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| Status |
Public on Apr 22, 2024 |
| Title |
TnpB homologs exapted from transposons are RNA-guided transcription factors |
| Organisms |
Enterobacter cloacae; Escherichia coli; Enterobacter sp. BIDMC93 |
| Experiment type |
Genome binding/occupancy profiling by high throughput sequencing
Expression profiling by high throughput sequencing
Other
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| Summary |
Transposon-encoded tnpB and iscB genes encode RNA-guided DNA nucleases that promote their own selfish spread through targeted DNA cleavage and homologous recombination. These widespread gene families were repeatedly domesticated over evolutionary timescales, leading to the emergence of diverse CRISPR-associated nucleases including Cas9 and Cas12. We set out to test the hypothesis that TnpB nucleases may have also been repurposed for novel, unexpected functions other than CRISPR-Cas. Here, using phylogenetics, structural predictions, comparative genomics, and functional assays, we uncover multiple instances of programmable transcription factors that we name TnpB-like nuclease-dead repressors (TldR). These proteins employ naturally occurring guide RNAs to specifically target conserved promoter regions of the genome, leading to potent gene repression in a mechanism akin to CRISPRi technologies invented by humans. Focusing on a TldR clade found broadly in Enterobacteriaceae, we discover that bacteriophages exploit the combined action of TldR and an adjacently encoded phage gene to alter the expression and composition of the host flagellar assembly, a transformation with the potential to impact motility, phage susceptibility, and host immunity. Collectively, this work showcases the diverse molecular innovations that were enabled through repeated exaptation of transposon-encoded genes, and reveals the evolutionary trajectory of diverse RNA-guided transcription factors.
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| Overall design |
E. coli genome ChIP-seq profiles for 16 RNA-guided endonuclease-dead proteins. 17 samples were sequenced in total. The raw read file "Input_Eco_dTnpB_ChIP-seq_paired_raw" is a control sample which was used as a reference sample for MACS3 peak calling.
RIP-seq for a subset of the 16 RNA-guided endonuclease-dead proteins tested using ChIP. Total RNA-seq of several Enterobacter species encoding endogenous RNA-guided endonuclease-dead proteins.
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| Contributor(s) |
Wiegand T, Hoffmann FT, Walker MW, Tang S, Richard E, Le HC, Meers C, Sternberg SH |
| Citation(s) |
38926585 |
| |
| Submission date |
Oct 18, 2023 |
| Last update date |
Jul 22, 2024 |
| Contact name |
Samuel Henry Sternberg |
| E-mail(s) |
shsternberg@gmail.com
|
| Phone |
717-475-3658
|
| Organization name |
Columbia University
|
| Department |
Biochemistry and Molecular Biophysics
|
| Lab |
Sternberg Lab
|
| Street address |
701 W. 168th Street, HHSC 726
|
| City |
New York |
| State/province |
NY |
| ZIP/Postal code |
10032 |
| Country |
USA |
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| Platforms (4)
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| GPL21222 |
Illumina NextSeq 500 (Escherichia coli) |
| GPL28126 |
NextSeq 550 (Escherichia coli) |
| GPL34277 |
NextSeq 550 (Enterobacter cloacae) |
| GPL34278 |
NextSeq 550 (Enterobacter sp. BIDMC93) |
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| Samples (51)
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| Relations |
| BioProject |
PRJNA1029663 |
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