ABSTRACT
Alport syndrome is a hereditary kidney disease caused by collagen IV mutations that interfere with the formation and deposition of the α3α4α5 protomer into the glomerular basement membrane. In this issue, Yu et al. show that the chemical chaperone tauroursodeoxycholic acid prevented kidney structural changes and function decline in mice with a pathogenic missense Col4a3 mutation by increasing mutant α3α4α5 protomer glomerular basement membrane deposition and preventing podocyte apoptosis induced by endoplasmic reticulum stress.
Subject(s)
Autoantigens , Collagen Type IV , Glomerular Basement Membrane , Nephritis, Hereditary , Taurochenodeoxycholic Acid , Nephritis, Hereditary/genetics , Nephritis, Hereditary/drug therapy , Nephritis, Hereditary/pathology , Nephritis, Hereditary/metabolism , Animals , Collagen Type IV/genetics , Collagen Type IV/metabolism , Taurochenodeoxycholic Acid/pharmacology , Taurochenodeoxycholic Acid/therapeutic use , Mice , Glomerular Basement Membrane/pathology , Glomerular Basement Membrane/drug effects , Humans , Autoantigens/genetics , Autoantigens/metabolism , Endoplasmic Reticulum Stress/drug effects , Apoptosis/drug effects , Disease Models, Animal , Podocytes/drug effects , Podocytes/pathology , Podocytes/metabolism , Mutation, Missense , Molecular Chaperones/genetics , Molecular Chaperones/metabolismABSTRACT
Ectopic lymphoid structures (ELSs) in the rheumatoid synovial joints sustain autoreactivity against locally expressed autoantigens. We recently identified recombinant monoclonal antibodies (RA-rmAbs) derived from single, locally differentiated rheumatoid arthritis (RA) synovial B cells, which specifically recognize fibroblast-like synoviocytes (FLSs). Here, we aimed to identify the specificity of FLS-derived autoantigens fueling local autoimmunity and the functional role of anti-FLS antibodies in promoting chronic inflammation. A subset of anti-FLS RA-rmAbs reacting with a 60 kDa band from FLS extracts demonstrated specificity for HSP60 and partial cross-reactivity to other stromal autoantigens (i.e., calreticulin/vimentin) but not to citrullinated fibrinogen. Anti-FLS RA-rmAbs, but not anti-neutrophil extracellular traps rmAbs, exhibited pathogenic properties in a mouse model of collagen-induced arthritis. In patients, anti-HSP60 antibodies were preferentially detected in RA versus osteoarthritis (OA) synovial fluid. Synovial HSPD1 and CALR gene expression analyzed using bulk RNA-Seq and GeoMx-DSP closely correlated with the lympho-myeloid RA pathotype, and HSP60 protein expression was predominantly observed around ELS. Moreover, we observed a significant reduction in synovial HSP60 gene expression followed B cell depletion with rituximab that was strongly associated with the treatment response. Overall, we report that synovial stromal-derived autoantigens are targeted by pathogenic autoantibodies and are associated with specific RA pathotypes, with potential value for patient stratification and as predictors of the response to B cell-depleting therapies.
Subject(s)
Arthritis, Rheumatoid , Autoantigens , Chaperonin 60 , Germinal Center , Arthritis, Rheumatoid/immunology , Arthritis, Rheumatoid/pathology , Animals , Humans , Mice , Autoantigens/immunology , Autoantigens/genetics , Germinal Center/immunology , Germinal Center/pathology , Chaperonin 60/immunology , Chaperonin 60/genetics , Autoantibodies/immunology , Autoimmunity , Male , Synoviocytes/immunology , Synoviocytes/pathology , Synoviocytes/metabolism , Arthritis, Experimental/immunology , Arthritis, Experimental/pathology , Female , B-Lymphocytes/immunology , B-Lymphocytes/pathology , Tertiary Lymphoid Structures/immunology , Tertiary Lymphoid Structures/pathologyABSTRACT
Purpose: The purpose of this study was to investigate the ocular morphological characteristics of Col4a3-/- mice as a model of Alport syndrome (AS) and the potential pathogenesis. Methods: The expression of collagen IV at 8, 12, and 21 weeks of age was evaluated by immunohistochemistry in wild-type (WT) and Col4a3-/- mice. Hematoxylin and eosin (H&E) staining and thickness measurements were performed to assess the thickness of anterior lens capsule and retina. Ultrastructure analysis of corneal epithelial basement membrane, anterior lens capsule, internal limiting membrane (ILM), and retinal pigment epithelium (RPE) basement membrane was performed using transmission electron microscopy. Finally, Müller cell activation was evaluated by glial fibrillary acidic protein (GFAP) expression. Results: Collagen IV was downregulated in the corneal epithelial basement membrane and ILM of Col4a3-/- mice. The hemidesmosomes of Col4a3-/- mice corneal epithelium became flat and less electron-dense than those of the WT group. Compared with those of the WT mice, the anterior lens capsules of Col4a3-/- mice were thinner. Abnormal structure was detected at the ILM Col4a3-/- mice, and the basal folds of the RPE basement membrane in Col4a3-/- mice were thicker and shorter. The retinas of Col4a3-/- mice were thinner than those of WT mice, especially within 1000 µm away from the optic nerve. GFAP expression enhanced in each age group of Col4a3-/- mice. Conclusions: Our results suggested that Col4a3-/- mice exhibit ocular anomalies similar to patients with AS. Additionally, Müller cells may be involved in AS retinal anomalies. Translational Relevance: This animal model could provide an opportunity to understand the underlying mechanisms of AS ocular disorders and to investigate potential new treatments.
