Exosomal miR-19a derived from melanoma cell promotes the vemurafenib resistance of malignant melanoma through directly targeting LRIG1 to reactivate AKT and MAPK pathway.

Exosomes derived from neighboring v-raf murine sarcoma viral oncogene homolog B1 inhibitor (BRAFi)-resistant melanoma cells mediate the formation of resistance in melanoma cells sensitive to BRAFi. The function and molecular mechanisms of exosomal miRNA in BRAFi resistance of melanoma have not been studied. We found that the expression of miR-19a in BRAFi resistant melanoma cells was significantly higher than that in sensitive cells, and miR-19a contributes to the resistance of melanoma cells to BRAFi by targeting immunoglobulin-like domains protein 1 (LRIG1). miR-19a was highly enriched in exosomes secreted from BRAFi resistant melanoma cells, and these exosomal miR-19a promote the spread of BRAFi resistant. The reactivation of Protein kinase B (AKT) and mitogen-activated protein kinase (MAPK) pathways is the main reason for the BRAFi resistant of melanoma cells. We demonstrated that exosomal miR-19a derived from melanoma cell promotes the formation and spread of BRAFi resistant in melanoma through targeting LRIG1 to reactivate AKT and MAPK pathway. Therefore, miR-19a may serve as a potential therapeutic target in melanoma patients with acquired drug resistance.

Immunoreactivity of LMO7 and other molecular markers as potential prognostic factors in oropharyngeal squamous cell carcinoma.

BACKGROUND: Despite the better prognosis associated with human papillomavirus (HPV)-positive oropharyngeal squamous cell carcinoma (OPSCC), some patients experience relapse and succumb to the disease; thus, there is a need for biomarkers identifying these patients for intensified treatment. Leucine-rich repeats and immunoglobulin-like domain (LRIG) protein 1 is a negative regulator of receptor tyrosine kinase signaling and a positive prognostic factor in OPSCC. Studies indicate that LRIG1 interacts with the LIM domain 7 protein (LMO7), a stabilizer of adherence junctions. Its role in OPSCC has not been studied before. METHODS: A total of 145 patients diagnosed with OPSCC were enrolled. Immunohistochemical LMO7 expression and staining intensity were evaluated in the tumors and correlated with known clinical and pathological prognostic factors, such as HPV status and LRIG1, CD44, Ki67, and p53 expression. RESULTS: Our results show that high LMO7 expression is associated with significantly longer overall survival (OS) (p = 0.044). LMO7 was a positive prognostic factor for OS in univariate analysis (HR 0.515, 95% CI: 0.267-0.994, p = 0.048) but not in multivariate analysis. The LMO7 expression correlated with LRIG1 expression (p = 0.048), consistent with previous findings. Interestingly, strong LRIG1 staining intensity was an independent negative prognostic factor in the HPV-driven group of tumors (HR 2.847, 95% Cl: 1.036-7.825, p = 0.043). CONCLUSIONS: We show for the first time that high LMO7 expression is a positive prognostic factor in OPSCC, and we propose that LMO7 should be further explored as a biomarker. In contrast to previous reports, LRIG1 expression was shown to be an independent negative prognostic factor in HPV-driven OPSCC.

Dysregulation and antimetastatic function of circLRIG1 modulated by miR-214-3p/LRIG1 axis in bladder carcinoma.

CircLRIG1, a newly discovered circRNA, has yet to have its potential function and biological processes reported. This study explored the role of circLRIG1 in the development and progression of bladder carcinoma and its potential molecular mechanisms. Techniques such as qRT-PCR, Western blot, various cellular assays, and in vivo models were used to investigate mRNA and protein levels, cell behavior, molecular interactions, and tumor growth. The results showed that both circLRIG1 and LRIG1 were significantly reduced in bladder carcinoma tissues and cell lines. Low circLRIG1 expression was associated with poor patient prognosis. Overexpressing circLRIG1 inhibited bladder carcinoma cell growth, migration, and invasion, promoted apoptosis, and decreased tumor growth and metastasis in vivo. Importantly, circLRIG1 was found to sponge miR-214-3p, enhancing LRIG1 expression, and its overexpression also modulated protein levels of E-cadherin, N-cadherin, Vimentin, and LRIG1. Similar effects were observed with LRIG1 overexpression. Notably, a positive correlation was found between circLRIG1 and LRIG1 expression in bladder carcinoma tissues. Additionally, the tumor-suppressing effect of circLRIG1 was reversed by overexpressing miR-214-3p or silencing LRIG1. The study concludes that circLRIG1 suppresses bladder carcinoma progression by enhancing LRIG1 expression via sponging miR-214-3p, providing a potential strategy for early diagnosis and treatment of bladder carcinoma.

