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Disruption of rimP-SC, encoding a ribosome assembly cofactor, markedly enhances the production of several antibiotics in Streptomyces coelicolor.

Pan Y, Lu C, Dong H, Yu L, Liu G, Tan H - Microb. Cell Fact. (2013)

Bottom Line: Further experiments demonstrated that MetK was one of the reasons for the increment of antibiotics production.Likewise, disruption of rimP-SV of Streptomyces venezuelae also significantly stimulated jadomycin production, suggesting that enhanced antibiotics production might be widespread in many other Streptomyces species.These results established an important relationship between ribosome assembly cofactor and secondary metabolites biosynthesis and provided an approach for yield improvement of secondary metabolites in Streptomyces.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.

ABSTRACT

Background: Ribosome assembly cofactor RimP is one of the auxiliary proteins required for maturation of the 30S subunit in Escherichia coli. Although RimP in protein synthesis is important, its role in secondary metabolites biosynthesis has not been reported so far. Considering the close relationship between protein synthesis and the production of secondary metabolites, the function of ribosome assembly cofactor RimP on antibiotics production was studied in Streptomyces coelicolor and Streptomyces venezuelae.

Results: In this study, the rimP homologue rimP-SC was identified and cloned from Streptomyces coelicolor. Disruption of rimP-SC led to enhanced production of actinorhodin and calcium-dependent antibiotics by promoting the transcription of actII-ORF4 and cdaR. Further experiments demonstrated that MetK was one of the reasons for the increment of antibiotics production. In addition, rimP-SC disruption mutant could be used as a host to produce more peptidyl nucleoside antibiotics (polyoxin or nikkomycin) than the wild-type strain. Likewise, disruption of rimP-SV of Streptomyces venezuelae also significantly stimulated jadomycin production, suggesting that enhanced antibiotics production might be widespread in many other Streptomyces species.

Conclusion: These results established an important relationship between ribosome assembly cofactor and secondary metabolites biosynthesis and provided an approach for yield improvement of secondary metabolites in Streptomyces.

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Related in: MedlinePlus

Transcriptional analysis of genes encoding global activators and repressors as well as sigma factors by real-time RT-PCR. The transcriptional levels were detected after fermentation for 24, 48, 72, 96 and 120 h in GYM medium. Data were presented as the averages of the results of three independent experiments in triplicate. Error bars showed standard deviations.
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Figure 5: Transcriptional analysis of genes encoding global activators and repressors as well as sigma factors by real-time RT-PCR. The transcriptional levels were detected after fermentation for 24, 48, 72, 96 and 120 h in GYM medium. Data were presented as the averages of the results of three independent experiments in triplicate. Error bars showed standard deviations.

Mentions: To study how disruption of rimP-SC enhance ACT/CDA production in S. coelicolor, transcriptions of six global activator genes (absR1, adpA, afsR, atrA, metK and rnc) and three global repressor genes (phoP, ndgR and ssgA) as well as five sigma factor genes (bldN, sigE, sigH, sigR and sigT) were analyzed by real-time RT-PCR. The results indicated that the transcriptions of five activator genes (absR1, adpA, afsR, atrA and rnc) were increased and the transcriptions of three repressor genes (phoP, ndgR and ssgA) were decreased in rimP-SCDM (Figure 5). Although the transcriptional changes of these genes could explain the increase of ACT production, it was unclear whether rimP-SC affected antibiotics biosynthesis at the translational level. Therefore, metK and sigR, whose transcription was not changed significantly in comparison with the wild-type strain, were selected for further studying their translations. To check the syntheses of MetK and SigR, the flag-tagged system was used. The result of western blotting showed that the expression of MetK was much stronger in rimP-SCDM than M145 from 24 h to 72 h (Figure 6A). However, the transcription of metK in rimP-SCDM did not exceed M145 (Figure 6B). In addition, the expression of SigR did not show significant difference between the wild-type strain and rimP-SCDM (data not show). In agreement with previous report [15], our results also showed that over-expression of MetK obviously stimulated ACT production in advance and led to an increase of ACT production up to 140% compared with the control strain M145/pIJ10500 (Figure 6C). Therefore, disruption of rimP-SC increased translational level of proteins related to secondary metabolites, such as MetK which may be one example of many proteins. The complicated mechanism that disruption of rimP homologues led to enhanced production of antibiotics is intriguing, but remains unclear, so it is worthy of studying and is being explored deeply in our lab at present.