Subject(s)
Basement Membrane , Collagen Type IV , Disease Models, Animal , Mice, Knockout , Nephritis, Hereditary , Animals , Nephritis, Hereditary/pathology , Nephritis, Hereditary/genetics , Nephritis, Hereditary/metabolism , Collagen Type IV/genetics , Collagen Type IV/metabolism , Collagen Type IV/deficiency , Mice , Basement Membrane/metabolism , Basement Membrane/pathology , Basement Membrane/ultrastructure , Retinal Pigment Epithelium/pathology , Retinal Pigment Epithelium/metabolism , Retinal Pigment Epithelium/ultrastructure , Microscopy, Electron, Transmission , Mice, Inbred C57BL , Lens Capsule, Crystalline/metabolism , Lens Capsule, Crystalline/pathology , Lens Capsule, Crystalline/ultrastructure , Epithelium, Corneal/pathology , Epithelium, Corneal/ultrastructure , Epithelium, Corneal/metabolism , Glial Fibrillary Acidic Protein/metabolism , Glial Fibrillary Acidic Protein/genetics , Retina/pathology , Retina/metabolism , Retina/ultrastructure , Autoantigens/genetics , Autoantigens/metabolism , Ependymoglial Cells/pathology , Ependymoglial Cells/metabolism , Ependymoglial Cells/ultrastructure , Immunohistochemistry , MaleABSTRACT
Background and Objectives: Congenital thyroid dyshormonogenesis is caused by alterations in the synthesis of thyroid hormones in a newborn. Additionally, 10 to 20% of these cases are hereditary, caused by defects in proteins involved in hormonal synthesis. One of the most common causes is mutations in the thyroid peroxidase (TPO) enzyme gene, an autosomal recessive disease. We aimed to detect mutations of the TPO gene in 12 Chilean patients with congenital hypothyroidism due to dyshormonogenesis (CHD) and to characterize these patients clinically and molecularly. Materials and Methods: Twelve patients under 20 years of age with CHD, controlled at San Juan de Dios Hospital in Santiago, Chile, were selected according to the inclusion criteria: elevated neonatal TSH, persistent hypothyroidism, and thyroid normotopic by imaging study. Those with deafness, Down syndrome, and central or transient congenital hypothyroidism were excluded. Blood samples were taken for DNA extraction, and the 17 exons and exon-intron junctions of the TPO gene were amplified by PCR. The PCR products were sequenced by Sanger. Results: Two possibly pathogenic mutations of the TPO gene were detected: c.2242G>A (p.Val748Met) and c.1103C>T (p.Pro368Leu). These mutations were detected in 2 of 12 patients (16.6%): 1 was compound heterozygous c.1103C>T/c.2242G>A, and the other was heterozygous for c.2242G>A. In the diagnostic confirmation test, both patients presented diffuse hyper-uptake goiter on thyroid scintigraphy and high TSH in venous blood (>190 uIU/mL). Conclusions: The frequency of patients with possibly pathogenic mutations in TPO with CHD was 16.6%. Its study would allow for genetic counseling to be offered to the families of affected patients.
Subject(s)
Congenital Hypothyroidism , Iodide Peroxidase , Iron-Binding Proteins , Mutation , Humans , Congenital Hypothyroidism/genetics , Congenital Hypothyroidism/blood , Chile , Iodide Peroxidase/genetics , Female , Male , Iron-Binding Proteins/genetics , Autoantigens/genetics , Infant , Child , Adolescent , Child, Preschool , Infant, Newborn , Thyroid Dysgenesis/genetics , Thyroid Dysgenesis/complications , Thyroid Dysgenesis/bloodABSTRACT
Human telomerase assembly is a highly dynamic process. Using biochemical approaches, we find that LARP3 and LARP7/MePCE are involved in the early stage of human telomerase RNA (hTR) and that their binding to RNA is destabilized when the mature form is produced. LARP3 plays a negative role in preventing the processing of the 3'-extended long (exL) form and the binding of LARP7 and MePCE. Interestingly, the tertiary structure of the exL form prevents LARP3 binding and facilitates hTR biogenesis. Furthermore, low levels of LARP3 promote hTR maturation, increase telomerase activity, and elongate telomeres. LARP7 and MePCE depletion inhibits the conversion of the 3'-extended short (exS) form into mature hTR and the cytoplasmic accumulation of hTR, resulting in telomere shortening. Taken together our data suggest that LARP3 and LARP7/MePCE mediate the processing of hTR precursors and regulate the production of functional telomerase.
