RESUMO
Regulation of gene expression via microRNAs is known to promote the development of many types of cancer. In melanoma, miRNAs are globally up-regulated, and alterations of miRNA-processing enzymes have already been identified. However, mis-regulation of miRNA transport has not been analyzed in melanoma yet. We hypothesized that alterations in miRNA transport disrupt miRNA processing. Therefore, we investigated whether the pre-miRNA transporter Exportin-5 (XPO5) was involved in altered miRNA maturation and functional consequences in melanoma. We found that XPO5 is significantly over-expressed in melanoma compared with melanocytes. We showed enhanced XPO5 mRNA stability in melanoma cell lines which likely contributes to up-regulated XPO5 protein expression. In addition, we identified MEK signaling as a regulator of XPO5 expression in melanoma. Knockdown of XPO5 expression in melanoma cells led to decreased mature miRNA levels and drastic functional changes. Our data revealed that aberrant XPO5 expression is important for the maturation of miRNAs and the malignant behavior of melanoma cells. We suggest that the high abundance of XPO5 in melanoma leads to enhanced survival, proliferation and metastasis and thereby supports the aggressiveness of melanoma.
Assuntos
Regulação Neoplásica da Expressão Gênica , Carioferinas/metabolismo , Sistema de Sinalização das MAP Quinases , Melanoma/genética , MicroRNAs/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Sobrevivência Celular , Regulação para Baixo , Técnicas de Silenciamento de Genes , Humanos , Carioferinas/genética , Melanócitos/metabolismo , Melanoma/patologia , Microscopia de Fluorescência , Invasividade Neoplásica , Interferência de RNA , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/metabolismo , Regulação para CimaRESUMO
Piccolo is one of the largest cytomatrix proteins present at active zones of chemical synapses, where it is suggested to play a role in recruiting and integrating molecules relevant for both synaptic vesicle exo- and endocytosis. Here we examined the retina of a Piccolo-mutant mouse with a targeted deletion of exon 14 in the Pclo gene. Piccolo deficiency resulted in its profound loss at conventional chemical amacrine cell synapses but retinal ribbon synapses were structurally and functionally unaffected. This led to the identification of a shorter, ribbon-specific Piccolo variant, Piccolino, present in retinal photoreceptor cells, bipolar cells, as well as in inner hair cells of the inner ear. By RT-PCR analysis and the generation of a Piccolino-specific antibody we show that non-splicing of intron 5/6 leads to premature translation termination and generation of the C-terminally truncated protein specifically expressed at active zones of ribbon synapse containing cell types. With in situ proximity ligation assays we provide evidence that this truncation leads to the absence of interaction sites for Bassoon, Munc13, and presumably also ELKS/CAST, RIM2, and the L-type Ca(2) (+) channel which exist in the full-length Piccolo at active zones of conventional chemical synapses. The putative lack of interactions with proteins of the active zone suggests a function of Piccolino at ribbon synapses of sensory neurons different from Piccolo's function at conventional chemical synapses.
Assuntos
Proteínas do Citoesqueleto/metabolismo , Orelha , Regulação da Expressão Gênica , Neuropeptídeos/metabolismo , Retina/citologia , Células Receptoras Sensoriais/metabolismo , Sinapses/metabolismo , Processamento Alternativo , Sequência de Aminoácidos , Animais , Canais de Cálcio Tipo L , Proteínas de Transporte/metabolismo , Bovinos , Proteínas do Citoesqueleto/química , Proteínas do Citoesqueleto/deficiência , Proteínas do Citoesqueleto/genética , Éxons/genética , Humanos , Imuno-Histoquímica , Camundongos , Dados de Sequência Molecular , Mutação , Proteínas do Tecido Nervoso/metabolismo , Neuropeptídeos/química , Neuropeptídeos/deficiência , Neuropeptídeos/genética , Isoformas de Proteínas/química , Isoformas de Proteínas/deficiência , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Ratos , Proteínas rab de Ligação ao GTP , Proteínas rab3 de Ligação ao GTP/metabolismoRESUMO
Melanoma is the most aggressive form of skin cancer, and the incidence of melanoma has been increasing faster than that of most other cancers. While the survival rate following surgical resection of early-stage primary tumors is nearly 100%, the survival of patients with metastasized tumors is strongly reduced, likely due to resistance to conventional therapies. Therefore, it is important to use new molecular approaches to develop new biomarkers to better prevent and diagnose melanoma. MicroRNAs (miRNAs) are short non-coding RNAs that post-transcriptionally regulate gene expression via repression of translation or direct degradation of their complementary mRNA. In this review, we summarize our current understanding of the involvement of miRNAs and their corresponding targets in melanomagenesis as well as the potential use of miRNAs as biomarkers.
Assuntos
Biomarcadores Tumorais/metabolismo , Melanoma/metabolismo , MicroRNAs/metabolismo , RNA Neoplásico/metabolismo , Neoplasias Cutâneas/metabolismo , Animais , Biomarcadores Tumorais/genética , Intervalo Livre de Doença , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Melanoma/genética , Melanoma/mortalidade , Melanoma/patologia , MicroRNAs/genética , Metástase Neoplásica , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Biossíntese de Proteínas/fisiologia , Estabilidade de RNA/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Neoplásico/genética , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/mortalidade , Neoplasias Cutâneas/patologia , Taxa de SobrevidaRESUMO
Loss of methylthioadenosine phosphorylase (MTAP) expression and a concomitant accumulation of 5'-methyl-thioadenosine (MTA) characterise several tumour entities including malignant melanoma. MTA affects cellular signalling, proliferation and migration not only of cancer but also surrounding cells including lymphocytes and stromal fibroblasts. The mode of action of MTA is still not known. Interestingly, MTA is a known potent inhibitor of protein arginine methyltransferases (PRMTs) and is used as a tool in studying activity and impact of PRMTs. This study aimed at analysing PRMTs in melanoma and the potential impact of MTA on tumourigenesis. Our findings demonstrate that expression of PRMT4/CARM1 and PRMT6 is deregulated in melanoma, whereas expression of the remaining PRMTs stays unchanged. General PRMT activity and, consequently, symmetric and asymmetric protein methylation are reduced significantly in melanoma cells and tissues. This is due to a loss of MTAP expression and accumulation of MTA. Reduction of protein methylation by MTA affects cell signalling and leads, for example, to an activation of extracellular signal-regulated kinase (ERK) activity. The effects of endogeneous MTA on PRMTs as presented in this study can strongly support the migratory and invasive phenotype of melanoma cells.