Promotes Melanoma Cell Proliferation Research Articles

H2A.Z chaperones converge on E2F target genes for melanoma cell proliferation.

High levels of H2A.Z promote melanoma cell proliferation and correlate with poor prognosis. However, the role of the two distinct H2A.Z histone chaperone complexes SRCAP and P400-TIP60 in melanoma remains unclear. Here, we show that individual subunit depletion of SRCAP, P400, and VPS72 (YL1) results in not only the loss of H2A.Z deposition into chromatin but also a reduction of H4 acetylation in melanoma cells. This loss of H4 acetylation is particularly found at the promoters of cell cycle genes directly bound by H2A.Z and its chaperones, suggesting a coordinated regulation between H2A.Z deposition and H4 acetylation to promote their expression. Knockdown of each of the three subunits downregulates E2F1 and its targets, resulting in a cell cycle arrest akin to H2A.Z depletion. However, unlike H2A.Z deficiency, loss of the shared H2A.Z chaperone subunit YL1 induces apoptosis. Furthermore, YL1 is overexpressed in melanoma tissues, and its upregulation is associated with poor patient outcome. Together, these findings provide a rationale for future targeting of H2A.Z chaperones as an epigenetic strategy for melanoma treatment.

Abstract 296: BRAFi-induced ROCK-mediated non-canonical nuclear β-catenin shuttling drives a phenotypic switch in cancer-associated fibroblasts

Abstract The discovery of BRAF mutations in around 60% of melanoma patients and the subsequent development of targeted inhibitors to counter the hyperactive BRAF-driven pro-tumorigenic signaling pathway have marked a significant milestone in melanoma therapy. However, despite this progress, the long-term outcomes remain unsatisfactory for many patients due to the occurrence of drug resistance. We previously reported that cancer-associated fibroblasts (CAFs) are stimulated by BRAF inhibitors (BRAFi) to drive matrix remodeling and therapeutic escape pathways in melanoma cells. However, the underlying mechanism by which CAFs are reprogrammed by BRAFi to obtain enhanced phenotypes remains elusive. Here, we present compelling evidence showing that BRAFi (PLX4032), but not CRAF inhibitor (GW5074) or pan-RAF inhibitor (RAF709), induces the accumulation of nuclear β-catenin in CAFs harboring wild-type BRAF, a process mediated by BRAF and CRAF but not ARAF isoform. BRAFi binding to BRAF and CRAF leads to robust RAF kinase dimerization, including BRAF and CRAF homodimer and BRAF: CRAF heterodimer. RAF dimers are subsequently recruited to the plasma membrane, where BRAF and CRAF are phosphorylated on residues Thr599 and Ser338, respectively, and activated in a RAS-GTP-dependent manner. RAF activation stimulates the downstream Rho kinase (ROCK) and MEK/ERK signaling pathways simultaneously. Notably, ablating RAS and RAF isoforms effectively suppressed BRAFi-induced activation of ROCK1/2 signaling in CAFs, further confirming that the ROCK pathway is a downstream effector of BRAFi-driven RAF activation and is intricately linked to increased nuclear β-catenin in CAFs. The pharmacological blockade of ROCK activity, using HA-1077, diminished BRAFi-induced nuclear β-catenin accumulation in CAFs, suggesting BRAFi-driven cytoskeletal remodeling plays a pivotal role in the nuclear shuttling of β-catenin. Using a mouse model that mimics human BRAF-mutant melanoma, we found that elevated level of nuclear β-catenin drives the conversion of fibroblasts into α-SMA-positive CAFs, leading to elevated collagen deposition and contributing to melanoma progression in vivo. In summary, BRAFi reprograms the transcriptional activity in CAFs by driving increased nuclear β-catenin to increase their ability to remodel the tumor ECM and promote melanoma cell proliferation. Collectively, the data offer new insights into the molecular mechanisms that underlie the paradoxical reprogramming of CAFs by BRAFi in melanoma therapy through the non-canonical β-catenin pathway. Citation Format: Bruna da Silva Soley, Radhika Athalye, Ryan Zhang, Thomas Andl, Yuhang Zhang. BRAFi-induced ROCK-mediated non-canonical nuclear β-catenin shuttling drives a phenotypic switch in cancer-associated fibroblasts [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 296.