Disruption of rimP-SC, encoding a ribosome assembly cofactor, markedly enhances the production of several antibiotics in Streptomyces coelicolor.

Pan Y, Lu C, Dong H, Yu L, Liu G, Tan H - Microb. Cell Fact. (2013)

Transcriptional analysis of genes encoding global activators and repressors as well as sigma factors by real-time RT-PCR. The transcriptional levels were detected after fermentation for 24, 48, 72, 96 and 120 h in GYM medium. Data were presented as the averages of the results of three independent experiments in triplicate. Error bars showed standard deviations.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3716926&req=5

Figure 5: Transcriptional analysis of genes encoding global activators and repressors as well as sigma factors by real-time RT-PCR. The transcriptional levels were detected after fermentation for 24, 48, 72, 96 and 120 h in GYM medium. Data were presented as the averages of the results of three independent experiments in triplicate. Error bars showed standard deviations.
Mentions: To study how disruption of rimP-SC enhance ACT/CDA production in S. coelicolor, transcriptions of six global activator genes (absR1, adpA, afsR, atrA, metK and rnc) and three global repressor genes (phoP, ndgR and ssgA) as well as five sigma factor genes (bldN, sigE, sigH, sigR and sigT) were analyzed by real-time RT-PCR. The results indicated that the transcriptions of five activator genes (absR1, adpA, afsR, atrA and rnc) were increased and the transcriptions of three repressor genes (phoP, ndgR and ssgA) were decreased in rimP-SCDM (Figure 5). Although the transcriptional changes of these genes could explain the increase of ACT production, it was unclear whether rimP-SC affected antibiotics biosynthesis at the translational level. Therefore, metK and sigR, whose transcription was not changed significantly in comparison with the wild-type strain, were selected for further studying their translations. To check the syntheses of MetK and SigR, the flag-tagged system was used. The result of western blotting showed that the expression of MetK was much stronger in rimP-SCDM than M145 from 24 h to 72 h (Figure 6A). However, the transcription of metK in rimP-SCDM did not exceed M145 (Figure 6B). In addition, the expression of SigR did not show significant difference between the wild-type strain and rimP-SCDM (data not show). In agreement with previous report [15], our results also showed that over-expression of MetK obviously stimulated ACT production in advance and led to an increase of ACT production up to 140% compared with the control strain M145/pIJ10500 (Figure 6C). Therefore, disruption of rimP-SC increased translational level of proteins related to secondary metabolites, such as MetK which may be one example of many proteins. The complicated mechanism that disruption of rimP homologues led to enhanced production of antibiotics is intriguing, but remains unclear, so it is worthy of studying and is being explored deeply in our lab at present.

Bottom Line: Further experiments demonstrated that MetK was one of the reasons for the increment of antibiotics production.Likewise, disruption of rimP-SV of Streptomyces venezuelae also significantly stimulated jadomycin production, suggesting that enhanced antibiotics production might be widespread in many other Streptomyces species.These results established an important relationship between ribosome assembly cofactor and secondary metabolites biosynthesis and provided an approach for yield improvement of secondary metabolites in Streptomyces.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.

ABSTRACT

Background: Ribosome assembly cofactor RimP is one of the auxiliary proteins required for maturation of the 30S subunit in Escherichia coli. Although RimP in protein synthesis is important, its role in secondary metabolites biosynthesis has not been reported so far. Considering the close relationship between protein synthesis and the production of secondary metabolites, the function of ribosome assembly cofactor RimP on antibiotics production was studied in Streptomyces coelicolor and Streptomyces venezuelae.

Results: In this study, the rimP homologue rimP-SC was identified and cloned from Streptomyces coelicolor. Disruption of rimP-SC led to enhanced production of actinorhodin and calcium-dependent antibiotics by promoting the transcription of actII-ORF4 and cdaR. Further experiments demonstrated that MetK was one of the reasons for the increment of antibiotics production. In addition, rimP-SC disruption mutant could be used as a host to produce more peptidyl nucleoside antibiotics (polyoxin or nikkomycin) than the wild-type strain. Likewise, disruption of rimP-SV of Streptomyces venezuelae also significantly stimulated jadomycin production, suggesting that enhanced antibiotics production might be widespread in many other Streptomyces species.

Conclusion: These results established an important relationship between ribosome assembly cofactor and secondary metabolites biosynthesis and provided an approach for yield improvement of secondary metabolites in Streptomyces.

Show MeSH
Related in: MedlinePlus