Bioinformatics analysis of small RNAs in Helicobacter pylori and the role of NAT‑67 under tinidazole treatment

Du,Jie; Zhang,Wang; Li,Xiao‑Hui; Li,Yuan‑Jian

doi:10.3892/mmr.2020.11232

Bioinformatics analysis of small RNAs in Helicobacter pylori and the role of NAT‑67 under tinidazole treatment

Authors:
- Jie Du
- Wang Zhang
- Xiao‑Hui Li
- Yuan‑Jian Li
View Affiliations

Affiliations: Department of Pharmacy, Xiangya Hospital, Central South University, Changsha, Hunan 410008, P.R. China, Department of Pharmacology, School of Pharmaceutical Sciences, Central South University, Changsha, Hunan 410078, P.R. China
Published online on: June 15, 2020 https://doi.org/10.3892/mmr.2020.11232
Pages: 1227-1234
Copyright: © Du et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

Helicobacter pylori (Hp) infection is a major cause of gastrointestinal disease. However, the pathogenesis of gastric mucosa injury by Hp has remained elusive. Small non‑coding RNA (sRNA) is a type of widespread RNA in prokaryotic organisms and regulates bacterial growth, reproduction and virulence. In the present study, Hp sRNA profiles were generated to reveal the sequences and possible functions of sRNA by bioinformatics analysis. The role of sRNA in tinidazole (TNZ) treatment was also explored. Total sRNAs of HP26695 were sequenced using an Illumina HiSeq2000. Detected Tags were then compared with a known sRNA database to build an sRNA profile. Reverse transcription‑quantitative (RT‑q)PCR products were sequenced directly and agarose gel electrophoresis was used to identify NAT‑67 and 5'ureB‑sRNA in HP. Furthermore, HP was treated with TNZ for 6, 12 and 24 h. The bacterial concentration was measured, the expression of NAT‑67, 5'ureB‑sRNA and ceuE was determined by RT‑qPCR and superoxide dismutase (SOD) activity and reactive oxygen species (ROS) production were detected. A total of 163 sRNA tags were predicted in Hp through bioinformatics analysis. Among them, 35 tags were evolutionarily aconserved in different Hp strains. By target prediction, it was indicated that certain candidate sRNAs were associated with bacterial oxidative stress, virulence and chemotaxis. It was also observed that NAT‑67 and 5'ureB‑sRNA were downregulated in TNZ‑treated HP. TNZ treatment inhibited the growth of Hp, which was accompanied by downregulation of ceuE and SOD activity, as well as upregulation of ROS. RNA sequencing and bioinformatics are valuable in predicting the expression profile and function of sRNA in HP. sRNA‑targeted genes may be associated with virulence, oxidative stress and chemokines. Downregulation of NAT‑67 by TNZ may be involved in Hp oxidative stress regulation, which may comprise one of the mechanisms of the antibacterial effects of TNZ.