Circular RNA circFCHO2(hsa_circ_0002490) promotes the proliferation of melanoma by directly binding to DND1

Circular RNAs (circRNAs) have been documented to play crucial roles in the biology of various cancers. However, their investigation in melanoma is still at an early stage, particularly as a broader mechanism beyond acting as miRNA sponges needs to be explored. We report here that circFCHO2(hsa_circ_0002490), a circRNA encompassing exons 19 and 20 of the FCHO2 gene, exhibited a consistent overexpression in melanoma tissues. Furthermore, elevated circFCHO2 levels demonstrated a positive correlation with the malignant phenotype and poor prognosis among the 158 melanoma patients studied. Besides, we observed that heightened levels of circFCHO2 promoted melanoma cell proliferation, migration, and invasion in vitro, along with contributing to tumor growth in vivo. Furthermore, we found differences in the secondary structure of circFCHO2 compared to most other circular RNA structures. It has fewer miRNA binding sites, while it has more RNA binding protein binding sites. We therefore speculate that circFCHO2 may have a function of interacting with RNA binding proteins. Mechanistically, it was confirmed by fluorescence in situ hybridization (FISH), RNA-pull down, RNA immunoprecipitation (RIP), and western blotting assays that circFCHO2 interacts with dead end protein homolog 1 (DND1) and reverses the inhibition of the PI3K/AKT signaling pathway by binding to DND1. Our findings reveal that circFCHO2 drives melanoma progression by regulating the PI3K/AKT signaling pathway through direct binding to DND1 and may serve as a potential diagnostic biomarker and therapeutic target for the treatment of melanoma.Graphical Headlights: CircFCHO2 is highly expressed in melanoma. High circFCHO2 levels were positively correlated with poor prognosis in 158 melanoma patients. CircFCHO2 is involved in the regulation of the PI3K/AKT signalling pathway by binding to DND1. CircFCHO2 could serve as a potential biomarker and therapeutic target for the management of melanoma.

ALKBH5 facilitates the progression of skin cutaneous melanoma via mediating ABCA1 demethylation and modulating autophagy in an m6A-dependent manner.

Background: N6-methyladenosine (m6A) is the most common and abundant mRNA modification, playing an essential role in biological processes and tumor development. However, the role of m6A methylation in skin cutaneous melanoma (SKCM) is not yet clear. This study analyzed the expression of m6A-related functional genes in SKCM and aimed to explore the key demethylase ALKBH5 mediated m6A modification and its potential mechanism in human SKCM. Methods: Based on public databases, the m6A-related gene expression landscape in SKCM was portrayed. MeRIP-Seq and RNA-Seq were used to recognize the downstream target of ALKBH5. In vivo and in vitro functional phenotype and rescue functional experiments were performed to explore the mechanism of the ALKBH5-m6A-ABCA1 axis in SKCM. Results: We found ALKBH5 upregulated in SKCM, associated with poor prognosis. ALKBH5 can promote melanoma cell proliferation, colony formation, migration, and invasion and inhibit autophagy in vitro, facilitating tumor growth and metastasis in vivo. We identified ABCA1, a membrane protein that assists cholesterol efflux, as a downstream target of ALKBH5-mediated m6A demethylation. Finally, our data demonstrated that ALKBH5 promoted SKCM via mediating ABCA1 downregulation by reducing ABCA1 mRNA stability in an m6A-dependent manner. Conclusion: Our findings exhibited the functional value of the key demethylase ALKBH5 mediated m6A modification in the progression of SKCM, suggesting the ALKBH5-m6A-ABCA1 axis as a potential therapeutic target in SKCM.

