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The novel cis-encoded antisense RNA AsrC positively regulates the expression of rpoE-rseABC operon and thus enhances the motility of Salmonella enterica serovar typhi.

Zhang Q, Zhang Y, Zhang X, Zhan L, Zhao X, Xu S, Sheng X, Huang X - Front Microbiol (2015)

Bottom Line: We found that AsrC increased the levels of rseC mRNA and protein.The expression of rpoE was also increased in S. typhi after overexpression of AsrC, which was dependent on rseC.Thus, we propose that AsrC increased RseC level and indirectly activating RpoE which can initiate fliA expression and promote the motility of S. typhi.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Jiangsu University - School of Medicine Zhenjiang, China ; Danyang People's Hospital of Jiangsu Province Danyang, China.

ABSTRACT
Bacterial non-coding RNAs are essential in many cellular processes, including response to environmental stress, and virulence. Deep sequencing analysis of the Salmonella enterica serovar typhi (S. typhi) transcriptome revealed a novel antisense RNA transcribed in cis on the strand complementary to rseC, an activator gene of sigma factor RpoE. In this study, expression of this antisense RNA was confirmed in S. typhi by Northern hybridization. Rapid amplification of cDNA ends and sequence analysis identified an 893 bp sequence from the antisense RNA coding region that covered all of the rseC coding region in the reverse direction of transcription. This sequence of RNA was named as AsrC. After overexpression of AsrC with recombinantant plasmid in S. typhi, the bacterial motility was increased obviously. To explore the mechanism of AsrC function, regulation of rseC and rpoE expression by AsrC was investigated. We found that AsrC increased the levels of rseC mRNA and protein. The expression of rpoE was also increased in S. typhi after overexpression of AsrC, which was dependent on rseC. Thus, we propose that AsrC increased RseC level and indirectly activating RpoE which can initiate fliA expression and promote the motility of S. typhi.

No MeSH data available.


Related in: MedlinePlus

Expression of AsrC under stress conditions. (A) Northern hybridization and (B) qRT-PCR of total RNA isolated from Salmonella typhi cells grown in LB to OD600 0.4 and subjected for 30 min to acid stress (HCl: pH 4.5), oxidative stress (H2O2: 4 mM hydrogen peroxide) or osmotic shock (NaCl: 0.3 M NaCl). Internal reference was 5S rRNA. ∗∗P < 0.01 compared with control group.
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Figure 3: Expression of AsrC under stress conditions. (A) Northern hybridization and (B) qRT-PCR of total RNA isolated from Salmonella typhi cells grown in LB to OD600 0.4 and subjected for 30 min to acid stress (HCl: pH 4.5), oxidative stress (H2O2: 4 mM hydrogen peroxide) or osmotic shock (NaCl: 0.3 M NaCl). Internal reference was 5S rRNA. ∗∗P < 0.01 compared with control group.

Mentions: To understand the expression characteristics of asrC, RNA was harvested from wild-type S. typhi at different times and under different stress conditions and analyzed by Northern hybridization and qRT-PCR. Total RNA from a S. typhi wildtype strain was collected at OD600 0.3, 0.8, 1.2, and 1.8, representing bacterial growth phases from lag through stationary. Two oligonucleotide probes specific to different regions of the asrC gene (Figure 1B) were designed for Northern hybridization. AsrC RNA was detected by both probes mainly in late log and early stationary phase (Figure 2A). The expression pattern of asrC was observed by qRT-PCR (Figure 2B). Expression of asrC was also determined under stress conditions reflecting the Salmonella environment upon invasion of a host or within macrophages. Total RNA was extracted from wild-type after cells were subjected to acidic, oxidative, or osmotic stress. Northern hybridization and qRT-PCR analysis showed that expression of asrC was reduced in acidic, oxidative conditions (Figures 3A,B). These results suggested that the novel asRNA AsrC was expressed in S. typhi and that expression was regulated by environmental conditions.


