RESUMO
RNA half-life is closely related to its cellular physiological function, so stability determinants may have regulatory functions. Micro(mi)RNAs have primarily been studied with respect to post-transcriptional mRNA regulation and target degradation. Here we study the impact of the tumour suppressive melanoma miRNA miR-211 on transcriptome stability and phenotype in the non-pigmented melanoma cell line, A375. Using 5'-bromouridine IP chase (BRIC)-seq, transcriptome-wide RNA stability profiles revealed highly regulated genes and pathways important in this melanoma cell line. By combining BRIC-seq, RNA-seq and in silico predictions, we identified both existing and novel direct miR-211 targets. We validated DUSP3 as one such novel miR-211 target, which itself sustains colony formation and invasion in A375 cells via MAPK/PI3K signalling. miRNAs have the capacity to control RNA turnover as a gene expression mechanism, and RNA stability profiling is an excellent tool for interrogating functionally relevant gene regulatory pathways and miRNA targets when combined with other high-throughput and in silico approaches.
Assuntos
Regulação Neoplásica da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Melanoma/genética , MicroRNAs/genética , Interferência de RNA , Transcriptoma , Linhagem Celular Tumoral , Biologia Computacional/métodos , Progressão da Doença , Perfilação da Expressão Gênica , Ontologia Genética , Humanos , Melanoma/diagnóstico , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Estadiamento de Neoplasias , Fosfatidilinositol 3-Quinases/metabolismo , Estabilidade de RNA , RNA Mensageiro/genética , Transdução de SinaisRESUMO
Actin, cytoskeleton protein forming microfilaments, play a crucial role in cellular motility. Here we show that exposure to very low levels of polarized light guide their orientation in-vivo within the live cell. Using a simple model to describe the role of actin-filament orientation in directional cellular motion, we demonstrate that the actin polymerization/depolymerization mechanism develops primarily along this direction and, under certain conditions, can lead to guidance of the cell movement. Our results also show a dose dependent increase in actin activity in direct correspondence to the level of laser irradiance. We found that total expression of Tau protein, which stabilize microtubules, was decreased by the irradiance, indicating that exposure to the light may change the activity of kinase, leading to increased cell activity.