Subject(s)
Autoantigens , RNA , Ribonucleoproteins , SS-B Antigen , Telomerase , Humans , Autoantigens/metabolism , Autoantigens/genetics , HeLa Cells , Protein Binding , Ribonucleoproteins/metabolism , Ribonucleoproteins/genetics , RNA/metabolism , RNA/genetics , Telomerase/metabolism , Telomerase/genetics , Telomere/metabolism , Telomere/genetics , Telomere ShorteningABSTRACT
Bullous pemphigoid (BP) is a type 2 inflammation- and immunity-driven skin disease, yet a comprehensive understanding of the immune landscape, particularly immune-stromal crosstalk in BP, remains elusive. Herein, using single-cell RNA sequencing (scRNA-seq) and in vitro functional analyzes, we pinpoint Th2 cells, dendritic cells (DCs), and fibroblasts as crucial cell populations. The IL13-IL13RA1 ligand-receptor pair is identified as the most significant mediator of immune-stromal crosstalk in BP. Notably, fibroblasts and DCs expressing IL13RA1 respond to IL13-secreting Th2 cells, thereby amplifying Th2 cell-mediated cascade responses, which occurs through the specific upregulation of PLA2G2A in fibroblasts and CCL17 in myeloid cells, creating a positive feedback loop integral to immune-stromal crosstalk. Furthermore, PLA2G2A and CCL17 contribute to an increased titer of pathogenic anti-BP180-NC16A autoantibodies in BP patients. Our work provides a comprehensive insight into BP pathogenesis and shows a mechanism governing immune-stromal interactions, providing potential avenues for future therapeutic research.
Subject(s)
Chemokine CCL17 , Dendritic Cells , Fibroblasts , Pemphigoid, Bullous , Single-Cell Analysis , Th2 Cells , Humans , Pemphigoid, Bullous/immunology , Pemphigoid, Bullous/genetics , Single-Cell Analysis/methods , Fibroblasts/metabolism , Fibroblasts/immunology , Dendritic Cells/immunology , Dendritic Cells/metabolism , Chemokine CCL17/genetics , Chemokine CCL17/metabolism , Th2 Cells/immunology , Autoantibodies/immunology , Transcriptome , Interleukin-13/metabolism , Interleukin-13/genetics , Interleukin-13/immunology , Non-Fibrillar Collagens/immunology , Non-Fibrillar Collagens/genetics , Non-Fibrillar Collagens/metabolism , Inflammation/immunology , Inflammation/genetics , Inflammation/metabolism , Gene Expression Profiling/methods , Male , Female , Autoantigens/immunology , Autoantigens/metabolism , Autoantigens/genetics , Collagen Type XVII , Myeloid Cells/metabolism , Myeloid Cells/immunology , Stromal Cells/metabolism , Stromal Cells/immunologyABSTRACT
La-related proteins (LARPs) are a family of RNA-binding proteins that share a conserved La motif (LaM) domain. LARP1 plays a role in regulating ribosomal protein synthesis and stabilizing mRNAs and has a unique structure without an RNA binding RRM domain adjoining the LaM domain. In this study, we investigated the physical basis for LARP1 specificity for poly(A) sequences and observed an unexpected bias for sequences with single guanines. Multiple guanine substitutions did not increase the affinity, demonstrating preferential recognition of singly guanylated sequences. We also observed that the cyclic di-nucleotides in the cCAS/STING pathway, cyclic-di-GMP and 3',3'-cGAMP, bound with sub-micromolar affinity. Isothermal titration measurements were complemented by high-resolution crystal structures of the LARP1 LaM with six different RNA ligands, including two stereoisomers of a phosphorothioate linkage. The selectivity for singly substituted poly(A) sequences suggests LARP1 may play a role in the stabilizing effect of poly(A) tail guanylation. [Figure: see text].
Subject(s)
Poly A , Protein Binding , Ribonucleoproteins , SS-B Antigen , Ribonucleoproteins/metabolism , Ribonucleoproteins/chemistry , Ribonucleoproteins/genetics , Poly A/metabolism , Poly A/chemistry , Humans , Models, Molecular , Binding Sites , Autoantigens/metabolism , Autoantigens/chemistry , Autoantigens/genetics , Crystallography, X-Ray , Protein Domains , Cyclic GMP/metabolism , Cyclic GMP/analogs & derivatives , Cyclic GMP/chemistry , RNA, Messenger/metabolism , RNA, Messenger/chemistry , RNA, Messenger/geneticsABSTRACT
The mammalian Golgi is composed of stacks that are laterally connected into a continuous ribbon-like structure. The integrity and function of the ribbon is disrupted under stress conditions, but the molecular mechanisms remain unclear. Here we show that the ribbon is maintained by biomolecular condensates of RNA and the Golgi matrix protein GM130 (GOLGA2). We identify GM130 as a membrane-bound RNA-binding protein, which directly recruits RNA and associated RNA-binding proteins to the Golgi membrane. Acute degradation of RNA or GM130 in cells disrupts the ribbon. Under stress conditions, RNA dissociates from GM130 and the ribbon is disjointed, but after the cells recover from stress the ribbon is restored. When overexpressed in cells, GM130 forms RNA-dependent liquid-like condensates. GM130 contains an intrinsically disordered domain at its amino terminus, which binds RNA to induce liquid-liquid phase separation. These co-condensates are sufficient to link purified Golgi membranes, reconstructing lateral linking of stacks into a ribbon-like structure. Together, these studies show that RNA acts as a structural biopolymer that together with GM130 maintains the integrity of the Golgi ribbon.