LINC00662 promotes melanoma progression by competitively binding miR-107 and activating the β-catenin signaling pathway.

Melanoma is a highly malignant tumor in the body. Long non-coding RNAs (lncRNAs) have been reported to be involved in the development of various tumors. Emerging evidence demonstrates the critical role of lncRNAs in melanoma development. In this study, we aimed to investigate the expression, biological function and regulatory mechanism of LINC00662 in melanomas. First, we found that LINC00662 was up-regulated in melanoma tissues and cell lines. High expression of LINC00662 in melanomas was associated with a poor patient prognosis. Silencing of LINC00662 suppressed the proliferation, migration, and invasion of melanoma cells in vitro and in vivo, while overexpression of LINC00662 promoted melanoma cell proliferation in vitro. Bioinformatics analysis, dual-luciferase assay, and RIP assay confirmed that LINC00662 competitively regulated miR-107. Silencing of LINC00662 upregulated miR-107 expression in a melanoma cell line. Inhibition of miR-107 significantly reversed the inhibitory effect of LINC00662 silencing on cell proliferation and migration. Furthermore, POU3F2 was validated as a downstream target of LINC00662/miR107 and was downregulated when LINC00662 was silenced. Overexpressing POU3F2 attenuated the effect of si-LINC00662 on cellular functions. In addition, the results also showed that the β-catenin pathway was involved in a si-LINC00662-induced function in melanoma. Overall, our results confirmed that LINC00662 promoted melanoma progression by sponging miR107 and inducing POU3F2, highlighting the mechanism of the LINC00662/miR-107/POU3F2 axis in melanoma cell proliferation and invasion.

Super-enhancer-driven expression of BAHCC1 promotes melanoma cell proliferation and genome stability

Super-enhancers (SEs) are stretches of enhancers ensuring a high level of expression of key genes associated with cell function. The identification of cancer-specific SE-driven genes is a powerful means for the development of innovative therapeutic strategies. Here, we identify a MITF/SOX10/TFIIH-dependent SE promoting the expression of BAHCC1 in a broad panel of melanoma cells. BAHCC1 is highly expressed in metastatic melanoma and is required for tumor engraftment, growth, and dissemination. Integrative genomics analyses reveal that BAHCC1 is a transcriptional regulator controlling expression of E2F/KLF-dependent cell-cycle and DNA-repair genes. BAHCC1 associates with BRG1-containing remodeling complexes at the promoters of these genes. BAHCC1 silencing leads to decreased cell proliferation and delayed DNA repair. Consequently, BAHCC1 deficiency cooperates with PARP inhibition to induce melanoma cell death. Our study identifies BAHCC1 as an SE-driven gene expressed in melanoma and demonstrates how its inhibition can be exploited as a therapeutic target.

SOX10 Loss Sensitizes Melanoma Cells to Cytokine-Mediated Inflammatory Cell Death.

These data support a role for SOX10 expression altering the response to T cell-mediated cell death and contribute to a broader understanding of the interaction between immune cells and melanoma cells.

CSTF2T up-regulates IGHG1 by binding to ZEB1 to promote melanoma cell proliferation, migration, and invasion

