RESUMO
Drosophila melanogaster Down syndrome cell adhesion molecule 1 (Dscam1) encodes 19,008 diverse ectodomain isoforms via the alternative splicing of exon 4, 6, and 9 clusters. However, whether individual isoforms or exon clusters have specific significance is unclear. Here, using phenotype-diversity correlation analysis, we reveal the redundant and specific roles of Dscam1 diversity in neuronal wiring. A series of deletion mutations were performed from the endogenous locus harboring exon 4, 6, or 9 clusters, reducing to 396 to 18,612 potential ectodomain isoforms. Of the 3 types of neurons assessed, dendrite self/non-self discrimination required a minimum number of isoforms (approximately 2,000), independent of exon clusters or isoforms. In contrast, normal axon patterning in the mushroom body and mechanosensory neurons requires many more isoforms that tend to associate with specific exon clusters or isoforms. We conclude that the role of the Dscam1 diversity in dendrite self/non-self discrimination is nonspecifically mediated by its isoform diversity. In contrast, a separate role requires variable domain- or isoform-related functions and is essential for other neurodevelopmental contexts, such as axonal growth and branching. Our findings shed new light on a general principle for the role of Dscam1 diversity in neuronal wiring.
Assuntos
Síndrome de Down , Proteínas de Drosophila , Animais , Drosophila melanogaster/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Molécula 1 de Adesão Celular/genética , Molécula 1 de Adesão Celular/metabolismo , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Síndrome de Down/genética , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Neurônios/metabolismoRESUMO
Glucosidase II beta subunit (GluIIß), encoded from PRKCSH, is a subunit of the glucosidase II enzyme responsible for quality control of N-linked glycoprotein folding and suppression of GluIIß led to inhibitory effect of the receptor tyrosine kinase (RTKs) activities known to be critical for survival and development of cancer. In this study, we investigated the effect of GluIIß knockout on the global gene expression of cancer cells and its impact on functions of immune cells. GluIIß knockout lung adenocarcinoma A549 cell line was generated using CRISPR/Cas9-based genome editing system and subjected to transcriptomic analysis. Among 23,502 expressed transcripts, 1068 genes were significantly up-regulated and 807 genes greatly down-regulated. The KEGG enrichment analysis showed significant down-regulation of genes related extracellular matrix (ECM), ECM-receptor interaction, cytokine-cytokine receptor interaction and cell adhesion molecules (CAMs) in GluIIß knockout cells. Of 9 CAMs encoded DEG identified by KEGG enrichment analysis, real time RT-PCR confirmed 8 genes to be significantly down-regulated in all 3 different GluIIß knockout clones, which includes cadherin 4 (CDH4), cadherin 2 (CDH2), versican (VCAN), integrin subunit alpha 4 (ITGA4), endothelial cell-selective adhesion molecule (ESAM), CD274 (program death ligand-1 (PD-L1)), Cell Adhesion Molecule 1 (CADM1), and Nectin Cell Adhesion Molecule 3 (NECTIN3). Whereas PTPRF (Protein Tyrosine Phosphatase Receptor Type F) was significantly decreased only in 1 out of 3 knockout clones. Microscopic analysis revealed distinctively different cell morphology of GluIIß knockout cells with lesser cytoplasmic and cell surface area compared to parental A549 cells and non-targeted transfected cells.Further investigations revealed that Jurkat E6.1 T cells or human peripheral blood mononuclear cells (PBMCs) co-cultured with GluIIß knockout A549 exhibited significantly increased viability and tumor cell killing activity compared to those co-cultured with non-target transfected cells. Analysis of cytokine released from Jurkat E6.1 T cells co-cultured with GluIIß knockout A549 cells showed significant increased level of angiogenin and significant decreased level of ENA-78. In conclusion, knockout of GluIIß from cancer cells induced altered gene expression profile that improved anti-tumor activities of co-cultured T lymphocytes and PBMCs thus suppression of GluIIß may represent a novel approach of boosting anti-tumor immunity.