Subject(s)
Autoantigens , Golgi Apparatus , Membrane Proteins , RNA , Golgi Apparatus/metabolism , Humans , Autoantigens/metabolism , Autoantigens/genetics , Autoantigens/chemistry , Membrane Proteins/metabolism , Membrane Proteins/genetics , Membrane Proteins/chemistry , RNA/metabolism , RNA/genetics , RNA-Binding Proteins/metabolism , RNA-Binding Proteins/genetics , RNA-Binding Proteins/chemistry , HeLa Cells , Biomolecular Condensates/metabolism , Protein Binding , Intracellular Membranes/metabolism , Animals , HEK293 CellsABSTRACT
BACKGROUND: Alport syndrome (AS) is an inherited nephropathy caused by mutations in the type IV collagen genes. It is clinically characterized by damage to the eyes, ears and kidneys. Diagnosis of AS is hampered by its atypical clinical picture, particularly when the typical features, include persistent hematuria and microscopic changes in the glomerular basement membrane (GBM), are the only clinical manifestations in the patient. METHODS: We screened 10 families with suspected AS using whole exome sequencing (WES) and analyzed the harmfulness, conservation, and protein structure changes of mutated genes. In further, we performed in vitro functional analysis of two missense mutations in the COL4A5 gene (c.2359G > C, p.G787R and c.2605G > A, p.G869R). RESULTS: We identified 11 pathogenic variants in the type IV collagen genes (COL4A3, COL4A4 and COL4A5). These pathogenic variants include eight missense mutations, two nonsense mutations and one frameshift mutation. Notably, Family 2 had digenic mutations in the COL4A3 (p.G1170A) and UMOD genes (p.M229K). Family 3 had a digenic missense mutation (p.G997E) in COL4A3 and a frameshift mutation (p.P502L fs*151) in COL4A4. To our knowledge, four of the 11 mutations are novel mutations. In addition, we found that COL4A5 mutation relation mRNA levels were significantly decreased in HEK 293 T cell compared to control, while the cellular localization remained the same. CONCLUSIONS: Our research expands the spectrum of COL4A3-5 pathogenic variants, which is helpful for clinical and scientific research.
Subject(s)
Autoantigens , Collagen Type IV , Nephritis, Hereditary , Pedigree , Humans , Nephritis, Hereditary/genetics , Nephritis, Hereditary/pathology , Collagen Type IV/genetics , Autoantigens/genetics , Female , Male , Adult , Mutation , Exome Sequencing , Middle Aged , Mutation, Missense , HEK293 CellsABSTRACT
Cellular communication (CC) influences tumor development by mediating intercellular junctions between cells. However, the role and underlying mechanisms of CC in malignant transformation remain unknown. Here, we investigated the spatiotemporal heterogeneity of CC molecular expression during malignant transformation. It was found that although both tight junctions (TJs) and gap junctions (GJs) were involved in maintaining the tumor microenvironment (TME), they exhibited opposite characteristics. Mechanistically, for epithelial cells (parenchymal component), the expression of TJ molecules consistently decreased during normal-cancer transformation and is a potential oncogenic factor. For fibroblasts (mesenchymal component), the expression of GJs consistently increased during normal-cancer transformation and is a potential oncogenic factor. In addition, the molecular profiles of TJs and GJs were used to stratify colorectal cancer (CRC) patients, where subtypes characterized by high GJ levels and low TJ levels exhibited enhanced mesenchymal signals. Importantly, we propose that leiomodin 1 (LMOD1) is biphasic, with features of both TJs and GJs. LMOD1 not only promotes the activation of cancer-associated fibroblasts (CAFs) but also inhibits the Epithelial-mesenchymal transition (EMT) program in cancer cells. In conclusion, these findings demonstrate the molecular heterogeneity of CC and provide new insights into further understanding of TME heterogeneity.
