Journal of Biotechnology 222 (2016) 9–10
Contents lists available at ScienceDirect
Journal of Biotechnology journal homepage: www.elsevier.com/locate/jbiotec
Genome announcement
Complete genome sequence of Streptomyces globisporus C-1027, the producer of an enediyne antibiotic lidamycin Xingxing Li, Xuan Lei, Cong Zhang, Zhibo Jiang, Yuanyuan Shi, Songmei Wang, Lifei Wang ∗∗ , Bin Hong ∗ Key Laboratory of Biotechnology of Antibiotics of Ministry of Health, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China
a r t i c l e
i n f o
Article history: Received 25 January 2016 Accepted 2 February 2016 Available online 4 February 2016 Keywords: Streptomyces globisporus C-1027 Genome sequence Lidamycin
a b s t r a c t Streptomyces globisporus C-1027 produces a nine-membered enediyne antitumor antibiotic lidamycin. Here we report the complete genome sequence of S. globisporus C-1027, which consists of a 7,608,611 bp linear chromosome, a 167,754 bp linear plasmid SGLP1 and a 7,234 bp circular plasmid pSGL1. The biosynthetic gene cluster for lidamycin was located in the linear plasmid SGLP1. Genome analysis also revealed a number of genes related to biosynthesis of diverse secondary metabolites. The genome sequence of C-1027 will enable us to disclose biosynthetic pathways of these secondary metabolites and discover new natural products with potential applications notably in human health. © 2016 Elsevier B.V. All rights reserved.
Streptomyces globisporus C-1027 (CPCC 200270, deposited at the China Pharmaceutical Culture Collection) is a Gram-positive bacterium, originally isolated from a soil sample collected in the Hubei Province of China (Hu et al., 1988). It produces a potent antitumor antibiotic lidamycin, originally called C-1027, which belongs to the nine-membered enediyne family and is investigated in Phase II clinical trials in China (Shao and Zhen, 2008). Lidamycin biosynthetic gene cluster has been cloned and characterized (Liu et al., 2002). Its biosynthesis regulated by a complex circuit involving at least four cluster-situated regulators: SgcR1, SgcR2, SgcR3 and SgcR (Chen et al., 2010, 2011; Ma, 2007; Wang, 2008). Recently, a pleiotropic regulator AtrA, located outside of the lidamycin cluster, was characterized to have a role in regulating lidamycin production (Li et al., 2015). The complete genome sequence information of S. globisporus C-1027 may contribute to the further understanding of the global regulatory mechanisms for lidamycin biosynthesis. The complete genome was obtained by assembling a combination of reads using illumine Hiseq 2000 sequence platform. A total of 622 contigs were ordered using SOAPdenovo 1.05 (Li et al., 2010), and aligned with paired BAC end sequences using synteny with the genome of S. griseus subsp. griseus NBRC 13350 (GenBank accession no. AP009493), which is closely related to S. globisporus C-1027.
∗ Corresponding author. Fax: +86 10 63017302. ∗∗ Corresponding author. Fax: +86 10 63017302. E-mail addresses: [email protected], [email protected] (L. Wang), [email protected], [email protected] (B. Hong). http://dx.doi.org/10.1016/j.jbiotec.2016.02.004 0168-1656/© 2016 Elsevier B.V. All rights reserved.
Table 1 Genomic features of S. globisporus C-1027. Features
Chromosome
Plasmid SGLP1
Plasmid pSGL1
Length (bp) G + C content (%) CDS rRNA genes (operons) tRNA genes GenBank accession no.