Assuntos
Moléculas de Adesão Celular , Leucócitos Mononucleares , alfa-Glucosidases , Humanos , Células A549 , Moléculas de Adesão Celular/genética , Perfilação da Expressão Gênica , Citocinas , Adesão Celular , Molécula 1 de Adesão CelularRESUMO
Measles virus (MeV), the causative agent of measles, is an enveloped RNA virus of the family Paramyxoviridae, which remains an important cause of childhood morbidity and mortality. MeV has two envelope glycoproteins, the hemagglutinin (H) and fusion (F) proteins. During viral entry or virus-mediated fusion between infected cells and neighboring susceptible cells, the head domain of the H protein initially binds to its receptors, signaling lymphocytic activation molecule family member 1 (SLAM) and nectin-4, and then the stalk region of the H protein transmits the fusion-triggering signal to the F protein. MeV may persist in the human brain and cause a fatal neurodegenerative disease, subacute sclerosing panencephalitis (SSPE). Recently, we showed, using in vitro cell culture, that cell adhesion molecule (CADM) 1 and CADM2 are host factors that trigger hyperfusogenic mutant F proteins, causing cell-to-cell fusion and the transfer of the MeV genome between neurons. Unlike conventional receptors, CADM1 and CADM2 interact in cis (on the same membrane) with the H protein and then trigger membrane fusion. Here, we show that alanine substitutions in part of the stalk region (positions 171-175) abolish the ability of the H protein to mediate membrane fusion triggered by CADM1 and CADM2, but not by SLAM. The recombinant hyperfusogenic MeV carrying this mutant H protein loses its ability to spread in primary mouse neurons as well as its neurovirulence in experimentally infected suckling hamsters. These results indicate that CADM1 and CADM2 are key molecules for MeV propagation in the brain and its neurovirulence in vivo. IMPORTANCE Measles is an acute febrile illness with skin rash. Despite the availability of highly effective vaccines, measles is still an important cause of childhood morbidity and mortality in many countries. The World Health Organization estimates that more than 120,000 people died from measles worldwide in 2021. Measles virus (MeV), the causative agent of measles, can also cause a fatal progressive neurological disorder, subacute sclerosing panencephalitis (SSPE), several years after acute infection. There is currently no effective treatment for this disease. In this study, using recombinant MeVs with altered receptor usage patterns, we show that cell adhesion molecule (CADM) 1 and CADM2 are host factors critical for MeV spread in neurons and its neurovirulence. These findings further our understanding of the molecular mechanism of MeV neuropathogenicity.
Assuntos
Sarampo , Doenças Neurodegenerativas , Panencefalite Esclerosante Subaguda , Cricetinae , Humanos , Camundongos , Animais , Vírus do Sarampo/fisiologia , Panencefalite Esclerosante Subaguda/genética , Hemaglutininas/metabolismo , Doenças Neurodegenerativas/metabolismo , Proteínas Virais de Fusão/genética , Proteínas Virais de Fusão/metabolismo , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Proteínas Recombinantes/metabolismo , Neurônios , Molécula 1 de Adesão Celular/metabolismoRESUMO
Measles virus (MeV), an enveloped RNA virus in the family Paramyxoviridae, usually causes acute febrile illness with skin rash but in rare cases persists in the brain, causing a progressive neurological disorder, subacute sclerosing panencephalitis (SSPE). MeV bears two envelope glycoproteins, the hemagglutinin (H) and fusion (F) proteins. The H protein possesses a head domain that initially mediates receptor binding and a stalk domain that subsequently transmits the fusion-triggering signal to the F protein. We recently showed that cell adhesion molecule 1 (CADM1; also known as IGSF4A, Necl-2, and SynCAM1) and CADM2 (also known as IGSF4D, Necl-3, and SynCAM2) are host factors enabling cell-cell membrane fusion mediated by hyperfusogenic F proteins of neuropathogenic MeVs as well as MeV spread between neurons lacking the known receptors. CADM1 and CADM2 interact in cis with the H protein on the same cell membrane, triggering hyperfusogenic F protein-mediated membrane fusion. Multiple isoforms of CADM1 and CADM2 containing various lengths of their stalk regions are generated by alternative splicing. Here, we show that only short-stalk isoforms of CADM1 and CADM2 predominantly expressed in the brain induce hyperfusogenic F protein-mediated membrane fusion. While the known receptors interact in trans with the H protein through its head domain, these isoforms can interact in cis even with the H protein lacking the head domain and trigger membrane fusion, presumably through its stalk domain. Thus, our results unveil a new mechanism of viral fusion triggering by host factors. IMPORTANCE Measles, an acute febrile illness with skin rash, is still an important cause of childhood morbidity and mortality worldwide. Measles virus (MeV), the causative agent of measles, may also cause a progressive neurological disorder, subacute sclerosing panencephalitis (SSPE), several years after acute infection. The disease is fatal, and no effective therapy is available. Recently, we reported that cell adhesion molecule 1 (CADM1) and CADM2 are host factors enabling MeV cell-to-cell spread in neurons. These molecules interact in cis with the MeV attachment protein on the same cell membrane, triggering the fusion protein and causing membrane fusion. CADM1 and CADM2 are known to exist in multiple splice isoforms. In this study, we report that their short-stalk isoforms can induce membrane fusion by interacting in cis with the viral attachment protein independently of its receptor-binding head domain. This finding may have important implications for cis-acting fusion triggering by host factors.
