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RNA-seq reveals the critical role of OtpR in regulating Brucella melitensis metabolism and virulence under acidic stress.

Liu W, Dong H, Li J, Ou Q, Lv Y, Wang X, Xiang Z, He Y, Wu Q - Sci Rep (2015)

Bottom Line: For instance, the virB operon encoding type IV secretion system was significantly down-regulated, and 36 known transcriptional regulators (e.g., vjbR and blxR) were differentially expressed in 16 MΔotpR.Selected RNA-seq results were experimentally confirmed by RT-PCR and RT-qPCR.Overall, these results deciphered differential phenomena associated with virulence, environmental stresses and cell morphology in 16 MΔotpR and 16 M, which provided important information for understanding the detailed OtpR-regulated interaction networks and Brucella pathogenesis.

View Article: PubMed Central - PubMed

Affiliation: 1] Key Laboratory of Animal Epidemiology and Zoonosis of Ministry of Agriculture, College of Veterinary Medicine, China Agricultural University, Beijing 100193, People's Republic of China [2] Unit for Laboratory Animal Medicine and Department of Microbiology and Immunology, The University of Michigan Medical School, Ann Arbor, MI 48109 [3] Center for Computational Medicine and Bioinformatics, The University of Michigan Medical School, Ann Arbor, MI 48109 [4] Beijing Senkang Biotech Development Co., LTD, Beijing 101400, People's Republic of China.

ABSTRACT
The response regulator OtpR is critical for the growth, morphology and virulence of Brucella melitensis. Compared to its wild type strain 16 M, B. melitensis 16 MΔotpR mutant has decreased tolerance to acid stress. To analyze the genes regulated by OtpR under acid stress, we performed RNA-seq whole transcriptome analysis of 16 MΔotpR and 16 M. In total, 501 differentially expressed genes were identified, including 390 down-regulated and 111 up-regulated genes. Among these genes, 209 were associated with bacterial metabolism, including 54 genes involving carbohydrate metabolism, 13 genes associated with nitrogen metabolism, and seven genes associated with iron metabolism. The 16 MΔotpR also decreased capacity to utilize different carbon sources and to tolerate iron limitation in culture experiments. Notably, OtpR regulated many Brucella virulence factors essential for B. melitensis intracellular survival. For instance, the virB operon encoding type IV secretion system was significantly down-regulated, and 36 known transcriptional regulators (e.g., vjbR and blxR) were differentially expressed in 16 MΔotpR. Selected RNA-seq results were experimentally confirmed by RT-PCR and RT-qPCR. Overall, these results deciphered differential phenomena associated with virulence, environmental stresses and cell morphology in 16 MΔotpR and 16 M, which provided important information for understanding the detailed OtpR-regulated interaction networks and Brucella pathogenesis.

No MeSH data available.


Related in: MedlinePlus

Functional categories of the differentially expressed genes in the otpR mutant as compared with the wild-type strain.Only genes that were up- or down-regulated by ≥2.0-fold are shown.
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f1: Functional categories of the differentially expressed genes in the otpR mutant as compared with the wild-type strain.Only genes that were up- or down-regulated by ≥2.0-fold are shown.

Mentions: To detect all the possible genes regulated by OtpR during acid stress, the Next-Generation Sequencing (NGS) technology was used to sequence the whole transcriptomic profiles of 16 MΔotpR and its wild-type strain 16 M. The raw sequence output of the two strain transcriptomes included 150 million reads in total. Approximately 50% reads were perfectly matched to the reference genome B. melitensis 16 M. Based on the genomic alignment, our analysis determined the expression of 3,163 genes in each strain. In total, 501 genes in B. melitensis were identified to be differentially expressed in OtpR (Fig. 1 and Supplemental Data 2). Among these genes, 390 genes were down-regulated and 111 genes were up-regulated in 16 MΔotpR compared to the 16 M control. Most of these differentially expressed genes were associated with carbohydrate metabolism (10.78%), energy metabolism (7.39%), amino acid metabolism (6.19%), nucleotide metabolism (4.59%), lipid metabolism (1.80%), membrane transport (7.39%) and transcription (7.19%) (Fig. 1).