7,608,611 71.6 7,078 18 (6) 66 CP013738
167,754 69.5 157 0 (0) 0 CP013739
7,234 70.9 9 0 (0) 0 CP013740
Finally, gaps were closed by PCR with Sanger sequencing. Putative protein-coding sequences were predicted using the Glimmer 3.0 (Delcher et al., 2007). Gene functional annotation was performed using BLASTP with KEGG, COG, Swiss-Port, TrEMBL, NR and GO databases. tRNA-encoding genes and rRNA operons were found by using tRNAscan software and rRNAmmer software. Repeat Masker software and CRISPR Finder software were used to find putative transposons and clustered regularly interspaced short palindromic repeats (CRISPRs). The genomic features are summarized in Table 1. S. globisporus C-1027 has a genome consisting of three replicons, a linear chromosome, a linear plasmid and a circular plasmid (Table 1). The linear chromosome is 7,608,611 bp with a 71.6% G + C content. 7,078 putative protein CDSs, 66 tRNA, 6 rRNA operons with the order 16S-23S-5S, 1 sRNA, and 19 CRISPRs were predicted on the chromosome. The large linear plasmid SGLP1, of 167,754 bp with a 69.5% G + C content, contains 4 CRISPRs and 157 putative genes, including the intact biosynthetic gene cluster for lidamycin. The circular plasmid pSGL1 (7,234 bp, 70.9% G + C content)
10
X. Li et al. / Journal of Biotechnology 222 (2016) 9–10
contains 9 putative CDS. Altogether 7,244 putative protein CDSs were identified in the genome containing 6,879,372 bp, accounting for 88.38% of the genome. On the complete genome, 117,075 bp repeat sequence and putative 49,291 bp transposon were found. Genome mining analysis using antiSMASH 3.0 (Weber et al., 2015) revealed the presence of 33 putative gene clusters responsible for the production of diverse secondary metabolites in the complete genome sequence of S. globisporus C-1027. Of these, the putative isorenieratene, griseobactin, desferrioxamines, ectoine and melanin are highly conserved in the closely related species S. griseus subsp. griseus NBRC 13350 and S. coelicolor A3(2). Some gene clusters related to biosynthesis of terpene, non-ribosomal peptide, polyketide and lantipeptide are not highly conserved with any known cluster, disclosing the potential of S. globisporus C-1027 to produce some new natural products. Nucleotide sequence accession numbers The complete genome sequence of S. globisporus C-1027 (CPCC 200270) has been deposited at GenBank under accession nos. CP013738, CP013739 and CP013740. Acknowledgments We thank Beijing Genomics Institute (Shenzhen, China) for illumine Hiseq sequencing, assembly and annotation of the genome. This work was supported by the National Natural Science Foundation of China (81302677, 81321004, 81402836, and 31170042), and National Mega-Project for Innovative Drugs (2012ZX09301002001-016 and 2015ZX09102007-016).
References Chen, Y., Yin, M., Horsman, G.P., Huang, S., Shen, B., 2010. Manipulation of pathway regulation in Streptomyces globisporus for overproduction of the enediyne antitumor antibiotic C-1027. J. Antibiot. (Tokyo) 63, 482–485. Chen, Y., Yin, M., Horsman, G.P., Shen, B., 2011. Improvement of the enediyne antitumor antibiotic C-1027 production by manipulating its biosynthetic pathway regulation in Streptomyces globisporus. J. Nat. Prod. 74, 420–424. Delcher, A.L., Bratke, K.A., Powers, E.C., Salzberg, S.L., 2007. Identifying bacterial genes and endosymbiont DNA with Glimmer. Bioinformatics 23, 673–679. Hu, J.L., Xue, Y.C., Xie, M.Y., Zhang, R., Otani, T., Minami, Y., Yamada, Y., Marunaka, T., 1988. A new macromolecular antitumor antibiotic, C-1027. I. Discovery, taxonomy of producing organism, fermentation and biological activity. J. Antibiot. (Tokyo) 41, 1575–1579. Li, R., Zhu, H., Ruan, J., Qian, W., Fang, X., Shi, Z., Li, Y., Li, S., Shan, G., Kristiansen, K., Yang, H., Wang, J., 2010. De novo assembly of human genomes with massively parallel short read sequencing. Genome Res. 20, 265–272. Li, X., Yu, T., He, Q., McDowall, K.J., Jiang, B., Jiang, Z., Wu, L., Li, G., Li, Q., Wang, S., Shi, Y., Wang, L., Hong, B., 2015. Binding of a biosynthetic intermediate to AtrA modulates the production of lidamycin by Streptomyces globisporus. Mol. Microbiol. 96, 1257–1271. Liu, W., Christenson, S.D., Standage, S., Shen, B., 2002. Biosynthesis of the enediyne antitumor antibiotic C-1027. Science 297, 1170–1173. Ma, D., 2007. Roles of sgcR and sgcM in Regulation of Enediyne Antibiotic C-1027 Production of Streptomyces globisporus C-1027. Peking Union Medical College, Beijing, pp. p. 83. Shao, R.G., Zhen, Y.S., 2008. Enediyne anticancer antibiotic lidamycin: chemistry, biology and pharmacology. Anticancer Agents Med. Chem. 8, 123–131. Wang, L., 2008. Roles of sgcR1, sgcR2 and sgcR3 in Positive Regulation of Enediyne Antibiotic C-1027 Production of Streptomyces globisporus C-1027. Peking Union Medical College, Beijing, p. 135. Weber, T., Blin, K., Duddela, S., Krug, D., Kim, H.U., Bruccoleri, R., Lee, S.Y., Fischbach, M.A., Muller, R., Wohlleben, W., Breitling, R., Takano, E., Medema, M.H., 2015. antiSMASH 3.0-a comprehensive resource for the genome mining of biosynthetic gene clusters. Nucleic Acids Res. 43, W237–243.
