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| Status |
Public on Nov 18, 2023 |
| Title |
Eukaryotic RNA-guided endonucleases evolved from a unique clade of bacterial enzymes |
| Organisms |
Escherichia coli; Spodoptera frugiperda |
| Experiment type |
Non-coding RNA profiling by high throughput sequencing
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| Summary |
RNA-guided endonucleases form the crux of diverse biological processes and technologies, including adaptive immunity, transposition, and genome editing. Some of these enzymes are components of insertion sequences (IS) in the IS200/IS605 and IS607 transposon families. Both IS families encode a TnpA transposase and TnpB nuclease, an RNA-guided enzyme ancestral to CRISPR-Cas12. In eukaryotes and their viruses, TnpB homologs occur as two distinct types, Fanzor1 and Fanzor2. We analyzed the evolutionary relationships between prokaryotic TnpBs and eukaryotic Fanzors, revealing that a clade of IS607 TnpBs with unusual active site arrangement found primarily in cyanobacteria likely gave rise to both types of Fanzors. The widespread nature of Fanzors imply that the properties of this particular group of IS607 TnpBs were particularly suited to adaptation and evolution in eukaryotes and their viruses. Biochemical analysis of a prokaryotic IS607 TnpB and virally encoded Fanzor1s revealed features that may have fostered co-evolution between TnpBs/Fanzors and their cognate transposases. These results provide insight into the evolutionary origins of a ubiquitous family of RNA-guided proteins that shows remarkable conservation across the three domains of life.
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| Overall design |
We performed small RNA-sequencing experiments to examine reRNA expression in vivo by infecting S. frugiperda larvae with SfAV. To do so, we pricked the S. frugiperda larvae with an SfAV contaminated needle to mimic oviposition of parasitoids, which effectively transmits the virus. As ascoviruses primarily replicate in the hemolymph, we isolated and sequenced small-RNAs from hemolymph samples from infected S. frugiperda larvae. Mapping of small-RNA reads to the SfAV genome revealed that the reRNA of the Fanzor was one of the most highly transcribed small RNAs in the SfAV genome (accession: AM398843). Consistent with previously reported kinetics of ascovirus infection, we found that reRNA abundance increased 5 days post infection (dpi) compared to 2 dpi, when infected larvae began to develop a cloudy hemolymph caused by budded viral vesicles. In both samples, we observed expression of an RNA that mapped to the right end (RE) of the transposons and extended further downstream into the flanking DNA sequence.
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| Contributor(s) |
Yoon PH, Skopintsev P, Shi H, Chen L, Adler BA, Al-Shimary M, Craig RJ, Loi KJ, DeTurk EC, Li Z, Amerasekara J, Trinidad M, Nisonoff H, Chen K, Lahiri A, Boger R, Jacobsen S, Banfield JF, Doudna JA |
| Citation(s) |
37971304 |
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| Submission date |
Oct 24, 2023 |
| Last update date |
Jan 02, 2024 |
| Contact name |
Marena Isabelle Trinidad |
| E-mail(s) |
marenatrinidad@berkeley.edu
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| Organization name |
Innovative Genomics Institute
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| Street address |
2151 Berkeley Way
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| City |
Berkeley |
| State/province |
CA |
| ZIP/Postal code |
94704 |
| Country |
USA |
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| Platforms (2) |
| GPL16085 |
Illumina MiSeq (Escherichia coli) |
| GPL33862 |
Illumina MiSeq (Spodoptera frugiperda) |
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| Samples (6)
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| Relations |
| BioProject |
PRJNA1031625 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE246134_AM398843_SFAV_IN_VIVO_2And5dpi_FIGURE_5_and_S5.fasta.gz |
42.3 Kb |
(ftp)(http) |
FASTA |
| GSE246134_HAGv-1_SF9_contig_FIGURE_S5.fasta.gz |
1.1 Kb |
(ftp)(http) |
FASTA |
| GSE246134_HvAV-1_SF9_contig_FIGURE_S5.fasta.gz |
1.0 Kb |
(ftp)(http) |
FASTA |
| GSE246134_ISXfa1_IS607_Contig_Figure3A.fasta.gz |
1.4 Kb |
(ftp)(http) |
FASTA |
| GSE246134_RAW.tar |
100.0 Kb |
(http)(custom) |
TAR (of BW) |
| GSE246134_SfAV-1_SF9_contig_FIGURE_S5.fasta.gz |
1.1 Kb |
(ftp)(http) |
FASTA |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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