The novel cis-encoded antisense RNA AsrC positively regulates the expression of rpoE-rseABC operon and thus enhances the motility of Salmonella enterica serovar typhi.

Zhang Q, Zhang Y, Zhang X, Zhan L, Zhao X, Xu S, Sheng X, Huang X - Front Microbiol (2015)

Expression of AsrC under stress conditions. (A) Northern hybridization and (B) qRT-PCR of total RNA isolated from Salmonella typhi cells grown in LB to OD600 0.4 and subjected for 30 min to acid stress (HCl: pH 4.5), oxidative stress (H2O2: 4 mM hydrogen peroxide) or osmotic shock (NaCl: 0.3 M NaCl). Internal reference was 5S rRNA. ∗∗P < 0.01 compared with control group.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4585123&req=5

Figure 3: Expression of AsrC under stress conditions. (A) Northern hybridization and (B) qRT-PCR of total RNA isolated from Salmonella typhi cells grown in LB to OD600 0.4 and subjected for 30 min to acid stress (HCl: pH 4.5), oxidative stress (H2O2: 4 mM hydrogen peroxide) or osmotic shock (NaCl: 0.3 M NaCl). Internal reference was 5S rRNA. ∗∗P < 0.01 compared with control group.
Mentions: To understand the expression characteristics of asrC, RNA was harvested from wild-type S. typhi at different times and under different stress conditions and analyzed by Northern hybridization and qRT-PCR. Total RNA from a S. typhi wildtype strain was collected at OD600 0.3, 0.8, 1.2, and 1.8, representing bacterial growth phases from lag through stationary. Two oligonucleotide probes specific to different regions of the asrC gene (Figure 1B) were designed for Northern hybridization. AsrC RNA was detected by both probes mainly in late log and early stationary phase (Figure 2A). The expression pattern of asrC was observed by qRT-PCR (Figure 2B). Expression of asrC was also determined under stress conditions reflecting the Salmonella environment upon invasion of a host or within macrophages. Total RNA was extracted from wild-type after cells were subjected to acidic, oxidative, or osmotic stress. Northern hybridization and qRT-PCR analysis showed that expression of asrC was reduced in acidic, oxidative conditions (Figures 3A,B). These results suggested that the novel asRNA AsrC was expressed in S. typhi and that expression was regulated by environmental conditions.

Bottom Line: We found that AsrC increased the levels of rseC mRNA and protein.The expression of rpoE was also increased in S. typhi after overexpression of AsrC, which was dependent on rseC.Thus, we propose that AsrC increased RseC level and indirectly activating RpoE which can initiate fliA expression and promote the motility of S. typhi.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Jiangsu University - School of Medicine Zhenjiang, China ; Danyang People's Hospital of Jiangsu Province Danyang, China.

ABSTRACT
Bacterial non-coding RNAs are essential in many cellular processes, including response to environmental stress, and virulence. Deep sequencing analysis of the Salmonella enterica serovar typhi (S. typhi) transcriptome revealed a novel antisense RNA transcribed in cis on the strand complementary to rseC, an activator gene of sigma factor RpoE. In this study, expression of this antisense RNA was confirmed in S. typhi by Northern hybridization. Rapid amplification of cDNA ends and sequence analysis identified an 893 bp sequence from the antisense RNA coding region that covered all of the rseC coding region in the reverse direction of transcription. This sequence of RNA was named as AsrC. After overexpression of AsrC with recombinantant plasmid in S. typhi, the bacterial motility was increased obviously. To explore the mechanism of AsrC function, regulation of rseC and rpoE expression by AsrC was investigated. We found that AsrC increased the levels of rseC mRNA and protein. The expression of rpoE was also increased in S. typhi after overexpression of AsrC, which was dependent on rseC. Thus, we propose that AsrC increased RseC level and indirectly activating RpoE which can initiate fliA expression and promote the motility of S. typhi.

No MeSH data available.


Related in: MedlinePlus