Lrig1 drives cryptogenesis and restrains proliferation during colon development.

Growth and specification of the mouse intestine occurs in utero and concludes after birth. Although numerous studies have examined this developmental process in the small intestine, far less is known about the cellular and molecular cues required for colon development. In this study, we examine the morphological events leading to crypt formation, epithelial cell differentiation, proliferation, and the emergence and expression of a stem and progenitor cell marker Lrig1. Through multicolor lineage tracing, we show Lrig1-expressing cells are present at birth and behave as stem cells to establish clonal crypts within 3 wk of life. In addition, we use an inducible knockout mouse to eliminate Lrig1 and show Lrig1 restrains proliferation within a critical developmental time window, without impacting colonic epithelial cell differentiation. Our study illustrates morphological changes during crypt development and the importance of Lrig1 in the developing colon.NEW & NOTEWORTHY Our studies define the importance of studying Lrig1 in colon development. We address a critical gap in the intestinal development literature and provide new information about the molecular cues that guide colon development. Using a novel, inducible knockout of Lrig1, we show Lrig1 is required for appropriate colon epithelial growth and illustrate the importance of Lrig1-expressing cells in the establishment of colonic crypts.

A member of the immunoglobulin superfamily lrig-1 might be involved in the immune priming of Scylla paramamosain in response to the infection and re-infection by Vibrio parahaemolyticus.

A member of the immunoglobulin superfamily designated leucine-rich repeats and immunoglobulin-like domains protein-1 (lrig-1) encoding a protein with 1109 amino acids with a characteristic IGc2 domain was identified from the transcriptome data of mud crab Scylla paramamosain. Lrig-1 contained: one signaling peptide; one LRR_NT domain; nine LRR domains; three LRR_TYP domains; one LRR_CT domain; three IGc2 regions; one transmembrane region; C-terminal cytoplasmic tail. lrig-1 was widely expressed in all tissues of mud crab and was responsive in hemocytes to first and second Vibrio parahaemolyticus infections. lrig-1 knockdown mediated by RNAi repressed expression of several antimicrobial peptides significantly. Its orthologs in 19 crustacean species were identified and showed high conservation. These results suggest that lrig-1 have a vital role in mud crabs against V. parahaemolyticus infection through expression of multiple antimicrobial peptides. The results obtained in the present study imply the potential roles the lrig-1 played in immune priming in crabs.

Inhibition of the bone morphogenetic protein pathway suppresses tumor growth through downregulation of epidermal growth factor receptor in MEK/ERK-dependent colorectal cancer.

The bone morphogenetic protein (BMP) pathway promotes differentiation and induces apoptosis in normal colorectal epithelial cells. However, its role in colorectal cancer (CRC) is controversial, where it can act as context-dependent tumor promoter or tumor suppressor. Here we have found that CRC cells reside in a BMP-rich environment based on curation of two publicly available RNA-sequencing databases. Suppression of BMP using a specific BMP inhibitor, LDN193189, suppresses the growth of select CRC organoids. Colorectal cancer organoids treated with LDN193189 showed a decrease in epidermal growth factor receptor, which was mediated by protein degradation induced by leucine-rich repeats and immunoglobulin-like domains protein 1 (LRIG1) expression. Among 18 molecularly characterized CRC organoids, suppression of growth by BMP inhibition correlated with induction of LRIG1 gene expression. Notably, knockdown of LRIG1 in organoids diminished the growth-suppressive effect of LDN193189. Furthermore, in CRC organoids, which are susceptible to growth suppression by LDN193189, simultaneous treatment with LDN193189 and trametinib, an FDA-approved MEK inhibitor, resulted in cooperative growth inhibition both in vitro and in vivo. Taken together, the simultaneous inhibition of BMP and MEK could be a novel treatment option in CRC cases, and evaluating in vitro growth suppression and LRIG1 induction by BMP inhibition using patient-derived organoids could offer functional biomarkers for predicting potential responders to this regimen.