Assuntos
Molécula 1 de Adesão Celular/metabolismo , Células Gigantes/virologia , Hemaglutininas Virais/metabolismo , Interações Hospedeiro-Patógeno , Vírus do Sarampo/fisiologia , Sarampo/metabolismo , Sarampo/virologia , Animais , Encéfalo/metabolismo , Encéfalo/virologia , Molécula 1 de Adesão Celular/genética , Células Cultivadas , Cricetinae , Modelos Biológicos , Ligação Proteica , Isoformas de Proteínas , Proteínas Virais de Fusão/metabolismoRESUMO
BACKGROUND: Methylation levels may be associated with and serve as markers to predict risk of progression of precancerous cervical lesions. We conducted an epigenome-wide association study (EWAS) of CpG methylation and progression to high-grade cervical intraepithelial neoplasia (CIN2 +) following an abnormal screening test. METHODS: A prospective US cohort of 289 colposcopy patients with normal or CIN1 enrollment histology was assessed. Baseline cervical sample DNA was analyzed using Illumina HumanMethylation 450K (n = 76) or EPIC 850K (n = 213) arrays. Participants returned at provider-recommended intervals and were followed up to 5 years via medical records. We assessed continuous CpG M values for 9 cervical cancer-associated genes and time-to-progression to CIN2+. We estimated CpG-specific time-to-event ratios (TTER) and hazard ratios using adjusted, interval-censored Weibull accelerated failure time models. We also conducted an exploratory EWAS to identify novel CpGs with false discovery rate (FDR) < 0.05. RESULTS: At enrollment, median age was 29.2 years; 64.0% were high-risk HPV-positive, and 54.3% were non-white. During follow-up (median 24.4 months), 15 participants progressed to CIN2+. Greater methylation levels were associated with a shorter time-to-CIN2+ for CADM1 cg03505501 (TTER = 0.28; 95%CI 0.12, 0.63; FDR = 0.03) and RARB Cluster 1 (TTER = 0.46; 95% CI 0.29, 0.71; FDR = 0.01). There was evidence of similar trends for DAPK1 cg14286732, PAX1 cg07213060, and PAX1 Cluster 1. The EWAS detected 336 novel progression-associated CpGs, including those located in CpG islands associated with genes FGF22, TOX, COL18A1, GPM6A, XAB2, TIMP2, GSPT1, NR4A2, and APBB1IP. CONCLUSIONS: Using prospective time-to-event data, we detected associations between CADM1-, DAPK1-, PAX1-, and RARB-related CpGs and cervical disease progression, and we identified novel progression-associated CpGs. IMPACT: Methylation levels at novel CpG sites may help identify individuals with ≤CIN1 histology at higher risk of progression to CIN2+ and inform risk-based cervical cancer screening guidelines.
Assuntos
Infecções por Papillomavirus , Displasia do Colo do Útero , Neoplasias do Colo do Útero , Feminino , Humanos , Estados Unidos , Adulto , Neoplasias do Colo do Útero/patologia , Estudos Prospectivos , Epigenoma , Detecção Precoce de Câncer , Metilação de DNA , Displasia do Colo do Útero/diagnóstico , Infecções por Papillomavirus/complicações , Papillomaviridae/genética , Molécula 1 de Adesão Celular/genéticaRESUMO
The interruption of normal cell cycle execution acts as an important part to the development of leukemia. It was reported that microRNAs (miRNAs) were closely related to tumorigenesis and progression, and their aberrant expression had been demonstrated to play a crucial role in numerous types of cancer. Our previous study showed that miR-1246 was preferentially overexpressed in chemo-resistant leukemia cell lines, and participated in process of cell cycle progression and multidrug resistant regulation. However, the underlying mechanism remains unclear. In present study, bioinformatics prediction and dual luciferase reporter assay indicated that CADM1 was a direct target of miR-1246. Evidently decreased expression of CADM1 was observed in relapsed primary leukemia patients and chemo-resistant cell lines. Our results furtherly proved that inhibition of miR-1246 could significantly enhance drug sensitivity to Adriamycin (ADM), induce cell cycle arrest at G0/G1 phase, promote cell apoptosis, and relieve its suppression on CADM1 in K562/ADM and HL-60/RS cells. Interference with CADM1 could reduce the increased drug sensitivity induced by miR-1246 inhibition, and notably restore drug resistance by promoting cell cycle progression and cell survival via regulating CDKs/Cyclins complexes in chemo-resistant leukemia cells. Above all, our results demonstrated that CADM1 attenuated the role of miR-1246 in promoting cell cycle progression and cell survival, thus influencing multidrug resistance within chemo-resistant leukemia cells via CDKs/Cyclins. Higher expression of miR-1246 and lower expression of CADM1 might be risk factors for leukemia.
Assuntos
Leucemia , MicroRNAs , Humanos , MicroRNAs/metabolismo , Células HL-60 , Doxorrubicina/farmacologia , Ciclo Celular/genética , Leucemia/tratamento farmacológico , Leucemia/genética , Ciclinas , Proliferação de Células , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/genética , Molécula 1 de Adesão Celular/genéticaRESUMO
T cell activation is a multifaceted process that depends on the activation of the T cell receptor (TCR). However, other coreceptors are also strictly necessary to provide co-signals and modulate the immune response. However, to date, most of these coreceptors are unknown in fish or their information is very limited. Therefore, in this work, we have identified the cytotoxic and regulatory T cell molecule, CRTAM, and its ligand, the cell adhesion molecule 1, CADM1, in European seabass (Dicentrarchus labrax) and gilthead seabream (Sparus aurata); and evaluated their transcriptional levels. Both putative proteins showed the canonical architecture observed in mammals, where CRTAM exhibited two immunoglobulin domains and CADM1, both the a and b forms, exhibited three of these domains. In addition, phylogeny and synteny analyses showed their conservation throughout vertebrate evolution. We found constitutive expression of all three genes, with crtam and cadm1a being predominant in immune tissues such as spleen, thymus and head-kidney (HK), while cadm1b expression was more limited to the brain. In vitro, only the T cell mitogen phytohemagglutinin (PHA) up-regulated the transcription of crtam and cadm1a in HK leucocytes. Nodavirus (NNV) infection elicited an up-regulation of crtam and cadm1a in brain and HK, appearing earlier in seabream than in seabass, which could explain the resistance of seabream to the development of nodavirus disease. In addition, they are up-regulated during the innate cell-mediated cytotoxic response in seabream but not in seabass. Altogether, our data seem to indicate that CRTAM is more related to the innate cytotoxicity in seabream and more in the specific and T cell-mediated cytotoxicity in seabass. Our results highlight the importance of CRTAM and CADM1 as important molecules in the activation of T lymphocytes in seabass and seabream, but further studies are needed.
