title: SmD1 Modulates the miRNA Pathway Independently of Its Pre-mRNA Splicing Function.
abstract: microRNAs (miRNAs) are a class of endogenous regulatory RNAs that play a key role in myriad biological processes. Upon transcription, primary miRNA transcripts are sequentially processed by Drosha and Dicer ribonucleases into ~22-24 nt miRNAs. Subsequently, miRNAs are incorporated into the RNA-induced silencing complexes (RISCs) that contain Argonaute (AGO) family proteins and guide RISC to target RNAs via complementary base pairing, leading to post-transcriptional gene silencing by a combination of translation inhibition and mRNA destabilization. Select pre-mRNA splicing factors have been implicated in small RNA-mediated gene silencing pathways in fission yeast, worms, flies and mammals, but the underlying molecular mechanisms are not well understood. Here, we show that SmD1, a core component of the Drosophila small nuclear ribonucleoprotein particle (snRNP) implicated in splicing, is required for miRNA biogenesis and function. SmD1 interacts with both the microprocessor component Pasha and pri-miRNAs, and is indispensable for optimal miRNA biogenesis. Depletion of SmD1 impairs the assembly and function of the miRISC without significantly affecting the expression of major canonical miRNA pathway components. Moreover, SmD1 physically and functionally associates with components of the miRISC, including AGO1 and GW182. Notably, miRNA defects resulting from SmD1 silencing can be uncoupled from defects in pre-mRNA splicing, and the miRNA and splicing machineries are physically and functionally distinct entities. Finally, photoactivatable-ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) analysis identifies numerous SmD1-binding events across the transcriptome and reveals direct SmD1-miRNA interactions. Our study suggests that SmD1 plays a direct role in miRNA-mediated gene silencing independently of its pre-mRNA splicing activity and indicates that the dual roles of splicing factors in post-transcriptional gene regulation may be evolutionarily widespread.
authors: Tumor Initiation and Maintenance Program, Sanford-Burnham Medical Research Institute, La Jolla, California, United States of America; Development, Aging and Regeneration Program, Sanford-Burnham Medical Research Institute, La Jolla, California, United States of America.
authors: Xiao-Peng
authors: XP
authors: Xiong
authors: Development, Aging and Regeneration Program, Sanford-Burnham Medical Research Institute, La Jolla, California, United States of America.
authors: Georg
authors: G
authors: Vogler
authors: Tumor Initiation and Maintenance Program, Sanford-Burnham Medical Research Institute, La Jolla, California, United States of America; Development, Aging and Regeneration Program, Sanford-Burnham Medical Research Institute, La Jolla, California, United States of America.
authors: Krishna
authors: K
authors: Kurthkoti
authors: Department of Oncology, University of Oxford, Oxford, United Kingdom.
authors: Anastasia
authors: A
authors: Samsonova
authors: Tumor Initiation and Maintenance Program, Sanford-Burnham Medical Research Institute, La Jolla, California, United States of America; Development, Aging and Regeneration Program, Sanford-Burnham Medical Research Institute, La Jolla, California, United States of America.
authors: Rui
authors: R
authors: Zhou
mesh_terms: D000818:Animals; D060565:Argonaute Proteins / Q000235:genetics; D001483:Base Sequence; D002460:Cell Line; D029721:Drosophila Proteins / Q000235:genetics; D004331:Drosophila melanogaster / Q000235:genetics*; D005786:Gene Expression Regulation; D020868:Gene Silencing; D019476:Insect Proteins / Q000235:genetics*; D035683:MicroRNAs / Q000235:genetics*; D012323:RNA Processing, Post-Transcriptional / Q000235:genetics*; D012326:RNA Splicing / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics; D034743:RNA-Induced Silencing Complex / Q000235:genetics; D017423:Sequence Analysis, RNA
chemical_list: C489654:AGO1 protein, Drosophila; D060565:Argonaute Proteins; D029721:Drosophila Proteins; C513405:Gw protein, Drosophila; D019476:Insect Proteins; D035683:MicroRNAs; C534610:Pasha protein, Drosophila; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D034743:RNA-Induced Silencing Complex; C108780:Smd1 protein, insect