Differential miRNA Expression Contributes to Emergence of Multiple Cancer Stem Cell Subpopulations in Human Colorectal Cancer.

One reason for lack of efficacy in cancer therapeutics is tumor heterogeneity. We hypothesize that tumor heterogeneity arises due to emergence of multiple cancer stem cell (CSC) subpopulations because miRNAs regulate expression of stem cell genes in CSCs. Our goal was to determine if: i) multiple CSC subpopulations exist in a human CRC cell population, and ii) miRNAs are differentially expressed in the different CSC subpopulations. We discovered that at least four different CSC populations (ALDH1, CD166, LGR5, LRIG1) exist in the HT29 cell line. CSC subpopulations were quantified using co-staining for multiple stem cell markers, isolated using FACS, and analyzed by NanoString miRNA profiling. The miRNA expression pattern in each CSC subpopulation was analyzed relative to miRNA expression patterns in other CSC subpopulations. Messenger RNAs predicted to be targeted by the upregulated miRNAs in each CSC subpopulation were: 1) identified using bioinformatics analyses, and 2) classified according to their predicted functions using David functional annotation analyses. We found multiple CSC subpopulations with a unique miRNA signature in each CSC subpopulation. Notably, the miRNAs expressed within one CSC subpopulation are predicted to target and downregulate the CSC genes and pathways that establish the other CSC subpopulations. Moreover, mRNAs predicted to be targeted by miRNAs in the different CSC subpopulations have different cellular functional classifications. That different CSC subpopulations express miRNAs that are predicted to target CSC genes expressed in other CSC subpopulations provides a mechanism that might explain the co-existence of multiple CSC subpopulations, tumor heterogeneity, and cancer therapy resistance.

Lrig1 expression identifies quiescent stem cells in the ventricular-subventricular zone from postnatal development to adulthood and limits their persistent hyperproliferation.

BACKGROUND: We previously identified Leucine-rich repeats and immunoglobulin-like domains 1 (Lrig1) as a marker of long-term neurogenic stem cells in the lateral wall of the adult mouse brain. The morphology of the stem cells thus identified differed from the canonical B1 type stem cells, raising a question about their cellular origin. Thus, we investigated the development of these stem cells in the postnatal and juvenile brain. Furthermore, because Lrig1 is a known regulator of quiescence, we also investigated the effect(s) of its deletion on the cellular proliferation in the lateral wall. METHODS: To observe the development of the Lrig1-lineage stem cells, genetic inducible fate mapping studies in combination with thymidine analog administration were conducted using a previously published Lrig1T2A-iCreERT2 mouse line. To identify the long-term consequence(s) of Lrig1 germline deletion, old Lrig1 knock-out mice were generated using two different Lrig1 null alleles in the C57BL/6J background. The lateral walls from these mice were analyzed using an optimized whole mount immunofluorescence protocol and confocal microscopy. RESULTS: We observed the Lrig1-lineage labeled cells with morphologies consistent with neurogenic stem cell identity in postnatal, juvenile, and adult mouse brains. Interestingly, when induced at postnatal or juvenile ages, morphologically distinct cells were revealed, including cells with the canonical B1 type stem cell morphology. Almost all of the presumptive stem cells labeled were non-proliferative at these ages. In the old Lrig1 germline knock-out mice, increased proliferation was observed compared to wildtype littermates without concomitant increase in apoptosis. CONCLUSIONS: Once set aside during embryogenesis, the Lrig1-lineage stem cells remain largely quiescent during postnatal and juvenile development until activation in adult age. The absence of premature proliferative exhaustion in the Lrig1 knock-out stem cell niche during aging is likely due to a complex cascade of effects on the adult stem cell pool. Thus, we suggest that the adult stem cell pool size may be genetically constrained via Lrig1.

HYPOTHESIS: Do LRIG Proteins Regulate Stem Cell Quiescence by Promoting BMP Signaling?

