Genomic mapping of phosphorothioates reveals partial modification of short consensus sequences
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Author(s)
Cao, Bo
Chen, Chao
DeMott, Michael S.
Cheng, Qiuxiang
Clark, Tyson A.
Xiong, Xiaolin
Zheng, Xiaoqing
Levine, Stuart S.
Yuan, George
Boitano, Matthew
Date Issued
June 2014
Journal
Nature Communications
Publisher
Nature Publishing Group
Citation
Cao, Bo, Chao Chen, Michael S. DeMott, Qiuxiang Cheng, Tyson A. Clark, Xiaolin Xiong, Xiaoqing Zheng, et al. “Genomic Mapping of Phosphorothioates Reveals Partial Modification of Short Consensus Sequences.” Nat Comms 5 (June 5, 2014).
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Author's final manuscript
Abstract
Bacterial phosphorothioate (PT) DNA modifications are incorporated by Dnd proteins A-E and often function with DndF-H as a restriction-modification (R-M) system, as in Escherichia coli B7A. However, bacteria such as Vibrio cyclitrophicus FF75 lack dndF-H, which points to other PT functions. Here we report two novel, orthogonal technologies to map PTs across the genomes of B7A and FF75 with >90% agreement: single molecule, real-time sequencing and deep sequencing of iodine-induced cleavage at PT (ICDS). In B7A, we detect PT on both strands of G[subscript ps]AAC/G[subscript ps]TTC motifs, but with only 12% of 40,701 possible sites modified. In contrast, PT in FF75 occurs as a single-strand modification at C[subscript ps]CA, again with only 14% of 160,541 sites modified. Single-molecule analysis indicates that modification could be partial at any particular genomic site even with active restriction by DndF-H, with direct interaction of modification proteins with GAAC/GTTC sites demonstrated with oligonucleotides. These results point to highly unusual target selection by PT-modification proteins and rule out known R-M mechanisms.
MIT Department
Massachusetts Institute of Technology. Center for Environmental Health Sciences
Massachusetts Institute of Technology. Department of Biological Engineering
Massachusetts Institute of Technology. Department of Biology
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DOI of Published Version
https://doi.org/10.1038/ncomms4951