ObjectivesMelanoma progression involves multiple molecular pathways. In this study, we explored the effect of immunoglobulin heavy constant gamma 1 (IGHG1) on melanoma progression. MethodsIGHG1, zinc finger E-box binding homeobox 1 (ZEB1), and cleavage stimulation factor subunit 2 tau variant (CSTF2T) expression levels were measured by quantitative reverse-transcription polymerase chain reaction and western blot. BALB/c nude mice were subcutaneously injected with A375 cells to develop a transplantation tumor model, 4 weeks after which the tumor tissues were collected. Cell proliferation, cell invasion, and cell migration of A375 and SK-MEL-28 cells were measured after gain- and loss-of-function of IGHG1, ZEB1, and CSTF2T. Binding of ZEB1 to the IGHG1 promoter was assayed by chromatin immunoprecipitation and dual-luciferase reporter gene assays, while binding of CSTF2T to ZEB1 mRNA was investigated by RNA immunoprecipitation and RNA pull-down assays. ResultsIGHG1, ZEB1, and CSTF2T were all highly expressed in human melanoma cell lines. In A375 and SK-MEL-28 cells, ZEB1 binds directly to the IGHG1 promoter. ZEB1 silencing reduced IGHG1 expression and melanoma cell proliferative, invasive, and migratory properties, and these cellular effects were nullified by overexpression of IGHG1. CSTF2T binds directly to ZEB1 mRNA. Silencing of CSTF2T diminished ZEB1 expression and restrained melanoma cell proliferative, invasive, and migratory capabilities. The impacts of CSTF2T silencing on melanoma cells were counteracted by concomitant overexpression of ZEB1. CSTF2T knockdown reduced tumor volume and weight in nude mice. ConclusionsThe CSTF2T–ZEB1–IGHG1 axis promotes melanoma cell proliferation and invasion.

SOX10 modulated SMARCA4 dysregulation alleviates DNA replication stress in cutaneous melanoma.

Cutaneous melanoma (CM) is the most fatal type of skin cancer with a high potency of metastasis, yet the treatment for metastatic melanoma remains limited. In this study, we are devoted to addressing the prognostic value and underlying mechanism of DNA damage repair-related genes in CM. We utilized integrated bioinformatic approaches and machine learning models to identify a cluster of convergently expressed DNA damage repair-related genes in melanoma. With multivariate Cox regression, SMARCA4 (also known as BRG1) was identified as an independent prognostic marker for melanoma patients. Yet the expression of SMARCA4 is not altered with the pathological staging or the metastasis condition. SMARCA4 is an essential ATPase subunit of the mammalian SWI/SNF complex. Mechanistically, we demonstrated that SMARCA4 could resolve DNA replication stress and guarantee the proliferation of melanoma cells. Furthermore, we predicted the binding of different transcription factors on the SMARCA4 promoter and unveiled the modulated expression of SMARCA4 by SOX10 in melanoma. Together, we performed integrated approaches to identify SMARCA4 as a promising prognostic marker for melanoma, which was transcriptionally regulated by SOX10 and promoted melanoma cell proliferation by ameliorating DNA replication stress.

Promotion of Melanoma Cell Proliferation by Cyclic Straining through Regulatory Morphogenesis.

The genotype and phenotype of acral melanoma are obviously different from UV-radiation-induced melanoma. Based on the clinical data, mechanical stimulation is believed to be a potential cause of acral melanoma. In this case, it is desirable to clarify the role of mechanical stimulation in the progression of acral melanoma. However, the pathological process of cyclic straining that stimulates acral melanoma is still unclear. In this study, the influence of cyclic straining on melanoma cell proliferation was analyzed by using a specifically designed cell culture system. In the results, cyclic straining could promote melanoma cell proliferation but was inefficient after the disruption of cytoskeleton organization. Therefore, the mechanotransduction mechanism of promoted proliferation was explored. Both myosin and actin polymerization were demonstrated to be related to cyclic straining and further influenced the morphogenesis of melanoma cells. Additionally, the activation of mechanosensing transcription factor YAP was related to regulatory morphogenesis. Furthermore, expression levels of melanoma-involved genes were regulated by cyclic straining and, finally, accelerated DNA synthesis. The results of this study will provide supplementary information for the understanding of acral melanoma.

RSK2 promotes melanoma cell proliferation and vemurafenib resistance via upregulating cyclin D1.