Contents lists available at ScienceDirect
Journal of Biotechnology journal homepage: www.elsevier.com/locate/jbiotec
Genome announcement
Complete genome sequence of Streptomyces globisporus C-1027, the producer of an enediyne antibiotic lidamycin Xingxing Li, Xuan Lei, Cong Zhang, Zhibo Jiang, Yuanyuan Shi, Songmei Wang, Lifei Wang ∗∗ , Bin Hong ∗ Key Laboratory of Biotechnology of Antibiotics of Ministry of Health, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China
a r t i c l e
i n f o
Article history: Received 25 January 2016 Accepted 2 February 2016 Available online 4 February 2016 Keywords: Streptomyces globisporus C-1027 Genome sequence Lidamycin
a b s t r a c t Streptomyces globisporus C-1027 produces a nine-membered enediyne antitumor antibiotic lidamycin. Here we report the complete genome sequence of S. globisporus C-1027, which consists of a 7,608,611 bp linear chromosome, a 167,754 bp linear plasmid SGLP1 and a 7,234 bp circular plasmid pSGL1. The biosynthetic gene cluster for lidamycin was located in the linear plasmid SGLP1. Genome analysis also revealed a number of genes related to biosynthesis of diverse secondary metabolites. The genome sequence of C-1027 will enable us to disclose biosynthetic pathways of these secondary metabolites and discover new natural products with potential applications notably in human health. © 2016 Elsevier B.V. All rights reserved.
Streptomyces globisporus C-1027 (CPCC 200270, deposited at the China Pharmaceutical Culture Collection) is a Gram-positive bacterium, originally isolated from a soil sample collected in the Hubei Province of China (Hu et al., 1988). It produces a potent antitumor antibiotic lidamycin, originally called C-1027, which belongs to the nine-membered enediyne family and is investigated in Phase II clinical trials in China (Shao and Zhen, 2008). Lidamycin biosynthetic gene cluster has been cloned and characterized (Liu et al., 2002). Its biosynthesis regulated by a complex circuit involving at least four cluster-situated regulators: SgcR1, SgcR2, SgcR3 and SgcR (Chen et al., 2010, 2011; Ma, 2007; Wang, 2008). Recently, a pleiotropic regulator AtrA, located outside of the lidamycin cluster, was characterized to have a role in regulating lidamycin production (Li et al., 2015). The complete genome sequence information of S. globisporus C-1027 may contribute to the further understanding of the global regulatory mechanisms for lidamycin biosynthesis. The complete genome was obtained by assembling a combination of reads using illumine Hiseq 2000 sequence platform. A total of 622 contigs were ordered using SOAPdenovo 1.05 (Li et al., 2010), and aligned with paired BAC end sequences using synteny with the genome of S. griseus subsp. griseus NBRC 13350 (GenBank accession no. AP009493), which is closely related to S. globisporus C-1027.
∗ Corresponding author. Fax: +86 10 63017302. ∗∗ Corresponding author. Fax: +86 10 63017302. E-mail addresses: [email protected], [email protected] (L. Wang), [email protected], [email protected] (B. Hong). http://dx.doi.org/10.1016/j.jbiotec.2016.02.004 0168-1656/© 2016 Elsevier B.V. All rights reserved.
Table 1 Genomic features of S. globisporus C-1027. Features
Chromosome
Plasmid SGLP1
Plasmid pSGL1
Length (bp) G + C content (%) CDS rRNA genes (operons) tRNA genes GenBank accession no.
