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The RNA 5’ phosphatase PIR-1 cooperates with dicer to produce endogenous small RNAs and suppress viral replication in C. elegans
Publication . Chaves, Daniel Marques de Almeida de Melo; Fonseca, M. Carmo, 1959-; Mello, Craig C., 1960-
Most organisms utilize small RNAs (sRNA) to control diverse aspects of development, reproduction and physiology by regulating gene expression at the transcriptional and post-transcriptional levels. The essential ribonuclease Dicer is a key enzyme in the production of several types of sRNAs. Prior work on the nematode Caenorhabditis elegans (C. elegans) has uncovered PIR-1 – a small protein conserved in all metazoans – as an interacting partner of Dicer in vivo. The human ortholog of PIR-1 has RNA 5' tri- and diphosphatase activities in vitro, but its biological role remains unclear. With the intent of finding its function, we characterized various aspects of C. elegans PIR-1. We found this enzyme to be essential for general growth and development, germline proliferation, and sperm maturation. We confirmed that PIR-1 associates with Dicer in vivo and expanded its repertoire of known interactions. Profiling of sRNAs from pir-1 loss-of-function animals by high-throughput sequencing revealed that PIR-1 is required for the production of 26G-RNAs during spermatogenesis, a class of Dicer-dependent sRNAs. 26G-RNAs are essential to promote appropriate sperm development, in agreement with the pir-1 mutant sperm defect. Additionally, we discovered a second, 26G-RNA-independent role for PIR-1, in which it cooperates with Dicer and other canonical RNA interference (RNAi) pathway components to suppress the replication of the C. elegans Orsay RNA virus. By demonstrating that PIR-1 functions as its human counterpart in vitro, and that a pir-1 transgene with a mutated phosphatase active site cannot rescue any of the mutant defects, we concluded that PIR-1 acts as an RNA phosphatase in vivo. This is the first study in which concrete biological functions are assigned to this enzyme. Given its high degree of conservation, these results provide a solid basis for studies on the multiple functions of PIR-1 in more complex animals.
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Fundação para a Ciência e a Tecnologia
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SFRH
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SFRH/BD/17629/2004