Subject(s)
Cancer-Associated Fibroblasts , Cell Communication , Colorectal Neoplasms , Epithelial-Mesenchymal Transition , Gene Expression Regulation, Neoplastic , Tumor Microenvironment , Animals , Humans , Cancer-Associated Fibroblasts/metabolism , Cancer-Associated Fibroblasts/pathology , Cell Line, Tumor , Colorectal Neoplasms/pathology , Colorectal Neoplasms/metabolism , Colorectal Neoplasms/genetics , Epithelial-Mesenchymal Transition/genetics , Gap Junctions/metabolism , Membrane Proteins/metabolism , Membrane Proteins/genetics , Spatio-Temporal Analysis , Tight Junctions/metabolism , Cytoskeletal Proteins/genetics , Cytoskeletal Proteins/metabolism , Autoantigens/genetics , Autoantigens/metabolismABSTRACT
Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer and it lacks specific therapeutic targets and effective treatment protocols. By analyzing a proteomic TNBC dataset, we found significant upregulation of sideroflexin 1 (SFXN1) in tumor tissues. However, the precise function of SFXN1 in TNBC remains unclear. Immunoblotting was performed to determine SFXN1 expression levels. Label-free quantitative proteomics and liquid chromatography-tandem mass spectrometry were used to identify the downstream targets of SFXN1. Mechanistic studies of SFXN1 and cellular inhibitor of PP2A (CIP2A) were performed using immunoblotting, immunofluorescence staining, and reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Functional experiments were used to investigate the role of SFXN1 in TNBC cells. SFXN1 was significantly overexpressed in TNBC tumor tissues and was associated with unfavorable outcomes in patients with TNBC. Functional experiments demonstrated that SFXN1 promoted TNBC growth and metastasis in vitro and in vivo. Mechanistic studies revealed that SFXN1 promoted TNBC progression by inhibiting the autophagy receptor TOLLIP (toll interacting protein)-mediated autophagic degradation of CIP2A. The pro-tumorigenic effect of SFXN1 overexpression was partially prevented by lapatinib-mediated inhibition of the CIP2A/PP2A/p-AKT pathway. These findings may provide a new targeted therapy for patients with TNBC.
Subject(s)
Autoantigens , Autophagy , Cation Transport Proteins , Lapatinib , Membrane Proteins , Triple Negative Breast Neoplasms , Animals , Female , Humans , Mice , Antineoplastic Agents/pharmacology , Autoantigens/metabolism , Autoantigens/genetics , Autophagy/drug effects , Cell Line, Tumor , Cell Proliferation/drug effects , Disease Progression , Gene Expression Regulation, Neoplastic/drug effects , Intracellular Signaling Peptides and Proteins/metabolism , Intracellular Signaling Peptides and Proteins/genetics , Lapatinib/pharmacology , Membrane Proteins/metabolism , Membrane Proteins/genetics , Proteolysis/drug effects , Signal Transduction/drug effects , Triple Negative Breast Neoplasms/drug therapy , Triple Negative Breast Neoplasms/pathology , Triple Negative Breast Neoplasms/metabolism , Triple Negative Breast Neoplasms/genetics , Xenograft Model Antitumor Assays , Cation Transport Proteins/genetics , Cation Transport Proteins/metabolismABSTRACT
Alport syndrome and autosomal dominant polycystic kidney disease are monogenic causes of chronic kidney disease and end-stage kidney failure. We present a case of a man in his 60s with progressive chronic kidney disease, bilateral sensorineural hearing loss and multiple renal cysts. Genetic analysis revealed a heterozygous variant in COL4A3 (linked to Alport syndrome) and in the GANAB gene (associated with a milder form of autosomal dominant polycystic kidney disease). Although each variant confers a mild risk of developing end-stage kidney disease, the patient presented a pronounced and accelerated progression of chronic kidney disease, which goes beyond what would be predicted by adding up their individual effects. This suggests a potential synergic effect of both variants, which warrants further investigation.
Subject(s)
Collagen Type IV , Nephritis, Hereditary , Polycystic Kidney, Autosomal Dominant , Humans , Nephritis, Hereditary/genetics , Nephritis, Hereditary/complications , Nephritis, Hereditary/diagnosis , Male , Polycystic Kidney, Autosomal Dominant/genetics , Polycystic Kidney, Autosomal Dominant/complications , Collagen Type IV/genetics , Middle Aged , Autoantigens/genetics , Disease Progression , Kidney Failure, Chronic/genetics , Kidney Failure, Chronic/etiology , Hearing Loss, Sensorineural/genetics , Hearing Loss, Sensorineural/diagnosisABSTRACT
BACKGROUND: Early embryonic arrest and fragmentation (EEAF) is a common cause of female infertility, but the genetic causes remain to be largely unknown. CIP2A encodes the cellular inhibitor of PP2A, playing a crucial role in mitosis and mouse oocyte meiosis. METHODS: Exome sequencing and Sanger sequencing were performed to identify candidate causative genes in patients with EEAF. The pathogenicity of the CIP2A variant was assessed and confirmed in cultured cell lines and human oocytes through Western blotting, semi-quantitative RT-PCR, TUNEL staining, and fluorescence localization analysis. FINDINGS: We identified CIP2A (c.1510C > T, p.L504F) as a novel disease-causing gene in human EEAF from a consanguineous family. L504 is highly conserved throughout evolution. The CIP2A variant (c.1510C > T, p.L504F) reduced the expression level of the mutant CIP2A protein, leading to the abnormal aggregation of mutant CIP2A protein and cell apoptosis. Abnormal aggregation of CIP2A protein and chromosomal dispersion occurred in the patient's oocytes and early embryos. We further replicated the patient phenotype by knockdown CIP2A in human oocytes. Additionally, CIP2A deficiency resulted in decreased levels of phosphorylated ERK1/2. INTERPRETATION: We first found that the CIP2A loss-of-function variant associate with female infertility characterized by EEAF. Our findings suggest the uniqueness and importance of CIP2A gene in human oocyte and early embryo development. FUNDING: This work was supported by National Key Research and Development Program of China (2023YFC2706302), the National Natural Science Foundation of China (81000079, 81170165, and 81870959), the HUST Academic Frontier Youth Team (2016QYTD02), and the Key Research of Huazhong University of Science and Technology, Tongji Hospital (2022A20).
