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Title: Functional and structural analysis of a highly-expressed Yersinia pestis small RNA following infection of cultured macrophages

Non-coding small RNAs (sRNAs) are found in practically all bacterial genomes and play important roles in regulating gene expression to impact bacterial metabolism, growth, and virulence. We performed transcriptomics analysis to identify sRNAs that are differentially expressed in Yersinia pestis that invaded the human macrophage cell line THP-1, compared to pathogens that remained extracellular in the presence of host. Using ultra high-throughput sequencing, we identified 37 novel and 143 previously known sRNAs in Y. pestis. In particular, the sRNA Ysr170 was highly expressed in intracellular Yersinia and exhibited a log2 fold change ~3.6 higher levels compared to extracellular bacteria. We found that knock-down of Ysr170 expression attenuated infection efficiency in cell culture and growth rate in response to different stressors. In addition, we applied selective 2’-hydroxyl acylation analyzed by primer extension (SHAPE) analysis to determine the secondary structure of Ysr170 and observed structural changes resulting from interactions with the aminoglycoside antibiotic gentamycin and the RNA chaperone Hfq. Interestingly, gentamicin stabilized helix 4 of Ysr170, which structurally resembles the native gentamicin 16S ribosomal binding site. Lastly, we modeled the tertiary structure of Ysr170 binding to gentamycin using RNA motif modeling. Integration of these experimental and structural methods can provide further insightmore » into the design of small molecules that can inhibit function of sRNAs required for pathogen virulence.« less
Authors:
 [1] ; ORCiD logo [1] ;  [2] ; ORCiD logo [1] ;  [3] ;  [1] ;  [4] ; ORCiD logo [1] ; ORCiD logo [1] ; ORCiD logo [1] ;  [1] ;  [5]
  1. Los Alamos National Lab. (LANL), Los Alamos, NM (United States)
  2. Univ. of Utah, Salt Lake City, UT (United States); Los Alamos National Lab. (LANL), Los Alamos, NM (United States)
  3. SRA International, Atlanta, GA (United States); Los Alamos National Lab. (LANL), Los Alamos, NM (United States)
  4. Viome Inc., Los Alamos, NM (United States); Los Alamos National Lab. (LANL), Los Alamos, NM (United States)
  5. East Carolina Univ. Brody School of Medicine, Greenville, NC (United States)
Publication Date:
Report Number(s):
LA-UR-16-20099
Journal ID: ISSN 1932-6203
Grant/Contract Number:
AC52-06NA25396
Type:
Accepted Manuscript
Journal Name:
PLoS ONE
Additional Journal Information:
Journal Volume: 11; Journal Issue: 12; Journal ID: ISSN 1932-6203
Publisher:
Public Library of Science
Research Org:
Los Alamos National Lab. (LANL), Los Alamos, NM (United States)
Sponsoring Org:
USDOE Laboratory Directed Research and Development (LDRD) Program
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; biological science; Yersinia pestis; RNA folding; RNA structure; bacterial pathogens; RNA sequencing; intracellular pathogens; Yersinia; Yersinia pseudotuberculosis
OSTI Identifier:
1351188