Leucine-rich repeats and immunoglobulin-like domains (LRIG) proteins are evolutionarily conserved integral membrane proteins. Mammalian LRIG1 regulates stem cell quiescence in various tissue compartments, including compartments in the epidermis, oral mucosa, intestines, neural system, and incisors. The planarian LRIG1 homolog regulates the quiescence of multipotent neoblasts. The mechanism through which LRIG proteins regulate stem cell quiescence has not been well documented, although it is generally assumed that LRIG1 regulates the epidermal growth factor receptor (EGFR) or other receptor tyrosine kinases. However, Lrig-null (Lrig1-/-;Lrig2-/-; and Lrig3-/-) mouse embryonic fibroblasts (MEFs) have been recently found to exhibit apparently normal receptor tyrosine kinase functions. Moreover, bone morphogenetic protein (BMP) signaling has been shown to depend on LRIG1 and LRIG3 expression. BMPs are well-known regulators of stem cell quiescence. Here, we hypothesize that LRIG1 might regulate stem cell quiescence by promoting BMP signaling. HYPOTHESIS: Based on recent findings, it is hypothesized that LRIG1 regulates stem cell quiescence in mammalian tissues as well as in planarian neoblasts by promoting BMP signaling.

Differential miRNA Expression Contributes to Emergence of Multiple Cancer Stem Cell Subpopulations in Human Colorectal Cancer.

One reason for lack of efficacy in cancer therapeutics is tumor heterogeneity. We hypothesize that tumor heterogeneity arises due to emergence of multiple Cancer Stem Cell (CSC) subpopulations because miRNAs regulate expression of stem cell genes in CSCs. Our goal was to determine if: i) multiple CSC subpopulations exist in a human CRC cell population, and ii) miRNAs are differentially expressed in the different CSC subpopulations. We discovered that at least four different CSC populations (ALDH1, CD166, LGR5, and LRIG1) exist in the HT29 cell line. CSC subpopulations were quantified using co-staining for multiple stem cell markers, isolated using FACS, and analyzed by NanoString miRNA profiling. The miRNA expression pattern in each CSC subpopulation was analyzed relative to miRNA expression patterns in other CSC subpopulations. Messenger RNAs predicted to be targeted by the up-regulated miRNAs in each CSC subpopulation were: 1) identified using bioinformatics analyses, and 2) classified according to their predicted functions using David functional annotation analyses. We found multiple CSC subpopulations with a unique miRNA signature in each CSC subpopulation. Notably, the miRNAs expressed within one CSC subpopulation are predicted to target and down-regulate the CSC genes and pathways that establish the other CSC subpopulations. Moreover, mRNAs predicted to be targeted by miRNAs in the different CSC subpopulations have different cellular functional classifications. That different CSC subpopulations express miRNAs that are predicted to target CSC genes expressed in other CSC subpopulations provides a mechanism that might explain the co-existence of multiple CSC subpopulations, tumor heterogeneity, and cancer therapy resistance.

MicroRNA-218-5p accelerates malignant behaviors of breast cancer through LRIG1

Abstract Objective: MicroRNAs play crucial roles in the pathogenesis of cancers. MiRNA-218-5p may act as either an oncogene or a tumor suppressor, but its role in the pathogenesis of Breast Cancer (BC) remains unclear. Methods: Infiltrative breast ductal carcinoma as well as corresponding adjacent normal samples were collected from 30 patients. Mimics and inhibitors of miRNA-218-5p or corresponding negative controls were transfected into BC cells. miRNA-218-5p expression was detected by quantitative PCR. The effects of miRNA-218-5p on the malignant behaviors of BC were assessed. Dual-luciferase reporter assay was employed to evaluate the binding of miRNA-218-5p to LRIG1. Results: BC tissues showed higher miRNA-218-5p expression as compared to the adjacent normal tissues. Ectopic miRNA-218-5p expression accelerated the cell cycle, cell growth and migration of BC, while repressed cell apoptosis. Interestingly, ectopic miRNA-218-5p expression down-regulated LRIG1 expression, and miRNA-218-5p could bind to LRIG1. Also, our study indicated that miRNA-218-5p up-regulated ErbB2 and EGFR expression by targeting LRIG1, suggesting that the LRIG1-mediated signaling pathway contributed to the pro-tumor effects of miRNA-218-5p on BC. Conclusion: MiRNA-218-5p up-regulates ErbB2 and EGFR expression by suppressing LRIG1 expression, thus promoting the malignant behaviors of BC. miRNA-218-5p may exert a pro-tumor effect on BC and serve as a therapeutic target for BC treatment.

Single-molecule array assay reveals the prognostic impact of plasma LRIG1 in ovarian carcinoma.