Subject(s)
Autoantigens , Infertility, Female , Intracellular Signaling Peptides and Proteins , Membrane Proteins , Oocytes , Humans , Female , Autoantigens/genetics , Autoantigens/metabolism , Infertility, Female/genetics , Infertility, Female/pathology , Infertility, Female/metabolism , Membrane Proteins/genetics , Membrane Proteins/metabolism , Oocytes/metabolism , Intracellular Signaling Peptides and Proteins/genetics , Intracellular Signaling Peptides and Proteins/metabolism , Apoptosis/genetics , Loss of Function Mutation , Adult , Exome Sequencing , Animals , Pedigree , MiceABSTRACT
OBJECTIVE: To diagnose and explore the genetic etiology of a neonate with Hereditary epidermolysis bullosa. METHODS: A neonate who was admitted to Suqian Hospital Affiliated to Xuzhou Medical University on July 10, 2021 was selected as the study subject. Peripheral blood samples were collected from the child and his parents for the extraction of genomic DNA. And target gene capture and next-generation sequencing were carried out. Candidate variants were verified by Sanger sequencing and pathogenicity analysis. RESULTS: The child was found to harbor compound heterozygous variants of the COL17A1 gene, namely c.997C>T (p.Q333X) and c.3481dupT (p.Y1161fs*2), which were respectively inherited from his father and mother. Both variants were predicted to be pathogenic. CONCLUSION: The child was diagnosed with Generalized atrophic benign epidermolysis bullosa due to the compound heterozygous variants of the COL17A1 gene.
Subject(s)
Collagen Type XVII , Non-Fibrillar Collagens , Humans , Male , Infant, Newborn , Non-Fibrillar Collagens/genetics , Autoantigens/genetics , Mutation , Heterozygote , Epidermolysis Bullosa/genetics , FemaleABSTRACT
COL4A3/A4/A5 mutations have been identified as critical causes of Alport syndrome and other genetic chronic kidney diseases. However, the underlying pathogenesis remains unclear, and specific treatments are lacking. Here, we constructed a transgenic Alport syndrome mouse model by generating a mutation (Col4a3 p.G799R) identified previously from one large Alport syndrome family into mice. We observed that the mutation caused a pathological decrease in intracellular and secreted collagen IV α3α4α5 heterotrimers. The mutant collagen IV α3 chains abnormally accumulated in the endoplasmic reticulum and exhibited defective secretion, leading to persistent endoplasmic reticulum stress in vivo and in vitro. RNA-seq analysis revealed that the MyD88/p38 MAPK pathway plays key roles in mediating subsequent inflammation and apoptosis signaling activation. Treatment with tauroursodeoxycholic acid, a chemical chaperone drug that functions as an endoplasmic reticulum stress inhibitor, effectively suppressed endoplasmic reticulum stress, promoted secretion of the α3 chains, and inhibited the activation of the MyD88/p38 MAPK pathway. Tauroursodeoxycholic acid treatment significantly improved kidney function in vivo. These results partly clarified the pathogenesis of kidney injuries associated with Alport syndrome, especially in glomeruli, and suggested that tauroursodeoxycholic acid might be useful for the early clinical treatment of Alport syndrome.