7,608,611 71.6 7,078 18 (6) 66 CP013738
167,754 69.5 157 0 (0) 0 CP013739
7,234 70.9 9 0 (0) 0 CP013740
Finally, gaps were closed by PCR with Sanger sequencing. Putative protein-coding sequences were predicted using the Glimmer 3.0 (Delcher et al., 2007). Gene functional annotation was performed using BLASTP with KEGG, COG, Swiss-Port, TrEMBL, NR and GO databases. tRNA-encoding genes and rRNA operons were found by using tRNAscan software and rRNAmmer software. Repeat Masker software and CRISPR Finder software were used to find putative transposons and clustered regularly interspaced short palindromic repeats (CRISPRs). The genomic features are summarized in Table 1. S. globisporus C-1027 has a genome consisting of three replicons, a linear chromosome, a linear plasmid and a circular plasmid (Table 1). The linear chromosome is 7,608,611 bp with a 71.6% G + C content. 7,078 putative protein CDSs, 66 tRNA, 6 rRNA operons with the order 16S-23S-5S, 1 sRNA, and 19 CRISPRs were predicted on the chromosome. The large linear plasmid SGLP1, of 167,754 bp with a 69.5% G + C content, contains 4 CRISPRs and 157 putative genes, including the intact biosynthetic gene cluster for lidamycin. The circular plasmid pSGL1 (7,234 bp, 70.9% G + C content)
10
X. Li et al. / Journal of Biotechnology 222 (2016) 9–10
contains 9 putative CDS. Altogether 7,244 putative protein CDSs were identified in the genome containing 6,879,372 bp, accounting for 88.38% of the genome. On the complete genome, 117,075 bp repeat sequence and putative 49,291 bp transposon were found. Genome mining analysis using antiSMASH 3.0 (Weber et al., 2015) revealed the presence of 33 putative gene clusters responsible for the production of diverse secondary metabolites in the complete genome sequence of S. globisporus C-1027. Of these, the putative isorenieratene, griseobactin, desferrioxamines, ectoine and melanin are highly conserved in the closely related species S. griseus subsp. griseus NBRC 13350 and S. coelicolor A3(2). Some gene clusters related to biosynthesis of terpene, non-ribosomal peptide, polyketide and lantipeptide are not highly conserved with any known cluster, disclosing the potential of S. globisporus C-1027 to produce some new natural products. Nucleotide sequence accession numbers The complete genome sequence of S. globisporus C-1027 (CPCC 200270) has been deposited at GenBank under accession nos. CP013738, CP013739 and CP013740. Acknowledgments We thank Beijing Genomics Institute (Shenzhen, China) for illumine Hiseq sequencing, assembly and annotation of the genome. This work was supported by the National Natural Science Foundation of China (81302677, 81321004, 81402836, and 31170042), and National Mega-Project for Innovative Drugs (2012ZX09301002001-016 and 2015ZX09102007-016).
References Chen, Y., Yin, M., Horsman, G.P., Huang, S., Shen, B., 2010. Manipulation of pathway regulation in Streptomyces globisporus for overproduction of the enediyne antitumor antibiotic C-1027. J. Antibiot. (Tokyo) 63, 482–485. Chen, Y., Yin, M., Horsman, G.P., Shen, B., 2011. Improvement of the enediyne antitumor antibiotic C-1027 production by manipulating its biosynthetic pathway regulation in Streptomyces globisporus. J. Nat. Prod. 74, 420–424. Delcher, A.L., Bratke, K.A., Powers, E.C., Salzberg, S.L., 2007. Identifying bacterial genes and endosymbiont DNA with Glimmer. Bioinformatics 23, 673–679. Hu, J.L., Xue, Y.C., Xie, M.Y., Zhang, R., Otani, T., Minami, Y., Yamada, Y., Marunaka, T., 1988. A new macromolecular antitumor antibiotic, C-1027. I. Discovery, taxonomy of producing organism, fermentation and biological activity. J. Antibiot. (Tokyo) 41, 1575–1579. Li, R., Zhu, H., Ruan, J., Qian, W., Fang, X., Shi, Z., Li, Y., Li, S., Shan, G., Kristiansen, K., Yang, H., Wang, J., 2010. De novo assembly of human genomes with massively parallel short read sequencing. Genome Res. 20, 265–272. Li, X., Yu, T., He, Q., McDowall, K.J., Jiang, B., Jiang, Z., Wu, L., Li, G., Li, Q., Wang, S., Shi, Y., Wang, L., Hong, B., 2015. Binding of a biosynthetic intermediate to AtrA modulates the production of lidamycin by Streptomyces globisporus. Mol. Microbiol. 96, 1257–1271. Liu, W., Christenson, S.D., Standage, S., Shen, B., 2002. Biosynthesis of the enediyne antitumor antibiotic C-1027. Science 297, 1170–1173. Ma, D., 2007. Roles of sgcR and sgcM in Regulation of Enediyne Antibiotic C-1027 Production of Streptomyces globisporus C-1027. Peking Union Medical College, Beijing, pp. p. 83. Shao, R.G., Zhen, Y.S., 2008. Enediyne anticancer antibiotic lidamycin: chemistry, biology and pharmacology. Anticancer Agents Med. Chem. 8, 123–131. Wang, L., 2008. Roles of sgcR1, sgcR2 and sgcR3 in Positive Regulation of Enediyne Antibiotic C-1027 Production of Streptomyces globisporus C-1027. Peking Union Medical College, Beijing, p. 135. Weber, T., Blin, K., Duddela, S., Krug, D., Kim, H.U., Bruccoleri, R., Lee, S.Y., Fischbach, M.A., Muller, R., Wohlleben, W., Breitling, R., Takano, E., Medema, M.H., 2015. antiSMASH 3.0-a comprehensive resource for the genome mining of biosynthetic gene clusters. Nucleic Acids Res. 43, W237–243.