BRAF inhibitors are commonly used in targeted therapies for melanoma patients harboring BRAFV600E mutant. Despite the benefit of vemurafenib therapy, acquired resistance during or after treatment remains a major obstacle in BRAFV600E mutant melanoma. Here we found that RSK2 is overexpressed in melanoma cells and the high expression of RSK2 indicates poor overall survival (OS) in melanoma patients. Overexpression of RSK2 leads to vemurafenib resistance, and the deletion of RSK2 inhibits cell proliferation and sensitizes melanoma cells to vemurafenib. Mechanistically, RSK2 enhances the phosphorylation of FOXO1 by interacting with FOXO1 and promoting its subsequent degradation, leading to upregulation of cyclin D1 in melanoma cells. These results not only reveal the presence of a RSK2-FOXO1-cyclin D1 signaling pathway in melanoma, but also provide a potential therapeutic strategy to enhance the efficacy of vemurafenib against cancer.

Microphthalmia Transcription Factor almost Thirty Years after: Its Role in Melanogenesis and its Plant-Derived Inhibitors

It has been almost thirty years since the first publication on microphthalmia-associated transcription factor (MITF) in 1993. MITF, which plays an important role in the melanogenesis process, is an interesting target for melanoma therapy, due to its associates with melanoma survival. MITF promotes melanoma cell proliferation, whereas the sustained suppression of MITF expression causes aging. MITF contributes to differentiation, which involves breaking out of the cell cycle and triggering a melanogenesis, and this function appears to often persist during melanoma development given the frequently observed high pigmented lesions, even in the late stages of melanoma. Several drugs that could inhibit MITF e.g. histone deacetylase inhibitors, such as sodium butyrate and trichostatin A, have been proven could suppress M-MITF expression in melanoma cells. H1-receptor antagonists, particularly loratadine, could downregulate MITF and tyrosinase in melanocytes. Some plants can inhibit MITF e.g Gentiana veitchiorum Hemsl., Thymelaea hirsuta, Argania spinosa L. In this review, we update the information about MITF and describe the mechanism of its inhibitors in preventing melanogenesis.

A novel 7‑hypoxia‑related long non‑coding RNA signature associated with prognosis and proliferation in melanoma.

Hypoxia-related long non-coding RNAs (lncRNAs) are important indicators of the poor prognosis of cancers. The present study aimed to explore the potential relationship between melanoma and hypoxia-related lncRNAs. The transcriptome and clinical data of patients with melanoma were downloaded from The Cancer Genome Atlas database. The prognostic hypoxia-related lncRNAs were screened out using Pearson's correlation test and univariate Cox analysis. As a result, a hypoxia-related-lncRNA signature based on the expression of 7 lncRNAs was constructed, with one unfavourable [MIR205 host gene (MIR205HG)] and six favourable (T cell receptor β variable 11-2, HLA-DQB1 antisense RNA 1, AL365361.1, AC004847.1, ubiquitin specific peptidase 30 antisense RNA 1 and AC022706.1) lncRNAs as prognostic factors for melanoma. Patients with melanoma were divided into high- and low-risk groups based on the risk score obtained. Survival analyses were performed to assess the prognostic value of the present risk model. Potential tumour-associated biological pathways associated with the present signature were explored using gene set enrichment analysis. The CIBERSORT algorithm demonstrated the important role of the hypoxia-related lncRNAs in regulating tumour-infiltrating immune cells. Clinical samples collected from our center partly confirmed our findings. Cell Counting Kit-8 and flow cytometry assays indicated the suppression of proliferation of melanoma cells following inhibition of MIR205HG expression. Indicators of the canonical Wnt/β-catenin signalling pathway were detected by western blotting. The present study demonstrated that MIR205HG could promote melanoma cell proliferation partly via the canonical Wnt/β-catenin signalling pathway. These findings indicated a 7-hypoxia-related-lncRNA signature that can serve as a novel predictor of melanoma prognosis.

Immune Landscape and an RBM38-Associated Immune Prognostic Model with Laboratory Verification in Malignant Melanoma.

