Discrete subcellular partitioning of human retrotransposon RNAs despite a common mechanism of genome insertion.
Despite the immense significance retrotransposons have had for genome evolution much about their biology is unknown, including the processes of forming their ribonucleoprotein (RNP) particles and transporting them about the cell. Suppression of retrotransposon expression, together with the presence of retrotransposon sequence within numerous mRNAs, makes tracking endogenous L1 RNP particles in cells problematic. We overcame these difficulties by assaying in living and fixed cells tagged-RNPs generated from constructs expressing retrotransposition-competent L1s. In this way, we demonstrate for the first time the subcellular colocalization of L1 RNA and proteins ORF1p and ORF2p, and show their targeting together to cytoplasmic foci. Foci are often associated with markers of cytoplasmic stress granules. Furthermore, mutation analyses reveal that ORF1p can direct L1 RNP distribution within the cell. We also assayed RNA localization of the non-autonomous retrotransposons Alu and SVA. Despite a requirement for the L1 integration machinery, each manifests unique features of subcellular RNA distribution. In nuclei Alu RNA forms small round foci partially associated with marker proteins for coiled bodies, suborganelles involved in the processing of non-coding RNAs. SVA RNA patterning is distinctive, being cytoplasmic but without prominent foci and concentrated in large nuclear aggregates that often ring nucleoli. Such variability predicts significant differences in the life cycles of these elements.
Department of Genetics, University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA. jgoodier@mail.med.upenn.edu
John L
JL
Goodier
Prabhat K
PK
Mandal
Lili
L
Zhang
Haig H
HH
Kazazian
D002460:Cell Line; D003001:Cloning, Molecular; D003593:Cytoplasm / Q000378:metabolism*; D017931:DNA Primers; D005455:Fluorescent Antibody Technique; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D016366:Open Reading Frames; D010957:Plasmids / Q000235:genetics; D012313:RNA / Q000235:genetics / Q000378:metabolism*; D018626:Retroelements / Q000235:genetics*; D048348:Reverse Transcription / Q000235:genetics / Q000502:physiology*; D012261:Ribonucleoproteins / Q000235:genetics / Q000378:metabolism*
D017931:DNA Primers; D018626:Retroelements; D012261:Ribonucleoproteins; D012313:RNA