Subject(s)
Apoptosis , Autoantigens , Collagen Type IV , Disease Models, Animal , Endoplasmic Reticulum Stress , Mice, Transgenic , Mutation , Nephritis, Hereditary , Taurochenodeoxycholic Acid , p38 Mitogen-Activated Protein Kinases , Animals , Taurochenodeoxycholic Acid/pharmacology , Taurochenodeoxycholic Acid/therapeutic use , Endoplasmic Reticulum Stress/drug effects , Nephritis, Hereditary/genetics , Nephritis, Hereditary/drug therapy , Nephritis, Hereditary/pathology , Collagen Type IV/genetics , Collagen Type IV/metabolism , p38 Mitogen-Activated Protein Kinases/metabolism , Autoantigens/genetics , Autoantigens/metabolism , Apoptosis/drug effects , Myeloid Differentiation Factor 88/genetics , Myeloid Differentiation Factor 88/metabolism , Humans , Signal Transduction/drug effects , Mice , Mice, Inbred C57BL , Male , Kidney/pathology , Kidney/drug effects , Kidney/metabolismABSTRACT
Motile cilia on the cell surface produce fluid flows in the body and abnormalities in motile cilia cause primary ciliary dyskinesia. Dynein axonemal assembly factor 6 (DNAAF6), a causative gene of primary ciliary dyskinesia, was isolated as an interacting protein with La ribonucleoprotein 6 (LARP6) that regulates ciliogenesis in multiciliated cells (MCCs). In MCCs of Xenopus embryos, LARP6 and DNAAF6 were colocalized in biomolecular condensates termed dynein axonemal particles and synergized to control ciliogenesis. Moreover, tubulin alpha 1c-like mRNA encoding α-tubulin protein, that is a major component of ciliary axoneme, was identified as a target mRNA regulated by binding LARP6. While DNAAF6 was necessary for high α-tubulin protein expression near the apical side of Xenopus MCCs during ciliogenesis, its mutant, which abolishes binding with LARP6, was unable to restore the expression of α-tubulin protein near the apical side of MCCs in Xenopus DNAAF6 morphant. These results indicated that the binding of LARP6 and DNAAF6 in dynein axonemal particles regulates highly expressed α-tubulin protein near the apical side of Xenopus MCCs during ciliogenesis.
Subject(s)
Cilia , Ribonucleoproteins , Tubulin , Xenopus Proteins , Xenopus laevis , Cilia/metabolism , Animals , Ribonucleoproteins/metabolism , Ribonucleoproteins/genetics , Tubulin/metabolism , Xenopus Proteins/metabolism , Xenopus Proteins/genetics , Humans , SS-B Antigen , Autoantigens/metabolism , Autoantigens/genetics , Protein Binding , Axoneme/metabolism , RNA, Messenger/metabolism , RNA, Messenger/geneticsABSTRACT
TAR DNA-binding protein 43 (TDP-43) proteinopathy in brain cells is the hallmark of amyotrophic lateral sclerosis (ALS) but its cause remains elusive. Asparaginase-like-1 protein (ASRGL1) cleaves isoaspartates, which alter protein folding and susceptibility to proteolysis. ASRGL1 gene harbors a copy of the human endogenous retrovirus HML-2, whose overexpression contributes to ALS pathogenesis. Here we show that ASRGL1 expression was diminished in ALS brain samples by RNA sequencing, immunohistochemistry, and western blotting. TDP-43 and ASRGL1 colocalized in neurons but, in the absence of ASRGL1, TDP-43 aggregated in the cytoplasm. TDP-43 was found to be prone to isoaspartate formation and a substrate for ASRGL1. ASRGL1 silencing triggered accumulation of misfolded, fragmented, phosphorylated and mislocalized TDP-43 in cultured neurons and motor cortex of female mice. Overexpression of ASRGL1 restored neuronal viability. Overexpression of HML-2 led to ASRGL1 silencing. Loss of ASRGL1 leading to TDP-43 aggregation may be a critical mechanism in ALS pathophysiology.
Subject(s)
Amyotrophic Lateral Sclerosis , Asparaginase , DNA-Binding Proteins , Neurons , TDP-43 Proteinopathies , Animals , Female , Humans , Male , Mice , Amyotrophic Lateral Sclerosis/metabolism , Amyotrophic Lateral Sclerosis/genetics , Amyotrophic Lateral Sclerosis/pathology , Asparaginase/genetics , Asparaginase/metabolism , Autoantigens/genetics , Autoantigens/metabolism , Brain/metabolism , Brain/pathology , DNA-Binding Proteins/metabolism , DNA-Binding Proteins/genetics , Motor Cortex/metabolism , Motor Cortex/pathology , Neurons/metabolism , Neurons/pathology , TDP-43 Proteinopathies/metabolism , TDP-43 Proteinopathies/pathology , TDP-43 Proteinopathies/genetics , Endogenous Retroviruses/genetics , Endogenous Retroviruses/metabolismABSTRACT
Recently, the progression of gastric cancer (GC), as one of the most ordinary malignant tumors, has been reported to be associated with circular RNAs. This study aimed to identify the role of circular RNA_LARP4 in GC. We performed real-time quantitative polymerase chain reaction (RT-qPCR) in 46 paired GC patients and GC cell lines to detect the expression of circular RNA_LARP4. Moreover, the role of circular RNA_LARP4 in GC proliferation was identified through proliferation assay and colony formation assay, while the role of circular RNA_LARP4 in GC metastasis was measured through scratch wound assay and transwell assay. Furthermore, the potential targets of circular RNA_LARP4 were predicted through bioinformatics methods and further identified by western blot assay and RT-qPCR. Circular RNA_LARP4 expression was remarkably lower in GC tissues compared with that in adjacent samples. Besides, cell proliferation of GC was inhibited after overexpression of circular RNA_LARP4, while cell migration and invasion of GC was inhibited after overexpression of circular RNA_LARP4. Furthermore, Upstream frameshift 1 (UPF1) was predicted as the potential target of circular RNA_LARP4 and was upregulated via overexpression of circular RNA_LARP4 in GC. Circular RNA_LARP4 inhibits GC cell proliferation and metastasis via targeting UPF1 in vitro, which might provide a new tumor suppressor in GC development.