Simple SummaryThe primary treatment of malignant melanoma is a classical regimen of surgery combined with chemotherapy, targeted drugs, and immunotherapy. The purpose of this study was to explore the immune response mechanism of RNA binding protein RBM38 in the development of melanoma with the screening of effective immunodiagnostic models and targeted therapy. We found that RBM38, as an oncogene, promotes the proliferation, invasion, and migration of melanoma cells and is associated with immune infiltration and pathways. Our investigation presented the prognostic significance of RBM38-associated immune signature. In addition, this model may provide a potential strategy for improving the survival and immunotherapy of melanoma patients.Background: Current studies have revealed that RNA-binding protein RBM38 is closely related to tumor development, while its role in malignant melanoma remains unclear. Therefore, this research aimed to investigate the function of RBM38 in melanoma and the prognosis of the disease. Methods: Functional experiments (CCK-8 assay, cell colony formation, transwell cell migration/invasion experiment, wound healing assay, nude mouse tumor formation, and immunohistochemical analysis) were applied to evaluate the role of RBM38 in malignant melanoma. Immune-associated differentially expressed genes (DEGs) on RBM38 related immune pathways were comprehensively analyzed based on RNA sequencing results. Results: We found that high expression of RBM38 promoted melanoma cell proliferation, invasion, and migration, and RBM38 was associated with immune infiltration. Then, a five-gene (A2M, NAMPT, LIF, EBI3, and ERAP1) model of RBM38-associated immune DEGs was constructed and validated. Our signature showed superior prognosis capacity compared with other melanoma prognostic signatures. Moreover, the risk score of our signature was connected with the infiltration of immune cells, immune-regulatory proteins, and immunophenoscore in melanoma. Conclusions: We constructed an immune prognosis model using RBM38-related immune DEGs that may help evaluate melanoma patient prognosis and immunotherapy modalities.

EEF2K silencing inhibits tumour progression through repressing SPP1 and synergises with BET inhibitors in melanoma.

BackgroundDespite the remarkable breakthroughs achieved in the management of metastatic melanoma using immunotherapy and targeted therapies, long‐term clinical efficacy is often compromised due to dose‐limiting toxicity and innate or acquired resistance. Therefore, it is of vital importance to further explore the molecular mechanisms underlying melanoma progression and identify new targeted therapeutic approaches.MethodsThe function of eukaryotic elongation factor‐2 kinase (EEF2K) in melanoma were investigated in vitro and in vivo. RNA‐seq and chromatin immunoprecipitation (ChIP) assay were undertaken to explore the mechanisms. The antitumor effect of bromodomain and extra terminal domain (BET) inhibitors combined with cytarabine were assessed in melanoma both in vitro and in vivo.ResultsEEF2K silencing markedly attenuated the malignant phenotypes of melanoma cells, including proliferation, migration, invasion and metastasis. In contrast, EEF2K overexpression promoted melanoma cell proliferation, migration and invasion. Mechanistically, we demonstrated that EEF2K upregulates the phosphorylation of STAT3 (p‐STAT3) at Tyr705, which binds to the promoter region of SPP1 and enhances its transcription, thus facilitating melanoma progression. Transfection‐induced re‐expression of SPP1 partly negated the inhibitory effect of EEF2K silencing on melanoma, whereas inhibition of SPP1 or STAT3 significantly abolished the efficacy of EEF2K on melanoma cells. Intriguingly, EEF2K silencing combined with BET inhibitor treatment further inhibited cell proliferation and promoted apoptosis in melanoma. We further screened the US FDA‐approved antitumour drug library and identified cytarabine as a potential clinically applicable EEF2K inhibitor that could synergise with BET inhibitors in melanoma treatment.ConclusionEEF2K/p‐STAT3/SPP1 may be a novel oncogenic pathway in melanoma progression, which could be a target for novel combination therapy for melanoma.

Acetylation-dependent regulation of BRAF oncogenic function.

