Dissection of affinity captured LINE-1 macromolecular complexes
- Massachusetts General Hospital, United States
- Moscow Institute of Physics and Technology, Russian Federation
- The Rockefeller University, United States
- NYU Langone Medical Center, United States
- Johns Hopkins University School of Medicine, United States
- Novosibirsk State University, Russian Federation
Abstract
Long Interspersed Nuclear Element-1 (LINE-1, L1) is a mobile genetic element active in human genomes. L1-encoded ORF1 and ORF2 proteins bind L1 RNAs, forming ribonucleoproteins (RNPs). These RNPs interact with diverse host proteins, some repressive and others required for the L1 lifecycle. Using differential affinity purifications, quantitative mass spectrometry, and next generation RNA sequencing, we have characterized the proteins and nucleic acids associated with distinctive, enzymatically active L1 macromolecular complexes. Among them, we describe a cytoplasmic intermediate that we hypothesize to be the canonical ORF1p/ORF2p/L1-RNA-containing RNP, and we describe a nuclear population containing ORF2p, but lacking ORF1p, which likely contains host factors participating in target-primed reverse transcription.
Data availability
-
RNA-seq FASTAQ filesPublicly available at ProteomeXchange (accession no: PXD008542).
-
RNAs associated with affinity captured LINE-1 ribonucleoproteinsPublicly available at the NCBI Gene Expression Omnibus (accession no: GSE108270).
Article and author information
Author details
Funding
National Institutes of Health (P41GM109824)
- Michael P Rout
National Institutes of Health (P41GM103314)
- Brian T Chait
National Institutes of Health (P50GM107632)
- Jef D Boeke
National Institutes of Health (R01GM126170)
- John LaCava
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2018, Taylor et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 5,148
- views
-
- 832
- downloads
-
- 67
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Dissection of affinity captured LINE-1 macromolecular complexes
eLife 7:e30094.
https://doi.org/10.7554/eLife.30094
Further reading
-
- Cell Biology
- Genetics and Genomics
The mechanisms by which a retrotransposon called LINE-1 duplicates itself and spreads through the human genome are becoming clearer.
-
- Biochemistry and Chemical Biology
- Genetics and Genomics
Yerba mate (YM, Ilex paraguariensis) is an economically important crop marketed for the elaboration of mate, the third-most widely consumed caffeine-containing infusion worldwide. Here, we report the first genome assembly of this species, which has a total length of 1.06 Gb and contains 53,390 protein-coding genes. Comparative analyses revealed that the large YM genome size is partly due to a whole-genome duplication (Ip-α) during the early evolutionary history of Ilex, in addition to the hexaploidization event (γ) shared by core eudicots. Characterization of the genome allowed us to clone the genes encoding methyltransferase enzymes that catalyse multiple reactions required for caffeine production. To our surprise, this species has converged upon a different biochemical pathway compared to that of coffee and tea. In order to gain insight into the structural basis for the convergent enzyme activities, we obtained a crystal structure for the terminal enzyme in the pathway that forms caffeine. The structure reveals that convergent solutions have evolved for substrate positioning because different amino acid residues facilitate a different substrate orientation such that efficient methylation occurs in the independently evolved enzymes in YM and coffee. While our results show phylogenomic constraint limits the genes coopted for convergence of caffeine biosynthesis, the X-ray diffraction data suggest structural constraints are minimal for the convergent evolution of individual reactions.
