Bacterial small RNAs in the Genus Rickettsia

Casey L C Schroeder, Hema P. Narra, Mark Rojas, Abha Sahni, Jignesh Patel, Kamil Khanipov, Thomas Wood, Yuriy Fofanov, Sanjeev Sahni

Research output: Contribution to journalArticle

19 Citations (Scopus)

Abstract

Background: Rickettsia species are obligate intracellular Gram-negative pathogenic bacteria and the etiologic agents of diseases such as Rocky Mountain spotted fever (RMSF), Mediterranean spotted fever, epidemic typhus, and murine typhus. Genome sequencing revealed that R. prowazekii has ~25% non-coding DNA, the majority of which is thought to be either "junk DNA" or pseudogenes resulting from genomic reduction. These characteristics also define other Rickettsia genomes. Bacterial small RNAs, whose biogenesis is predominantly attributed to either the intergenic regions (trans-acting) or to the antisense strand of an open reading frame (cis-acting), are now appreciated to be among the most important post-transcriptional regulators of bacterial virulence and growth. We hypothesize that intergenic regions in rickettsial species encode for small, non-coding RNAs (sRNAs) involved in the regulation of its transcriptome, leading to altered virulence and adaptation depending on the host niche. Results: We employed a combination of bioinformatics and in vitro approaches to explore the presence of sRNAs in a number of species within Genus Rickettsia. Using the sRNA Identification Protocol using High-throughput Technology (SIPHT) web interface, we predicted over 1,700 small RNAs present in the intergenic regions of 16 different strains representing 13 rickettsial species. We further characterized novel sRNAs from typhus (R. prowazekii and R. typhi) and spotted fever (R. rickettsii and R. conorii) groups for their promoters and Rho-independent terminators using Bacterial Promoter Prediction Program (BPROM) and TransTermHP prediction algorithms, respectively. Strong σ70 promoters were predicted upstream of all novel small RNAs, indicating the potential for transcriptional activity. Next, we infected human microvascular endothelial cells (HMECs) with R. prowazekii for 3 h and 24 h and performed Next Generation Sequencing to experimentally validate the expression of 26 sRNA candidates predicted in R. prowazekii. Reverse transcriptase PCR was also used to further verify the expression of six putative novel sRNA candidates in R. prowazekii. Conclusions: Our results yield clear evidence for the expression of novel R. prowazekii sRNA candidates during infection of HMECs. This is the first description of novel small RNAs for a highly pathogenic species of Rickettsia, which should lead to new insights into rickettsial virulence and adaptation mechanisms.

Original languageEnglish (US)
Article number1075
JournalBMC Genomics
Volume16
Issue number1
DOIs
StatePublished - Dec 18 2015

Fingerprint

Bacterial RNA
Rickettsia
Small Untranslated RNA
Intergenic DNA
Epidemic Louse-Borne Typhus
Virulence
RNA
Endothelial Cells
Endemic Flea-Borne Typhus
Boutonneuse Fever
Rocky Mountain Spotted Fever
Genome
Pseudogenes
Computational Biology
Gram-Negative Bacteria
Reverse Transcriptase Polymerase Chain Reaction
Transcriptome
Open Reading Frames
Fever
Technology

Keywords

  • Bioinformatics
  • Deep-Sequencing
  • Endothelial cells
  • Rickettsia
  • SIPHT/sRNAPredict3
  • Small RNAs
  • Spotted Fever
  • Typhus

ASJC Scopus subject areas

  • Biotechnology
  • Genetics

Cite this

Schroeder, C. L. C., Narra, H. P., Rojas, M., Sahni, A., Patel, J., Khanipov, K., ... Sahni, S. (2015). Bacterial small RNAs in the Genus Rickettsia. BMC Genomics, 16(1), [1075]. https://doi.org/10.1186/s12864-015-2293-7

Bacterial small RNAs in the Genus Rickettsia. / Schroeder, Casey L C; Narra, Hema P.; Rojas, Mark; Sahni, Abha; Patel, Jignesh; Khanipov, Kamil; Wood, Thomas; Fofanov, Yuriy; Sahni, Sanjeev.

In: BMC Genomics, Vol. 16, No. 1, 1075, 18.12.2015.

Research output: Contribution to journalArticle

Schroeder, CLC, Narra, HP, Rojas, M, Sahni, A, Patel, J, Khanipov, K, Wood, T, Fofanov, Y & Sahni, S 2015, 'Bacterial small RNAs in the Genus Rickettsia', BMC Genomics, vol. 16, no. 1, 1075. https://doi.org/10.1186/s12864-015-2293-7
Schroeder CLC, Narra HP, Rojas M, Sahni A, Patel J, Khanipov K et al. Bacterial small RNAs in the Genus Rickettsia. BMC Genomics. 2015 Dec 18;16(1). 1075. https://doi.org/10.1186/s12864-015-2293-7
Schroeder, Casey L C ; Narra, Hema P. ; Rojas, Mark ; Sahni, Abha ; Patel, Jignesh ; Khanipov, Kamil ; Wood, Thomas ; Fofanov, Yuriy ; Sahni, Sanjeev. / Bacterial small RNAs in the Genus Rickettsia. In: BMC Genomics. 2015 ; Vol. 16, No. 1.
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AU - Khanipov, Kamil

AU - Wood, Thomas

AU - Fofanov, Yuriy

AU - Sahni, Sanjeev

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N2 - Background: Rickettsia species are obligate intracellular Gram-negative pathogenic bacteria and the etiologic agents of diseases such as Rocky Mountain spotted fever (RMSF), Mediterranean spotted fever, epidemic typhus, and murine typhus. Genome sequencing revealed that R. prowazekii has ~25% non-coding DNA, the majority of which is thought to be either "junk DNA" or pseudogenes resulting from genomic reduction. These characteristics also define other Rickettsia genomes. Bacterial small RNAs, whose biogenesis is predominantly attributed to either the intergenic regions (trans-acting) or to the antisense strand of an open reading frame (cis-acting), are now appreciated to be among the most important post-transcriptional regulators of bacterial virulence and growth. We hypothesize that intergenic regions in rickettsial species encode for small, non-coding RNAs (sRNAs) involved in the regulation of its transcriptome, leading to altered virulence and adaptation depending on the host niche. Results: We employed a combination of bioinformatics and in vitro approaches to explore the presence of sRNAs in a number of species within Genus Rickettsia. Using the sRNA Identification Protocol using High-throughput Technology (SIPHT) web interface, we predicted over 1,700 small RNAs present in the intergenic regions of 16 different strains representing 13 rickettsial species. We further characterized novel sRNAs from typhus (R. prowazekii and R. typhi) and spotted fever (R. rickettsii and R. conorii) groups for their promoters and Rho-independent terminators using Bacterial Promoter Prediction Program (BPROM) and TransTermHP prediction algorithms, respectively. Strong σ70 promoters were predicted upstream of all novel small RNAs, indicating the potential for transcriptional activity. Next, we infected human microvascular endothelial cells (HMECs) with R. prowazekii for 3 h and 24 h and performed Next Generation Sequencing to experimentally validate the expression of 26 sRNA candidates predicted in R. prowazekii. Reverse transcriptase PCR was also used to further verify the expression of six putative novel sRNA candidates in R. prowazekii. Conclusions: Our results yield clear evidence for the expression of novel R. prowazekii sRNA candidates during infection of HMECs. This is the first description of novel small RNAs for a highly pathogenic species of Rickettsia, which should lead to new insights into rickettsial virulence and adaptation mechanisms.

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