SUMMARYAberrant BRAF activation, including the BRAFV600E mutation, is frequently observed in human cancers. However, it remains largely elusive whether other types of post-translational modification(s) in addition to phosphorylation and ubiquitination-dependent regulation also modulate BRAF kinase activity. Here, we report that the acetyltransferase p300 activates the BRAF kinase by promoting BRAF K601 acetylation, a process that is antagonized by the deacetylase SIRT1. Notably, K601 acetylation facilitates BRAF dimerization with RAF proteins and KSR1. Furthermore, K601 acetylation promotes melanoma cell proliferation and contributes to BRAFV600E inhibitor resistance in BRAFV600E harboring melanoma cells. As such, melanoma patient-derived K601E oncogenic mutation mimics K601 acetylation to augment BRAF kinase activity. Our findings, therefore, uncover a layer of BRAF regulation and suggest p300 hyperactivation or SIRT1 deficiency as potential biomarkers to determine ERK activation in melanomas.

Bladder cancer-associated transcript 1 promotes melanoma cell proliferation and invasion via the miR-374b-5p/U2-associated factor homology motif kinase 1 axis.

Melanoma is a malignancy derived from melanocytes and is associated with high mortality rates worldwide. Long noncoding RNAs (lncRNAs) have been confirmed to be pivotal regulators in multiple types of cancer. Many lncRNAs are aberrantly expressed in tumors and perform vital functions in cancer progression. Nevertheless, the biological role of lncRNA bladder cancer-associated transcript 1 (BLACAT1) in melanoma progression remains unexplored. In this study, the collected data showed that BLACAT1 was highly expressed in melanoma. Mechanistically, miR-374b-5p bound to BLACAT1, and U2-associated factor homology motif kinase 1 (UHMK1) was a downstream target of miR-374b-5p. BLACAT1 upregulated UHMK1 expression by acting as a competing endogenous RNA for miR-374-5b. BLACAT1 deficiency resulted in the upregulation of miR-374b-5p expression and the downregulation of UHMK1 expression in melanoma cells. Moreover, BLACAT1 activated PI3K and AKT signaling by upregulating UHMK1 expression, as shown by western blotting analyses. Functionally, UHMK1 overexpression or miR-374b-5p knockdown reversed the suppressive effect of BLACAT1 depletion on melanoma cell proliferation and invasion. In conclusion, BLACAT1 promotes melanoma cell proliferation and invasion by upregulating UHMK1 expression via miR-374b-5p to activate the PI3K/AKT pathway. These results might provide promising insight into the investigation of prognostic biomarkers of melanoma.

UBTF facilitates melanoma progression via modulating MEK1/2-ERK1/2 signalling pathways by promoting GIT1 transcription

BackgroundUBTF is an HMGB-box DNA binding protein and a necessary Pol I/Pol II basal transcription factor. It has been found that UBTF involves in carcinogenesis and progression of a few cancers. Nevertheless, the the biological function and potential molecular mechanism of UBTF in melanoma are still not clear and need to be clarified.MethodsUBTF and GIT1 expressions in melanoma specimens and cell lines were examined by quantitative real-time PCR (qRT-PCR) and Western blot. MTT and colony formation assays were used to investigate the effects of UBTF and GIT1 on melanoma cell proliferation. Cell cycle and apoptosis assays were detected by flow cytometry. Tumor formation assay was used to analyze the effect of UBTF on melanoma growth. Bioinformatics predicting, chromatin immunoprecipitation (ChIP)-qRT-PCR and reporter gene assay were fulfilled for verifing GIT1 as UBTF targeting gene.ResultsHere we reported that UBTF mRNA and protein expressions were upregulated in primary melanoma specimens and cell lines. UBTF overexpression facilitated melanoma cell proliferation and cell cycle progression and restrained. Silencing UBTF suppressed cell multiplication, cell cycle progression and tumor growth, and promoted apoptosis. UBTF expression was positively related with GIT1 expression in human melanoma tissues. It was verified that UBTF promoted GIT1 transcription in melanoma cells through binding to the promoter region of GIT1. Furthermore, GIT1 overexpression promoted melanoma cell growth and suppressed apoptosis. Knockdown of GIT1 inhibited cell multiplication and induced apoptosis. Overexpression of GIT1 eliminated the effects of silencing UBTF on melanoma cells. Importantly, UBTF activated MEK1/2-ERK1/2 signalling pathways by upregulating GIT1 expression.ConclusionsOur study demonstrates that UBTF promotes melanoma cell proliferation and cell cycle progression by promoting GIT1 transcription, thereby activating MEK1/2-ERK1/2 signalling pathways. The findings indicate that UBTF plays a crucial function in melanoma and may be a potential therapeutic target for the treatment of this disease.