RNA-seq reveals the critical role of OtpR in regulating Brucella melitensis metabolism and virulence under acidic stress.

Liu W, Dong H, Li J, Ou Q, Lv Y, Wang X, Xiang Z, He Y, Wu Q - Sci Rep (2015)

Functional categories of the differentially expressed genes in the otpR mutant as compared with the wild-type strain.Only genes that were up- or down-regulated by ≥2.0-fold are shown.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4542472&req=5

f1: Functional categories of the differentially expressed genes in the otpR mutant as compared with the wild-type strain.Only genes that were up- or down-regulated by ≥2.0-fold are shown.
Mentions: To detect all the possible genes regulated by OtpR during acid stress, the Next-Generation Sequencing (NGS) technology was used to sequence the whole transcriptomic profiles of 16 MΔotpR and its wild-type strain 16 M. The raw sequence output of the two strain transcriptomes included 150 million reads in total. Approximately 50% reads were perfectly matched to the reference genome B. melitensis 16 M. Based on the genomic alignment, our analysis determined the expression of 3,163 genes in each strain. In total, 501 genes in B. melitensis were identified to be differentially expressed in OtpR (Fig. 1 and Supplemental Data 2). Among these genes, 390 genes were down-regulated and 111 genes were up-regulated in 16 MΔotpR compared to the 16 M control. Most of these differentially expressed genes were associated with carbohydrate metabolism (10.78%), energy metabolism (7.39%), amino acid metabolism (6.19%), nucleotide metabolism (4.59%), lipid metabolism (1.80%), membrane transport (7.39%) and transcription (7.19%) (Fig. 1).

Bottom Line: For instance, the virB operon encoding type IV secretion system was significantly down-regulated, and 36 known transcriptional regulators (e.g., vjbR and blxR) were differentially expressed in 16 MΔotpR.Selected RNA-seq results were experimentally confirmed by RT-PCR and RT-qPCR.Overall, these results deciphered differential phenomena associated with virulence, environmental stresses and cell morphology in 16 MΔotpR and 16 M, which provided important information for understanding the detailed OtpR-regulated interaction networks and Brucella pathogenesis.

View Article: PubMed Central - PubMed

Affiliation: 1] Key Laboratory of Animal Epidemiology and Zoonosis of Ministry of Agriculture, College of Veterinary Medicine, China Agricultural University, Beijing 100193, People's Republic of China [2] Unit for Laboratory Animal Medicine and Department of Microbiology and Immunology, The University of Michigan Medical School, Ann Arbor, MI 48109 [3] Center for Computational Medicine and Bioinformatics, The University of Michigan Medical School, Ann Arbor, MI 48109 [4] Beijing Senkang Biotech Development Co., LTD, Beijing 101400, People's Republic of China.

ABSTRACT
The response regulator OtpR is critical for the growth, morphology and virulence of Brucella melitensis. Compared to its wild type strain 16 M, B. melitensis 16 MΔotpR mutant has decreased tolerance to acid stress. To analyze the genes regulated by OtpR under acid stress, we performed RNA-seq whole transcriptome analysis of 16 MΔotpR and 16 M. In total, 501 differentially expressed genes were identified, including 390 down-regulated and 111 up-regulated genes. Among these genes, 209 were associated with bacterial metabolism, including 54 genes involving carbohydrate metabolism, 13 genes associated with nitrogen metabolism, and seven genes associated with iron metabolism. The 16 MΔotpR also decreased capacity to utilize different carbon sources and to tolerate iron limitation in culture experiments. Notably, OtpR regulated many Brucella virulence factors essential for B. melitensis intracellular survival. For instance, the virB operon encoding type IV secretion system was significantly down-regulated, and 36 known transcriptional regulators (e.g., vjbR and blxR) were differentially expressed in 16 MΔotpR. Selected RNA-seq results were experimentally confirmed by RT-PCR and RT-qPCR. Overall, these results deciphered differential phenomena associated with virulence, environmental stresses and cell morphology in 16 MΔotpR and 16 M, which provided important information for understanding the detailed OtpR-regulated interaction networks and Brucella pathogenesis.

No MeSH data available.


Related in: MedlinePlus