Regulation of Geldanamycin Biosynthesis by Cluster-Situated Transcription Factors and the Master Regulator PhoP

Abstract

Geldanamycin and the closely related herbimycins A, B, and C are benzoquinone-type ansamycins with antitumoral activity. They are produced by Streptomyces hygroscopicus var. geldanus, Streptomyces lydicus and Streptomyces autolyticus among other Streptomyces strains. Geldanamycins interact with the Hsp-90 chaperone, a protein that has a key role in tumorigenesis of human cells. Geldanamycin is a polyketide antibiotic and the polyketide synthase contain seven modules organized in three geldanamycin synthases genes named gdmAI, gdmAII, and gdmAIII. The loading domain of GdmI activates AHBA, and also related hydroxybenzoic acid derivatives, forming geldanamycin analogues. Three regulatory genes, gdmRI, gdmRII, and gdmRIII were found associated with the geldanamycin gene cluster in S. hygroscopicus strains. GdmRI and GdmRII are LAL-type (large ATP binding regulators of the LuxR family) transcriptional regulators, while GdmRIII belongs to the TetR-family. All three are positive regulators of geldanamycin biosynthesis and are strictly required for expression of the geldanamycin polyketide synthases. In S. autolyticus the gdmRIII regulates geldanamycin biosynthesis and also expression of genes in the elaiophylin gene cluster, an unrelated macrodiolide antibiotic. The biosynthesis of geldanamycin is very sensitive to the inorganic phosphate concentration in the medium. This regulation is exerted through the two components system PhoR-PhoP. The phoRP genes of S. hygroscopicus are linked to phoU encoding a transcriptional modulator. The phoP gene was deleted in S. hygroscopicus var geldanus and the mutant was unable to grow in SPG medium unless supplemented with 5 mM phosphate. Also, the S. hygroscopicus pstS gene involved in the high affinity phosphate transport was cloned, and PhoP binding sequences (PHO boxes), were found upstream of phoU, phoRP, and pstS; the phoRP-phoU sequences were confirmed by EMSA and nuclease footprinting protection assays. The PhoP binding sequence consists of 11 nucleotide direct repeat units that are similar to those found in S. coelicolor Streptomyces avermitilis and other Streptomyces species. The available genetic information provides interesting tools for modification of the biosynthetic and regulatory mechanisms in order to increase geldanamycin production and to obtain new geldanamycin analogues with better antitumor properties.

SenX3-RegX3, an Important Two-Component System, Regulates Strain Growth and Butenyl-spinosyn Biosynthesis in Saccharopolyspora pogona

“…The PhoR-PhoP system is conserved in all Streptomyces and most other sequenced actinomycetes ( Barreiro and Martínez-Castro, 2019 ). Many studies have demonstrated that this system plays an important regulatory role in cellular differentiation and secondary metabolite biosynthesis in actinomycetes ( Sola-Landa et al., 2003 , 2005 , 2013 ; Martín et al., 2019 ). In another system that senses and responds to phosphate, the sensor kinase SenX3 and the response regulator RegX3 were first identified and have been well studied in Mycobacterium ( Namugenyi et al., 2017 ; Rifat et al., 2014 ; Park et al., 2019 ).…”

The Application of Regulatory Cascades in Streptomyces: Yield Enhancement and Metabolite Mining

“…Overexpression of LAL-type regulatory factor samR in Streptomyces ambofaciens activated a 150 kb cryptic gene cluster, and promoted the discovery of a glycosylated macrolide product with anti-cancer activity (Laureti et al, 2011). Similarly, LAL-type transcriptional regulators of GdmRI and GdmRII in S. hygroscopicus are positive to geldanamycin biosynthesis (Martin et al, 2019). Furthermore, the LAL-like 63-amino acid protein AfsS and its homologs have been shown to relate closely with the induction of avermectin in S. avermitilis (Lee et al, 2000), pikromycin in S. venezuelae (Maharjan et al, 2009), and DXR in S. peucetius (Parajuli et al, 2005).…”

Identification of a Novel Bioactive Phenazine Derivative and Regulation ofphoPon Its Production inStreptomyces lomondensisS015

“…avermitilis, phoP negatively regulates the avermectin production, whereas phoP does not affect the production of oligomycin . The deletion of phoP appears to strongly inhibit cell growth but increase the production of geldanamycin in Streptomyces hygroscopicus . Moreover, the filipin production in Streptomyces filipinensis was greatly improved by deleting phoP .…”

“…These results indicated that phoP may participate in the regulation of secondary metabolites in S. lomondensis S015 as reported elsewhere. 29,31 To investigate the effects of phoP on CFTHP biosynthesis, it was disrupted in S. lomondensis S015 through a 466 bp internal deletion, as shown in Figure 4A. Also, the resulting strain S015ΔphoP was confirmed by PCR amplification (Figure 4B).…”

“…The regulatory genes are quite conserved in Streptomyces that phoP sequences among S. lividans, Streptomyces coelicolor, and Streptomyces avermitilis share highly identity. , Although several studies have developed deeper understanding about the influence of PhoP on the production of avermectin, filipin, geldanamycin, etc., − the regulation of PhoP in respect of phenazine biosynthesis still remains unclear.…”