Assuntos
Antineoplásicos , Bass , Dourada , Animais , Molécula 1 de Adesão Celular , Linfócitos T Reguladores , Ligantes , MamíferosRESUMO
Asbestos is a fibrous silicate mineral exhibiting biopersistence and carcinogenic properties and contributes to mesothelioma. Despite the concept of gene-environmental interaction in pathogenesis of mesothelioma, the possible pathophysiological changes of mesothelial cells simultaneously with SET domain containing 2 (SETD2) loss and asbestos exposure remains obscure. Herein, CRISPR/Cas9-mediated SETD2 knockout Met-5A mesothelial cells (Met-5ASETD2-KO ) were established and exposed with crocidolite, an amphibole asbestos. Cell viability of Met-5ASETD2-KO appeared to dramatically decrease with ≥2.5 µg/cm2 crocidolite exposure as compared with Met-5A, although no cytotoxicity and apoptosis changes of Met-5ASETD2-KO and Met-5A was evident with 1.25 µg/cm2 crocidolite exposure for 48 h. RNA sequencing uncovered top 50 differentially expressed genes (DEGs) between 1.25 µg/cm2 crocidolite exposed Met-5ASETD2-KO (Cro-Met-5ASETD2-KO ) and 1.25 µg/cm2 crocidolite exposed Met-5A (Cro-Met-5A), and ITGA4, THBS2, MYL7, RAC2, CADM1, and CLDN11 appeared to be the primary DEGs involved with adhesion in gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. Cro-Met-5ASETD2-KO had strong migration but mild adhesion behavior as compared with Cro-Met-5A. Additionally, crocidolite tended to increase migration of Met-5ASETD2-KO but inhibited migration of Met-5A when compared with their corresponding cells without crocidolite exposure, although no further adhesion property changes was evident for both cells in response to crocidolite. Therefore, crocidolite may affect adhesion-related gene expression and modify adhesion and migration behavior for SETD2-depleted Met-5A, which could provide preliminary insight regarding the potential role of SETD2 in the cell behavior of asbestos-related malignant mesothelial cell.
Assuntos
Amianto , Mesotelioma , Humanos , Asbesto Crocidolita/toxicidade , Asbesto Crocidolita/metabolismo , Epitélio , Amianto/toxicidade , Silicatos , Molécula 1 de Adesão Celular/metabolismoRESUMO
BACKGROUND: The specific pathogenesis of atrial fibrillation (AF) remains unclear. In this study, we examined the expression of differential messenger RNAs (mRNAs), circular RNAs (circRNAs), and long-stranded noncoding RNAs (lncRNAs) from human peripheral blood mononuclear cells to initially construct a circRNA/lncRNA-miRNA-mRNA ceRNA regulatory network to explore the pathogenesis of AF and to screen for potential biomarkers. METHODS: A total of four pairs of AF cases and healthy subjects were selected to detect differentially expressed mRNAs, circRNAs, and lncRNAs in peripheral blood mononuclear cells by microarray analysis. And 20 pairs of peripheral blood from AF patients and healthy subjects were selected for validation of mRNA, circRNA, and lncRNA by quantitative real-time PCR (qRT-PCR).The relevant ceRNA networks were constructed by GO and KEGG and correlation analysis. RESULTS: The results showed that compared with healthy subjects, there were 813 differentially expressed mRNAs (DEmRNAs) in peripheral blood monocytes of AF, including 445 upregulated genes and 368 downregulated genes, 120 differentially expressed circRNAs (DEcircRNAs), including 65 upregulated and 55 downregulated, 912 differentially expressed lncRNAs (DElncRNAs), including 531 upregulated and 381 downregulated lncRNAs. GO and KEGG analysis of DERNA revealed the biological processes and pathways involved in AF. Based on microarray data and predicted miRNAs, a ceRNA network containing 34 mRNAs, 212 circRNAs, 108 lncRNAs, and 38 miRNAs was constructed. CONCLUSION: We revealed a novel ceRNA network in AF and showed that downregulated XIST, circRNA_2773, and CADM1 were negatively correlated with miR-486-5p expression and had a potential targeting relationship with miR-486-5p.