BACKGROUND: Ovarian carcinoma is the eighth most common cause of cancer death in women worldwide. The disease is predominantly diagnosed at a late stage. This contributes to high recurrence rates, eventually leading to the development of treatment-resistant disease. Leucine-rich repeats and immunoglobulin-like domains protein 1 (LRIG1) is a transmembrane protein that functions as a tumor suppressor and regulator of growth factor signaling. LRIG1 levels have not been investigated in human plasma previously. MATERIALS AND METHODS: A quantitative LRIG1-specific single molecule array assay was developed and validated. LRIG1 levels were quantified in plasma samples from 486 patients with suspicious ovarian masses. RESULTS: Among women with ovarian carcinoma, LRIG1 levels were significantly elevated compared to women with benign or borderline type tumors. High LRIG1 plasma levels were associated with worse overall survival and shorter disease-free survival both in the group of all malignant cases and among the stage 3 cases only. LRIG1 was an independent prognostic factor in patients with stage 3 ovarian carcinoma. CONCLUSION: LRIG1 plasma levels were elevated in patients with ovarian carcinoma, and high levels were associated with poor prognosis, suggesting that LRIG1 might be an etiologic factor and a potentially useful biomarker in ovarian carcinoma.

Lrig1 regulates the balance between proliferation and quiescence in glioblastoma stem cells.

Patients with glioblastoma (GBM) face a dismal prognosis. GBMs are driven by glioblastoma stem cells (GSCs) that display a neural stem cell (NSC)-like phenotype. These glioblastoma stem cells are often in a quiescent state that evades current therapies, namely debulking surgery and chemo/radiotherapy. Leucine-rich repeats and immunoglobulin-like domains (LRIG) proteins have been implicated as regulators of growth factor signalling across many tissue stem cells. Lrig1 is highly expressed in gliomas and importantly, polymorphisms have been identified that are risk alleles for patients with GBM, which suggests some functional role in gliomagenesis. We previously reported that Lrig1 is a gatekeeper of quiescence exit in adult mouse neural stem cells, suppressing epidermal growth factor receptor signalling prior to cell cycle re-entry. Here, we perform gain- and loss-of-function studies to understand the function of Lrig1 in glioblastoma stem cells. Using a novel mouse glioblastoma stem cell model, we show that genetic ablation of Lrig1 in cultured GBM stem cells results in higher proliferation and loss of quiescence. In vivo, mice transplanted with glioblastoma stem cells lacking Lrig1 display lower survival compared to Lrig1 WT glioblastoma stem cells, with tumours displaying increased proportions of proliferative cells and reduced quiescent subpopulations. In contrast, Lrig1 overexpression in mouse glioblastoma stem cells results in enhanced quiescence and reduced proliferation, with impaired tumour formation upon orthotopic transplantation. Mechanistically, we find that Lrig1-null cells have a deficiency in BMP signalling responses that may underlie their lack of responsiveness to quiescence cues in vivo. These findings highlight important roles for Lrig1 in controlling responsiveness to both epidermal growth factor receptor and BMPR signalling, and hence the proportions of quiescent and proliferative subpopulations in GBMs.

The up-regulation of LRIG1 expression inhibits the proliferation, apoptosis and invasion of glioma cells.

OBJECTIVE: To illustrate the role of LRIG1 in regulating the Notch signaling pathway and glioma cell proliferation, apoptosis and invasion. METHODS: The glioma cells (U373) were divided into control group, NC group and LRIG1 group. After transfection, the CCK-8 assay, Transwell assay, and Flow cytometry were used to explore the biological function of LRIG1 in glioma cells. At the end, Western blot was used to detect the expression of LRIG1, Notch1, Hes1, Bcl-2, and Bax. RESULTS: The LRIG1 expression in U373 cells was remarkably lower than that in normal glial cells (P=0.019). The LRIG1 expression in the LRIG1 group was successfully increased when compared with that in the control group (P=0.004). The cell viability of the LRIG1 group was significantly lower than that of the NC group and control group at 24 h, 48 h, and 72 h (P=0.040, 0.025; P=0.041, 0.041; P=0.035, 0.035) respectively. Increased LRIG1 expression level in glioma cells strongly inhibits cell migration in transwell experiment. Flow cytometry results indicated that the apoptosis rate of the LRIG1 group was critically higher than that of the NC group and control group (P=0.003; P=0.003). According to results of Western blot, the expression levels of Notch1, Hes1, Hes5, and Jagged1 in LRIG1 group were dramatically higher than that in NC group and control group (P=0.006, 0.013; P=0.025, 0.026; P=0.001, 0.004; P=0.025, 0.027; P=0.029, 0.021) reespectively. While Bax expression in LRIG1 group was lower than that of NC group and control group (P=0.018, 0.021). CONCLUSION: The up-regulation of LRIG1 can inhibit the proliferation and migration of glioma cells and promote apoptosis by regulating the Notch signaling pathway.