View Figures

View References

1	Salama NR, Hartung ML and Müller A: Life in the human stomach: Persistence strategies of the bacterial pathogen Helicobacter pylori. Nat Rev Microbiol. 11:385–399. 2013. View Article : Google Scholar : PubMed/NCBI
2	Posselt G, Backert S and Wessler S: The functional interplay of Helicobacter pylori factors with gastric epithelial cells induces a multi-step process in pathogenesis. Cell Commun Signal. 11:772013. View Article : Google Scholar : PubMed/NCBI
3	Wassarman KM: 6S RNA: A small RNA regulator of transcription. Curr Opin Microbiol. 10:164–168. 2007. View Article : Google Scholar : PubMed/NCBI
4	Lalaouna D, Simoneau-Roy M, Lafontaine D and Massé E: Regulatory RNAs and target mRNA decay in prokaryotes. Biochim Biophys Acta. 1829:742–747. 2013. View Article : Google Scholar : PubMed/NCBI
5	Sharma CM, Hoffmann S, Darfeuille F, Reignier J, Findeiss S, Sittka A, Chabas S, Reiche K, Hackermüller J, Reinhardt R, et al: The primary transcriptome of the major human pathogen Helicobacter pylori. Nature. 464:250–255. 2010. View Article : Google Scholar : PubMed/NCBI
6	Pernitzsch SR, Tirier SM, Beier D and Sharma CM: A variable homopolymeric G-repeat defines small RNA-mediated posttranscriptional regulation of a chemotaxis receptor in Helicobacter pylori. Proc Natl Acad Sci USA. 111:E501–E510. 2014. View Article : Google Scholar : PubMed/NCBI
7	Wen Y, Feng J and Sachs G: Helicobacter pylori 5′ureB-sRNA, a cis-encoded antisense small RNA, negatively regulates ureAB expression by transcription termination. J Bacteriol. 195:444–452. 2013. View Article : Google Scholar : PubMed/NCBI
8	Xiao B, Li W, Guo G, Li BS, Liu Z, Tang B, Mao XH and Zou QM: Screening and identification of natural antisense transcripts in Helicobacter pylori by a novel approach based on RNase I protection assay. Mol Biol Rep. 36:1853–1858. 2009. View Article : Google Scholar : PubMed/NCBI
9	Xiao B, Li W, Guo G, Li B, Liu Z, Jia K, Guo Y, Mao X and Zou Q: Identification of small noncoding RNAs in Helicobacter pylori by a bioinformatics based approach. Curr Microbiol. 58:258–263. 2009. View Article : Google Scholar : PubMed/NCBI
10	Yi W, Jing F, Scott DR, Marcus EA and Sachs G: A cis-encoded antisense small RNA regulated by the HP0165-HP0166 two-component system controls expression of ureB in Helicobacter pylori. J bacterial. 193:40–51. 2011. View Article : Google Scholar
11	Calderón IL, Morales EH, Collao B, Calderón PF, Chahuán CA, Acuña LG, Gil F and Saavedra CP: Role of salmonella typhimurium small RNAs RyhB-1 and RyhB-2 in the oxidative stress response. Res Microbiol. 165:30–40. 2014. View Article : Google Scholar : PubMed/NCBI
12	Howden BP, Beaume M, Harrison PF, Hernandez D, Schrenzel J, Seemann T, Francois P and Stinear TP: Analysis of the small RNA transcriptional response in multidrug-resistant staphylococcus aureus after antimicrobial exposure. Antimicrob Agents Chemother. 57:3864–3874. 2013. View Article : Google Scholar : PubMed/NCBI
13	Eyraud A, Tattevin P, Chabelskaya S and Felden B: A small RNA controls a protein regulator involved in antibiotic resistance in staphylococcus aureus. Nucleic Acids Res. 42:4892–4905. 2014. View Article : Google Scholar : PubMed/NCBI
14	Pérez-Martínez I and Haas D: Azithromycin inhibits expression of the GacA-dependent small RNAs RsmY and RsmZ in pseudomonas aeruginosa. Antimicrob Agents Chemother. 55:3399–3405. 2011. View Article : Google Scholar : PubMed/NCBI
15	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI
16	Wassarman KM: Small RNAs in bacteria: Diverse regulators of gene expression in response to environmental changes. Cell. 109:141–144. 2002. View Article : Google Scholar : PubMed/NCBI
17	Perrais M, Rousseaux C, Ducourouble MP, Courcol R, Vincent P, Jonckheere N and Van Seuningen I: Helicobacter pylori urease and flagellin alter mucin gene expression in human gastric cancer cells. Gastric Cancer. 17:235–246. 2014. View Article : Google Scholar : PubMed/NCBI
18	Desnoyers G, Bouchard MP and Massé E: New insights into small RNA-dependent translational regulation in prokaryotes. Trends Genet. 29:92–98. 2013. View Article : Google Scholar : PubMed/NCBI
19	Salvail H, Caron MP, Bélanger J and Massé E: Antagonistic functions between the RNA chaperone Hfq and an sRNA regulate sensitivity to the antibiotic colicin. EMBO J. 32:2764–2778. 2013. View Article : Google Scholar : PubMed/NCBI
20	Ramirez-Peña E, Treviño J, Liu Z, Perez N and Sumby P: The group A Streptococcus small regulatory RNA FasX enhances streptokinase activity by increasing the stability of the ska mRNA transcript. Mol Microbiol. 78:1332–1347. 2010. View Article : Google Scholar : PubMed/NCBI
21	Tsugawa H, Suzuki H, Satoh K, Hirata K, Matsuzaki J, Saito Y, Suematsu M and Hibi T: Two amino acids mutation of ferric uptake regulator determines Helicobacter pylori resistance to metronidazole. Antioxid Redox Signal. 14:15–23. 2011. View Article : Google Scholar : PubMed/NCBI
22	Liang H, Zen K, Zhang J, Zhang CY and Chen X: New roles for microRNAs in cross-species communication. RNA Biol. 10:367–370. 2013. View Article : Google Scholar : PubMed/NCBI
23	Scott RB, Christopher N, Xie F, Wood JR and Zempleni J: MicroRNAs are absorbed in biologically meaningful amounts from nutritionally relevant doses of cow milk and affect gene expression in peripheral blood mononuclear cells, HEK-293 kidney cell cultures, and mouse livers. J Nutr. 144:1495–1500. 2013.
24	Zhou Z, Li X, Liu J, Dong L, Chen Q, Liu J, Kong H, Zhang Q, Qi X, Hou D, et al: Honeysuckle-encoded atypical microRNA2911 directly targets influenza A viruses. Cell Res. 25:39–49. 2015. View Article : Google Scholar : PubMed/NCBI
25	Li J, Zhang Y, Li D, Liu Y, Chu D, Jiang X, Hou D, Zen K and Zhang CY: Small non-coding RNAs transfer through mammalian placenta and directly regulate fetal gene expression. Protein Cell. 6:391–396. 2015. View Article : Google Scholar : PubMed/NCBI
26	Ando T, Suzuki H, Nishimura S, Tanaka T, Hiraishi A and Kikuchi Y: Characterization of extracellular RNAs produced by the marine photosynthetic bacterium rhodovulumsulfidophilum. J Biochem. 139:805–811. 2006. View Article : Google Scholar : PubMed/NCBI
27	Li T, Ling ZX and Wen S: Analysis of nucleic acids in lactobacillus culture filtrate. Chin J Microecol. 23:104–106. 2011.
28	Guo L, Sun B, Wu Q, Yang S and Chen F: miRNA-miRNA interaction implicates for potential mutual regulatory pattern. Gene. 511:187–194. 2012. View Article : Google Scholar : PubMed/NCBI
29	Matkovich SJ, Hu Y and Dorn GW II: Regulation of cardiac microRNAs by cardiac microRNAs. Circ Res. 113:62–71. 2013. View Article : Google Scholar : PubMed/NCBI