Assuntos
Fibrilação Atrial , MicroRNAs , RNA Longo não Codificante , Humanos , MicroRNAs/genética , RNA Circular/genética , RNA Mensageiro/genética , RNA Longo não Codificante/genética , Leucócitos Mononucleares/metabolismo , Redes Reguladoras de Genes , Biomarcadores , Molécula 1 de Adesão Celular/genéticaRESUMO
Nonalcoholic fatty liver disease (NAFLD) has become a widely studied subject due to its increasing prevalence and links to diseases such as type 2 diabetes and obesity. It has severe complications, including nonalcoholic steatohepatitis, cirrhosis, hepatocellular carcinoma, and portal hypertension that can lead to liver transplantation in some cases. To better prevent and treat this pathology, it is important to understand its underlying physiology. Here, we identify two main factors that play a crucial role in the pathophysiology of NAFLD: oxidative stress and the key role of carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1). We discuss the pathophysiology linking these factors to NAFLD pathophysiology.
Assuntos
Hepatopatia Gordurosa não Alcoólica , Estresse Oxidativo , Humanos , Antígeno Carcinoembrionário , Molécula 1 de Adesão Celular , Diabetes Mellitus Tipo 2/patologia , Hepatopatia Gordurosa não Alcoólica/metabolismoRESUMO
Cell adhesion proteins of the Cadm (SynCAM/Necl) family regulate myelination and the organization of myelinated axons. In the peripheral nervous system (PNS), intercellular contact between Schwann cells and their underlying axons is believed to be mediated by binding of glial Cadm4 to axonal Cadm3 or Cadm2. Nevertheless, given that distinct neurons express different combinations of the Cadm proteins, the identity of the functional axonal ligand for Cadm4 remains to be determined. Here, we took a genetic approach to compare the phenotype of Cadm4 null mice, which exhibit abnormal distribution of Caspr and Kv1 potassium channels, with mice lacking different combinations of Cadm1-Cadm3 genes. We show that in contrast to mice lacking the single Cadm1, Cadm2, or Cadm3 genes, genetic ablation of all three phenocopies the abnormalities detected in the absence of Cadm4. Similar defects were observed in double mutant mice lacking Cadm3 and Cadm2 (i.e., Cadm3-/-/Cadm2-/-) or Cadm3 and Cadm1 (i.e., Cadm3-/-/Cadm1-/-), but not in mice lacking Cadm1 and Cadm2 (i.e., Cadm1-/-/Cadm2-/-). Furthermore, axonal organization abnormalities were also detected in Cadm3 null mice that were heterozygous for the two other axonal Cadms. Our results identify Cadm3 as the main axonal ligand for glial Cadm4, and reveal that its absence could be compensated by the combined action of Cadm2 and Cadm1.SIGNIFICANCE STATEMENT Myelination by Schwann cells enables fast conduction of action potentials along motor and sensory axons. In these nerves, Schwann cell-axon contact is mediated by cell adhesion molecules of the Cadm family. Cadm4 in Schwann cells regulates axonal ensheathment and myelin wrapping, as well as the organization of the axonal membrane, but the identity of its axonal ligands is not clear. Here, we reveal that Cadm mediated axon-glia interactions depend on a hierarchical adhesion code that involves multiple family members. Our results provide important insights into the molecular mechanisms of axon-glia communication, and the function of Cadm proteins in PNS myelin.
Assuntos
Axônios/metabolismo , Molécula 1 de Adesão Celular/deficiência , Moléculas de Adesão Celular/deficiência , Comunicação Celular/fisiologia , Imunoglobulinas/deficiência , Fibras Nervosas Mielinizadas/metabolismo , Neuroglia/metabolismo , Animais , Molécula 1 de Adesão Celular/genética , Moléculas de Adesão Celular/genética , Imunoglobulinas/genética , Camundongos , Camundongos Knockout , Nervos Periféricos/metabolismoRESUMO
The initial step of organ infiltration of malignant cells is the interaction with host vascular endothelial cells, which is often mediated by specific combinations of cell adhesion molecules. Cell adhesion molecule 1 (CADM1) is overexpressed in adult T-cell leukemia/lymphoma (ATL) and provides a cell-surface diagnostic marker. CADM1 promotes the adhesion of ATL cells to vascular endothelial cells and multiple organ infiltration in mice. However, its binding partner on host cells has not yet been identified. In this study, we show that CADM1 promotes transendothelial migration of ATL cells in addition to the adhesion to vascular endothelial cells. Moreover, CADM1 enhances liver infiltration of mouse T-cell lymphoma cells, EL4, after tail vein injection, whereas a CADM1 mutant lacking adhesive activity did not. Among the known CADM1-binding proteins expressed in primary endothelial cells, only CADM1 and CADM4 could induce morphological extension of ATL cells when plated onto glass coated with these proteins. Furthermore, CADM1-mediated liver infiltration of EL4 cells was canceled in conventional and vascular endothelium-specific Cadm1 knockout mice, whereas it was not canceled in Cadm4 knockout mice. These results suggest that CADM1 on host vascular endothelial cells is required for organ infiltration of ATL and other T-cell lymphomas expressing CADM1.