Effects of RNA methylation N6-methyladenosine regulators on malignant progression and prognosis of melanoma

BackgroundMelanoma is an extremely aggressive type of skin cancer and experiencing a expeditiously rising mortality in a current year. Exploring new potential prognostic biomarkers and therapeutic targets of melanoma are urgently needed. The ambition of this research was to identify genetic markers and assess prognostic performance of N6-methyladenosine (m6A) regulators in melanoma.MethodsGene expression data and corresponding clinical informations of melanoma patients as well as sequence data of normal controls are collected from The Cancer Genome Atlas (TCGA) and the Genotype-Tissue Expression (GTEx) databases. Quantitative real-time PCR (qRT-PCR) analysis was carried out to detect the RNA expression of IGF2BP3 in A375 cell line, melanoma tissues, and normal tissues. Western blot, cell proliferation, and migration assays were performed to assess the ability of IGF2BP3 in A375 cell line.ResultsDifferently expressed m6A regulators between tumor samples and normal samples were analyzed. A three-gene prognostic signature including IGF2BP3, RBM15B, and METTL16 was constructed, and the risk score of this signature was identified to be an independent prognostic indicator for melanoma. In addition, IGF2BP3 was verified to promote melanoma cell proliferation and migration in vitro and associate with lymph node metastasis in clinical samples. Moreover, risk score and the expression of IGF2BP3 were positively associated with the infiltrating immune cells and these hub genes made excellent potential drug targets in melanoma.ConclusionWe identified the genetic changes in m6A regulatory genes and constructed a three-gene risk signature with distinct prognostic value in melanoma. This research provided new insights into the epigenetic understanding of m6A regulators and novel therapeutic strategies in melanoma.

LncRNA POU3F3 promotes melanoma cell proliferation by downregulating lncRNA MEG3

BackgroundLncRNA POU3F3 (POU3F3) is overexpressed and plays oncogenic roles in esophageal squamous-cell carcinomas. LncRNA MEG3 (MEG3) has been characterized as a tumor suppressive lncRNA in different types of cancer. Our preliminary deep sequencing analysis revealed the inverse correlation between POU3F3 and MEG2 across melanoma tissues, indicating the interaction between them in melanoma. Therefore, this study was performed to investigate the crosstalk between POU3F3 and MEG3 in melanoma.MethodsTumor and adjacent healthy tissues collected from 60 melanoma patients were subjected to RNA extractions and RT-qPCRs to analyze the differential expression of POU3F3 and MEG2 in melanoma. In melanoma cells, POU3F3 and MEG2 were overexpressed to study the interactions between them. CCK-8 assays were performed to analyze the roles of POU3F3 and MEG2 in regulating melanoma cell proliferation.ResultsWe found that POU3F3 was upregulated, while lncRNA MEG3 was downregulated in melanoma. Expression levels of POU3F3 and MEG3 were inversely correlated across tumor tissues. In vitro experiments showed that POU3F3 overexpression decreased MEG3 expression in melanoma cells, while MEG3 overexpression failed to affect POU3F3. POU3F3 overexpression increased melanoma cell proliferation, while MEG3 overexpression decreased melanoma cell proliferation. In addition, rescue experiments showed that MEG3 overexpression attenuated the enhancing effects of POU3F3 overexpression.ConclusionPOU3F3 may promote melanoma cell proliferation by downregulating MEG3.