroptosis sensitivity in hepatocytes. Further mechanistic studies found that Erastin can trigger de-O-GlcNAcylation of TFRC at serine 687 (Ser687), which diminishes the binding of ubiquitin E3 ligase membrane-associated RING-CH8 (MARCH8) and decreases polyubiquitination on lysine 665 (Lys665), thereby enhancing TFRC stability that favors labile iron accumulation. Therefore, our findings report O-GlcNAcylation on an important regulatory protein of ferroptosis and reveal an intriguing mechanism by which HCC ferroptosis is controlled by an iron metabolism pathway.