Assuntos
Molécula 1 de Adesão Celular/metabolismo , Endotélio Vascular , Linfoma de Células T , Animais , Adesão Celular , Molécula 1 de Adesão Celular/genética , Células Endoteliais/metabolismo , Endotélio Vascular/metabolismo , Humanos , Imunoglobulinas/química , Imunoglobulinas/genética , Linfoma de Células T/genética , CamundongosRESUMO
Transfer RNA-derived fragments are a group of small noncoding single-stranded RNA that play essential roles in multiple diseases. However, their biological functions in carcinogenesis are not well understood. In this study, 5'tRF-Gly was found to have significantly high expression in hepatocellular carcinoma (HCC), and the upregulation of 5'tRF-Gly was positively correlated with tumor size and tumor metastasis. Overexpression of 5'tRF-Gly induced increased growth rate and metastasis in HCC cells in vitro and in nude mice, while knockdown showed the opposite effect. Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) was confirmed to be a direct target of 5'tRF-Gly in HCC. In addition, the cytological effect of CEACAM1 knockdown proved to be similar to the overexpression of 5'tRF-Gly. Moreover, attenuation of CEACAM1 expression rescued the 5'tRF-Gly-mediated promoting effects on HCC cells. These data show that 5'tRF-Gly is a new tumor-promoting factor and could be a potential diagnostic biomarker or new therapeutic target for HCC.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , RNA Longo não Codificante , Animais , Antígenos CD , Antígeno Carcinoembrionário/genética , Antígeno Carcinoembrionário/metabolismo , Carcinoma Hepatocelular/patologia , Molécula 1 de Adesão Celular/metabolismo , Moléculas de Adesão Celular , Linhagem Celular Tumoral , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/patologia , Camundongos , Camundongos Nus , RNA , RNA Longo não Codificante/genética , RNA de TransferênciaRESUMO
Measles virus (MeV), an enveloped RNA virus in the family Paramyxoviridae, is still an important cause of childhood morbidity and mortality worldwide. MeV usually causes acute febrile illness with skin rash, but in rare cases persists in the brain, causing a progressive neurological disorder, subacute sclerosing panencephalitis (SSPE). The disease is fatal, and no effective therapy is currently available. Although transsynaptic cell-to-cell transmission is thought to account for MeV propagation in the brain, neurons do not express the known receptors for MeV. Recent studies have shown that hyperfusogenic changes in the MeV fusion (F) protein play a key role in MeV propagation in the brain. However, how such mutant viruses spread in neurons remains unexplained. Here, we show that cell adhesion molecule 1 (CADM1; also known as IGSF4A, Necl-2, and SynCAM1) and CADM2 (also known as IGSF4D, Necl-3, SynCAM2) are host factors that enable MeV to cause membrane fusion in cells lacking the known receptors and to spread between neurons. During enveloped virus entry, a cellular receptor generally interacts in trans with the attachment protein on the envelope. However, CADM1 and CADM2 interact in cis with the MeV attachment protein on the same cell membrane, causing the fusion protein triggering and membrane fusion. Knockdown of CADM1 and CADM2 inhibits syncytium formation and virus transmission between neurons that are both mediated by hyperfusogenic F proteins. Thus, our results unravel the molecular mechanism (receptor-mimicking cis-acting fusion triggering) by which MeV spreads transsynaptically between neurons, thereby causing SSPE. IMPORTANCE Measles virus (MeV), an enveloped RNA virus, is the causative agent of measles, which is still an important cause of childhood morbidity and mortality worldwide. Persistent MeV infection in the brain causes a fatal progressive neurological disorder, subacute sclerosing panencephalitis (SSPE), several years after acute infection. However, how MeV spreads in neurons, which are mainly affected in SSPE, remains largely unknown. In this study, we demonstrate that cell adhesion molecule 1 (CADM1) and CADM2 are host factors enabling MeV spread between neurons. During enveloped virus entry, a cellular receptor generally interacts in trans with the attachment protein on the viral membrane (envelope). Remarkably, CADM1 and CADM2 interact in cis with the MeV attachment protein on the same membrane, triggering the fusion protein and causing membrane fusion, as viral receptors usually do in trans. Careful screening may lead to more examples of such "receptor-mimicking cis-acting fusion triggering" in other viruses.
Assuntos
Molécula 1 de Adesão Celular/fisiologia , Moléculas de Adesão Celular/fisiologia , Vírus do Sarampo/patogenicidade , Panencefalite Esclerosante Subaguda/virologia , Internalização do Vírus , Animais , Linhagem Celular , Chlorocebus aethiops , Células Gigantes/virologia , Humanos , Camundongos , Células Vero , Proteínas do Envelope Viral/metabolismo , Proteínas Virais de Fusão/metabolismoRESUMO
ABSTRACT: High glucose-induced endothelial activation plays critical roles in the development of diabetic vascular complications. R-spondin 3 could inhibit inflammatory damage, and diabetic vascular inflammation is secondary to endothelial activation. In this article, we identify R-spondin 3 as a novel regulator of high glucose-induced endothelial activation. We found that the serum levels of R-spondin 3 were significantly reduced in type 2 diabetic patients and db/db mice. We observed that the increased expressions of vascular cell adhesion molecule-1, intercellular cell adhesion molecule-1, and monocyte chemoattractant protein-1 (endothelial activation makers) in high glucose-stimulated human umbilical vein endothelial cell lines (HUVECs) could be inhibited by overexpressing R-spondin 3 or human R-spondin 3 recombinant protein. Subsequently, high glucose-induced adhesion and migration of human myeloid leukemia mononuclear cells (THP-1 cells) to HUVECs were markedly suppressed by the overexpression of R-spondin 3 in HUVECs. Moreover, the inhibitory effect of R-spondin 3 on the expressions of vascular cell adhesion molecule-1, intercellular cell adhesion molecule-1, and monocyte chemoattractant protein-1 in high glucose-treated HUVECs could be blocked by knockdown of leucine-rich G protein-coupled receptor 4 (R-spondin 3 receptor) or the specific inhibitor of Wnt/ß-catenin pathway. Taken together, R-spondin 3 could suppress high glucose-induced endothelial activation through leucine-rich G protein-coupled receptor 4/Wnt/ß-catenin pathway.