Astrocyte-neuron crosstalk through Hedgehog signaling mediates cortical synapse development.

Neuron-glia interactions play a critical role in the regulation of synapse formation and circuit assembly. Here we demonstrate that canonical Sonic hedgehog (Shh) pathway signaling in cortical astrocytes acts to coordinate layer-specific synaptic connectivity. We show that the Shh receptor Ptch1 is expressed by cortical astrocytes during development and that Shh signaling is necessary and sufficient to promote the expression of genes involved in regulating synaptic development and layer-enriched astrocyte molecular identity. Loss of Shh in layer V neurons reduces astrocyte complexity and coverage by astrocytic processes in tripartite synapses; conversely, cell-autonomous activation of Shh signaling in astrocytes promotes cortical excitatory synapse formation. Furthermore, Shh-dependent genes Lrig1 and Sparc distinctively contribute to astrocyte morphology and synapse formation. Together, these results suggest that Shh secreted from deep-layer cortical neurons acts to specialize the molecular and functional features of astrocytes during development to shape circuit assembly and function.

Basal Cells in the Epidermis and Epidermal Differentiation.

A definite identification of epidermal stem cells is not known and the mechanism of epidermal differentiation is not fully understood. Toward both of these quests, considerable information is available from the research on lineage tracing and clonal growth analysis in the basal layer of the epidermis, on the hair follicle and the interfollicular epidermal stem cells, and on Wnt signaling along with its role in the developmental patterning and cell differentiation. In this paper, literature on the aforementioned research has been collated and analyzed. In addition, models of the basal layer cellular composition and the epidermal differentiation have been presented. Graphical Abstract.

LRIG1, a regulator of stem cell quiescence and a pleiotropic feedback tumor suppressor.

LRIG1, leucine-rich repeats and immunoglobulin-like domains protein 1, was discovered more than 20 years ago and has been shown to be downregulated or lost, and to function as a tumor suppressor in several cancers. Another well-reported biological function of LRIG1 is to regulate and help enforce the quiescence of adult stem cells (SCs). In both contexts, LRIG1 regulates SC quiescence and represses tumor growth via, primarily, antagonizing the expression and activities of ERBB and other receptor tyrosine kinases (RTKs). We have recently reported that in treatment-naïve human prostate cancer (PCa), LRIG1 is primarily regulated by androgen receptor (AR) and is prominently overexpressed. In castration-resistant PCa (CRPC), both LRIG1 and AR expression becomes heterogeneous and, frequently, discordant. Importantly, in both androgen-dependent PCa and CRPC models, LRIG1 exhibits tumor-suppressive functions. Moreover, LRIG1 induction inhibits the growth of pre-established AR+ and AR- PCa. Here, upon a brief introduction of the LRIG1 and the LRIG family, we provide an updated overview on LRIG1 functions in regulating SC quiescence and repressing tumor development. We further highlight the expression, regulation and functions of LRIG1 in treatment-naïve PCa and CRPC. We conclude by offering the perspectives of identifying novel cancer-specific LRIG1-interacting signaling partners and developing LRIG1-based anti-cancer therapeutics and diagnostic/prognostic biomarkers.

Aryl Hydrocarbon Receptor Defect Attenuates Mitogen-Activated Signaling through Leucine-Rich Repeats and Immunoglobulin-like Domains 1 (LRIG1)-Dependent EGFR Degradation.

