ABSTRACT
Despite the importance of microRNAs (miRNAs) in gene regulation, it is unclear how the miRNA-Argonaute complex--or miRNA-induced silencing complex (miRISC)--can regulate the translation of their targets in such diverse ways. We demonstrate here a direct interaction between the miRISC and the ribosome by showing that a constituent of the eukaryotic 40S subunit, receptor for activated C-kinase (RACK1), is important for miRNA-mediated gene regulation in animals. In vivo studies demonstrate that RACK1 interacts with components of the miRISC in nematodes and mammals. In both systems, the alteration of RACK1 expression alters miRNA function and impairs the association of the miRNA complex with the translating ribosomes. Our data indicate that RACK1 can contribute to the recruitment of miRISC to the site of translation, and support a post-initiation mode of miRNA-mediated gene repression.
Subject(s)
Caenorhabditis elegans Proteins/metabolism , Caenorhabditis elegans/genetics , Caenorhabditis elegans/metabolism , GTP-Binding Proteins/metabolism , MicroRNAs/metabolism , Neoplasm Proteins/metabolism , Receptors, Cell Surface/metabolism , Receptors, Cytoplasmic and Nuclear/metabolism , Animals , Gene Expression Regulation , Gene Silencing , HeLa Cells , Humans , MicroRNAs/genetics , Polyribosomes/metabolism , Protein Binding/physiology , RNA-Induced Silencing Complex/metabolism , Receptors for Activated C KinaseABSTRACT
Pulmonary arterial hypertension (PAH) is characterized by enhanced proliferation and reduced apoptosis of pulmonary artery smooth muscle cells (PASMCs). Because microRNAs have been recently implicated in the regulation of cell proliferation and apoptosis, we hypothesized that these regulatory molecules might be implicated in the etiology of PAH. In this study, we show that miR-204 expression in PASMCs is down-regulated in both human and rodent PAH. miR-204 down-regulation correlates with PAH severity and accounts for the proliferative and antiapoptotic phenotypes of PAH-PASMCs. STAT3 activation suppresses miR-204 expression, and miR-204 directly targets SHP2 expression, thereby SHP2 up-regulation, by miR-204 down-regulation, activates the Src kinase and nuclear factor of activated T cells (NFAT). STAT3 also directly induces NFATc2 expression. NFAT and SHP2 were needed to sustain PAH-PASMC proliferation and resistance to apoptosis. Finally, delivery of synthetic miR-204 to the lungs of animals with PAH significantly reduced disease severity. This study uncovers a new regulatory pathway involving miR-204 that is critical to the etiology of PAH and indicates that reestablishing miR-204 expression should be explored as a potential new therapy for this disease.
Subject(s)
MicroRNAs/physiology , Animals , Apoptosis/genetics , Apoptosis/physiology , Cell Proliferation , Familial Primary Pulmonary Hypertension , Gene Expression Regulation , Genetic Markers/genetics , Genetic Markers/physiology , Humans , Hypertension, Pulmonary/genetics , Hypertension, Pulmonary/physiopathology , Male , Mice , MicroRNAs/genetics , Muscle, Smooth, Vascular/metabolism , Muscle, Smooth, Vascular/physiopathology , Oligonucleotide Array Sequence Analysis , Pulmonary Artery/metabolism , Pulmonary Artery/physiopathology , Rats , Reverse Transcriptase Polymerase Chain Reaction , STAT3 Transcription Factor/physiology , Signal Transduction/genetics , Signal Transduction/physiology , src-Family Kinases/metabolism , src-Family Kinases/physiologyABSTRACT
A Tn402-like class 1 integron was recovered from a prawn-associated bacterium. One of its cassettes included methionine sulfoxide reductase genes, the first example of such genes being captured by an integron. The integron was flanked by direct repeats that resemble miniature inverted-repeat transposable element sequences. Excision of the integron by homologous recombination through these sequences was demonstrated.