Assuntos
Quimiocina CCL2 , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Via de Sinalização Wnt , beta Catenina , Animais , Molécula 1 de Adesão Celular , Glucose , Molécula 1 de Adesão Intercelular , Leucina , Camundongos , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Molécula 1 de Adesão de Célula Vascular , beta Catenina/metabolismoRESUMO
Anorexia nervosa (AN) is a devastating disorder with evidence of underexplored heritability. Twin and family studies estimate heritability (h2 ) to be 57%-64%, and genome-wide association studies (GWAS) reveal significant genetic correlations with psychiatric and anthropometric traits and a total of nine genome-wide significant loci. Whether significantly associated single nucleotide polymorphisms identified by GWAS are causal or tag true causal variants, remains to be elucidated. We propose a novel method for bridging this knowledge gap by fine-mapping short structural variants (SSVs) in and around GWAS-identified loci. SSV fine-mapping of loci associated with complex disorders such as schizophrenia, amyotrophic lateral sclerosis, and Alzheimer's disease has uncovered genetic risk markers, phenotypic variability between patients, new pathological mechanisms, and potential therapeutic targets. We analyze previous investigations' methods and propose utilizing an evaluation algorithm to prioritize 10 SSVs for each of the top two AN GWAS-identified loci followed by Sanger sequencing and fragment analysis via capillary electrophoresis to characterize these SSVs for case/control association studies. Success of previous SSV analyses in complex disorders and effective utilization of similar methodologies supports our proposed method. Furthermore, the structural and spatial properties of the 10 SSVs identified for each of the top two AN GWAS-associated loci, cell adhesion molecule 1 (CADM1) and NCK interacting protein with SH3 domain (NCKIPSD), are similar to previous studies. We propose SSV fine-mapping of AN-associated loci will identify causal genetic architecture. Deepening understandings of AN may lead to novel therapeutic targets and subsequently increase quality-of-life for individuals living with the illness. PUBLIC SIGNIFICANCE STATEMENT: Anorexia nervosa is a severe and complex illness, arising from a combination of environmental and genetic factors. Recent studies estimate the contribution of genetic variability; however, the specific DNA sequences and how they contribute remain unknown. We present a novel approach, arguing that the genetic variant class, short structural variants, could answer this knowledge gap and allow development of biologically targeted therapeutics, improving quality-of-life and patient outcomes for affected individuals.
Assuntos
Anorexia Nervosa , Anorexia Nervosa/genética , Estudos de Casos e Controles , Molécula 1 de Adesão Celular/genética , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Humanos , Fenótipo , Polimorfismo de Nucleotídeo ÚnicoRESUMO
INTRODUCTION AND OBJECTIVES: Long non-coding RNAs (lncRNAs) have great potential as therapeutic targets in hepatocellular carcinoma (HCC). In this study, we aimed to uncover the function and molecular mechanism of long intergenic non-protein coding RNA 1006 (LINC01006) in HCC. MATERIALS AND METHODS: Mice were injected with HCC cells in order to establish the HCC model. Quantitative reverse transcription polymerase chain reaction was used to determine the expression levels of LINC01006, cell adhesion molecule 1 (CADM1), and microRNA (miR)-194-5p in HCC tissues and cells. The cell proliferation, invasion, and migration abilities were assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide, transwell, and wound healing assays. The interrelation between LINC01006, miR-194-5p, and CADM1 was confirmed by a dual-luciferase reporter assay. Western blotting was employed to assess the relative protein expression level of CADM1. RESULTS: LINC01006 and CADM1 displayed upregulation, but miR-194-5p exhibited downregulation in HCC cells and tissues. Short hairpin (sh)-LINC01006 and miR-194-5p mimics repressed the proliferative, migratory, and invasive capacities of HCC cells, and injection of sh-LINC01006 restrained the growth of HCC tumours in mice. LINC01006 served as a competing endogenous RNA of miR-194-5p and was inversely correlated with miR-194-5p. CADM1 was targeted by miR-194-5p, inversely correlated with miR-194-5p, and positively associated with LINC01006. Furthermore, transfection of pcDNA-CADM1 or the miR-194-5p inhibitor reversed the suppressive effects of sh-LINC01006 on the proliferation, invasion, and migration abilities of HCC cells. CONCLUSIONS: Downregulation of LINC01006 repressed the development of HCC by sponging miR-194-5p to modulate the expression of CADM1, implying its potential as a therapeutic target for HCC.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , MicroRNAs , RNA Longo não Codificante , Animais , Carcinoma Hepatocelular/patologia , Molécula 1 de Adesão Celular/genética , Molécula 1 de Adesão Celular/metabolismo , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/patologia , Camundongos , MicroRNAs/genética , MicroRNAs/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismoRESUMO
BACKGROUND: The pathogenesis of connective tissue disease-associated interstitial lung disease (CTD-ILD) is unclear. This study aims to identify differentially expressed proteins (DEPs) in CTD-ILD to determine the potential role of these DEPs that may play in the pathogenesis of CTD-ILD and to offer potential therapeutic targets. METHODS: Bronchoalveolar lavage fluid (BALF) samples were collected from four patients with CTD-ILD and four patients without CTD-ILD. Label-free mass spectrometry-based relative quantification was used to identify the DEPs. Bioinformatics were used to determine the potential biological processes and signaling pathways associated with these DEPs. RESULTS: We found 65 upregulated DEPs including SFTPD, CADM1, ACSL4, TSTD1, CD163, LUM, SIGLEC1, CPB2, TGFBI and HGD, and 67 downregulated DEPs including SGSH, WIPF1, SIL1, RAB20, OAS3, GMPR2, PLBD1, DNAJC3, RNASET2 and OAS2. The results of GO functional annotation for the DEPs showed that the DEPS were mainly enriched in the binding, cellular anatomical entity, cellular processes, and biological regulation GO terms. The results of KEGG analyses showed that the pathways most annotated with the DEPs were complement and coagulation cascades, metabolic pathways, pathways in cancer, and PPAR signaling pathway. COG analyses further informed the functions associated with these DEPs, with most focused on signal transduction mechanisms; posttranslational modification, protein turnover, chaperones; intracellular trafficking, secretion, and vesicular transport; amino acid transport and metabolism; and lipid transport and metabolism. CONCLUSIONS: DEPs identified between patients with vs. without CTD-ILD may play important roles in the development of CTD-ILD and are potential new biomarkers for early diagnosis of CTD-ILD.
Assuntos
Doenças do Tecido Conjuntivo , Doenças Pulmonares Intersticiais , Biomarcadores , Líquido da Lavagem Broncoalveolar , Molécula 1 de Adesão Celular , Doenças do Tecido Conjuntivo/complicações , Doenças do Tecido Conjuntivo/diagnóstico , Proteínas do Citoesqueleto , Fatores de Troca do Nucleotídeo Guanina , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Doenças Pulmonares Intersticiais/complicações , Doenças Pulmonares Intersticiais/etiologia , Espectrometria de Massas , Proteínas rab de Ligação ao GTPRESUMO
Small-cell lung cancer (SCLC) is the most aggressive form of lung cancer and the leading cause of global cancer-related mortality. Despite the earlier identification of membrane-proximal cleavage of cell adhesion molecule 1 (CADM1) in cancers, the role of the membrane-bound fragment of CAMD1 (MF-CADM1) is yet to be clearly identified. In this study, we first isolated MF-CADM1-specific fully human single-chain variable fragments (scFvs) from the human synthetic scFv antibody library using the phage display technology. Following the selected scFv conversion to human immunoglobulin G1 (IgG1) scFv-Fc antibodies (K103.1-4), multiple characterization studies, including antibody cross-species reactivity, purity, production yield, and binding affinity, were verified. Finally, via intensive in vitro efficacy and toxicity evaluation studies, we identified K103.3 as a lead antibody that potently promotes the death of human SCLC cell lines, including NCI-H69, NCI-H146, and NCI-H187, by activated Jurkat T cells without severe endothelial toxicity. Taken together, these findings suggest that antibody-based targeting of MF-CADM1 may be an effective strategy to potentiate T cell-mediated SCLC death, and MF-CADM1 may be a novel potential therapeutic target in SCLC for antibody therapy.
Assuntos
Neoplasias Pulmonares , Anticorpos de Cadeia Única , Carcinoma de Pequenas Células do Pulmão , Molécula 1 de Adesão Celular/genética , Técnicas de Visualização da Superfície Celular , Humanos , Anticorpos de Cadeia Única/farmacologiaRESUMO
Cell adhesion molecule 1 (CADM1), which mediates intercellular adhesion between epithelial cells, is shown to be highly expressed in small-cell lung cancer (SCLC) and to enhance tumorigenicity of SCLC cells in nude mice. Here, we investigated the molecular mechanism underlying the oncogenic role of CADM1 in SCLC. CADM1 promoted colony formation of SCLC cells in soft agar. Analysis of deletion and point mutants of the conserved protein-binding motifs in CADM1 revealed that the 4.1 protein-binding motif in the cytoplasmic domain is responsible for the promotion of colony formation. Among the actin-binding 4.1 proteins, 4.1R was the only protein whose localization to the plasma membrane is dependent on CADM1 expression in SCLC cells. Knockdown of 4.1R suppressed the colony formation enhanced by CADM1, suggesting that 4.1R is required for the oncogenic role of CADM1 in SCLC. In primary SCLC, CADM1 expression was correlated with membranous localization of 4.1R, as was observed in a SCLC cell line. Moreover, membranous co-localization of CADM1 and 4.1R was associated with more advanced tumor stage. These results suggest that the formation of CADM1-4.1R complex would promote malignant features of SCLC.