Aryl hydrocarbon receptor (AHR) genomic pathway has been well-characterized in a number of respiratory diseases. In addition, the cytoplasmic AHR protein may act as an adaptor of E3 ubiquitin ligase. In this study, the physiological functions of AHR that regulate cell proliferation were explored using the CRISPR/Cas9 system. The doubling-time of the AHR-KO clones of A549 and BEAS-2B was observed to be prolonged. The attenuation of proliferation potential was strongly associated with either the induction of p27Kip1 or the impairment in mitogenic signal transduction driven by the epidermal growth factor (EGF) and EGF receptor (EGFR). We found that the leucine-rich repeats and immunoglobulin-like domains 1 (LRIG1), a repressor of EGFR, was induced in the absence of AHR in vitro and in vivo. The LRIG1 tends to degrade via a proteasome dependent manner by interacting with AHR in wild-type cells. Either LRIG1 or a disintegrin and metalloprotease 17 (ADAM17) were accumulated in AHR-defective cells, consequently accelerating the degradation of EGFR, and attenuating the response to mitogenic stimulation. We also affirmed low AHR but high LRIG1 levels in lung tissues of chronic obstructive pulmonary disease (COPD) patients. This might partially elucidate the sluggish tissue repairment and developing inflammation in COPD patients.

hsa_circ_0000006 induces tumorigenesis through miR-361-3p targeting immunoglobulin-like domains protein 1 (LRIG1) in osteosarcoma.

BACKGROUND: Osteosarcoma (OS) is considered to be the most highly prevalent bone tumor. In the progression of different human cancers, the role of circular RNAs (circRNAs) has been extensively studied. Microarray analysis has indicated that hsa_circ_0000006 expression was lower in OS, but the mechanism of hsa_circ_0000006 in regulating the progression of OS remains elusive. METHODS: The expression of cancer-related genes at the transcriptional and translational levels was assessed by RT-qPCR and western blotting (WB). Colony formation and Cell Counting Kit-8 (CCK-8) assays were used to evaluate the proliferative potential of cells. The transwell assay was used to examine the invasive and migratory potential of cells. Furthermore, dual-luciferase reporter (DLR) and RNA pull-down assays were performed for the validation of the targeting sites of hsa_circ_0000006, miR-361-3p, and the 3'-untranslated region (3'-UTR) of immunoglobulin-like domains protein 1 (LRIG1) mRNA. Moreover, the protein levels of epithelial-to-mesenchymal transition (EMT) markers were analyzed by WB. RESULTS: The expression of hsa_circ_0000006 and LRIG1 were found to be down-regulated in OS tissues and cells, while miR-361-3p was up-regulated. Knockdown of hsa_circ_0000006 promoted the progression and development of OS, as well as EMT. Furthermore, hsa_circ_0000006 was revealed as a sponge of miR-361-3p, which negatively regulates miR-361-3p expression. LRIG1 was found to be an miR-361-3p target. In OS cells, the LRIG1 expression level was decreased, with elevated expression of miR-361-3p. Advanced studies demonstrated that hsa_circ_0000006 regulates LRIG1 expression through sponging miR-361-3p, then promotes the tumorigenesis of OS. CONCLUSIONS: hsa_circ_0000006 is associated with the progression and development of OS through miR-361-3p by target LRIG1, which is a significant biomarker and effective therapeutic target for patients with OS.

LRIG1 is a positive prognostic marker in Merkel cell carcinoma and Merkel cell carcinoma expresses epithelial stem cell markers.

Merkel cell carcinoma (MCC) is a rare and aggressive neuroendocrine malignancy of the skin. The cell of origin of MCC is thus far unknown and proposed cells of origin include Merkel cells, pro-/pre- or pre-B cells, epithelial stem cells, and dermal stem cells. In this study, we aimed to shed further light on the possibility that a subset of MCC tumors arise from epithelial stem cells of the skin by examining the expression of hair follicle and epidermal stem cell markers in MCC and normal human skin. We also aimed to elucidate any correlation between the expression of these markers and tumor Merkel cell polyomavirus (MCPyV) status or other clinicopathological characteristics or patient survival. Expression of CK19, SOX9, LGR5, and LRIG1 in MCC and normal human skin was studied by immunohistochemistry, and the staining patterns or intensities were statistically correlated with patient, tumor, MCPyV, and survival parameters. In a cohort of 137 cases of MCC, we observed dot-like immunoexpression of CK19 in 30 cases (22.1%) and homogeneous expression in 103 cases (75.7%). We also observed positive immunoexpression of SOX9 in 21 cases (15.3%), LGR5 in 118 cases (86.1%), and LRIG1 in 117 cases (86.0%). Immunoexpression of LRIG1 was found to correlate with better overall and MCC-specific survival. We observed frequent immunoexpression of several hair follicle and epidermal stem cell markers in MCC and found LRIG1 to be a positive prognostic marker in MCC.