Subject(s)
Cell Movement , Cell Proliferation , Gene Expression Regulation, Neoplastic , RNA, Circular , Stomach Neoplasms , Female , Humans , Male , Middle Aged , Autoantigens/genetics , Autoantigens/metabolism , Cell Line, Tumor , Cell Movement/genetics , Cell Proliferation/genetics , Neoplasm Invasiveness/genetics , Neoplasm Metastasis , RNA/genetics , RNA/metabolism , RNA, Circular/genetics , RNA, Circular/metabolism , SS-B Antigen , Stomach Neoplasms/genetics , Stomach Neoplasms/pathology , Stomach Neoplasms/metabolism , Trans-Activators/genetics , Trans-Activators/metabolism , Up-Regulation/geneticsABSTRACT
One of the probable hypotheses for the onset of autoimmunity is molecular mimicry. This study aimed to determine the HLA-II risk alleles for developing Hashimoto's thyroiditis (HT) in order to analyze the molecular homology between candidate pathogen-derived epitopes and potentially self-antigens (thyroid peroxidase, TPO) based on the presence of HLA risk alleles. HLA-DRB1/-DQB1 genotyping was performed in 100 HT patients and 330 ethnically matched healthy controls to determine the predisposing/protective alleles for HT disease. Then, in silico analysis was conducted to examine the sequence homology between epitopes derived from autoantigens and four potentially relevant pathogens and their binding capacities to HLA risk alleles based on peptide docking analysis. We identified HLA-DRB1*03:01, *04:02, *04:05, and *11:04 as predisposing alleles and DRB1*13:01 as a potentially predictive allele for HT disease. Also, DRB1*11:04 ~ DQB1*03:01 (Pc = 0.002; OR, 3.97) and DRB1*03:01 ~ DQB1*02:01 (Pc = 0.004; OR, 2.24) haplotypes conferred a predisposing role for HT. Based on logistic regression analysis, carrying risk alleles increased the risk of HT development 4.5 times in our population (P = 7.09E-10). Also, ROC curve analysis revealed a high predictive power of those risk alleles for discrimination of the susceptible from healthy individuals (AUC, 0.70; P = 6.6E-10). Analysis of peptide sequence homology between epitopes of TPO and epitopes derived from four candidate microorganisms revealed a homology between envelop glycoprotein D of herpes virus and sequence 151-199 of TPO with remarkable binding capacity to HLA-DRB1*03:01 allele. Our findings indicate the increased risk of developing HT in those individuals carrying HLA risk alleles which can also be related to herpes virus infection.
Subject(s)
Alleles , Autoantigens , Epitopes , Genetic Predisposition to Disease , HLA-DQ beta-Chains , HLA-DRB1 Chains , Hashimoto Disease , Humans , Male , Female , Hashimoto Disease/genetics , Hashimoto Disease/immunology , Adult , Iran , HLA-DRB1 Chains/genetics , HLA-DQ beta-Chains/genetics , Autoantigens/immunology , Autoantigens/genetics , Epitopes/immunology , Epitopes/genetics , Middle Aged , Case-Control Studies , Iodide Peroxidase/genetics , Iodide Peroxidase/immunology , Haplotypes , Genotype , Gene FrequencyABSTRACT
Hepatitis C virus (HCV) infection is tightly connected to the lipid metabolism with lipid droplets (LDs) serving as assembly sites for progeny virions. A previous LD proteome analysis identified annexin A3 (ANXA3) as an important HCV host factor that is enriched at LDs in infected cells and required for HCV morphogenesis. To further characterize ANXA3 function in HCV, we performed proximity labeling using ANXA3-BioID2 as bait in HCV-infected cells. Two of the top proteins identified proximal to ANXA3 during HCV infection were the La-related protein 1 (LARP1) and the ADP ribosylation factor-like protein 8B (ARL8B), both of which have been previously described to act in HCV particle production. In follow-up experiments, ARL8B functioned as a pro-viral HCV host factor without localizing to LDs and thus likely independent of ANXA3. In contrast, LARP1 interacts with HCV core protein in an RNA-dependent manner and is translocated to LDs by core protein. Knockdown of LARP1 decreased HCV spreading without altering HCV RNA replication or viral titers. Unexpectedly, entry of HCV particles and E1/E2-pseudotyped lentiviral particles was reduced by LARP1 depletion, whereas particle production was not altered. Using a recombinant vesicular stomatitis virus (VSV)ΔG entry assay, we showed that LARP1 depletion also decreased entry of VSV with VSV, MERS, and CHIKV glycoproteins. Therefore, our data expand the role of LARP1 as an HCV host factor that is most prominently involved in the early steps of infection, likely contributing to endocytosis of viral particles through the pleiotropic effect LARP1 has on the cellular translatome.