Enhancement of toyocamycin production through increasing supply of precursor GTP in <i>Streptomyces diastatochromogenes</i> 1628

Zhang, Jinyao; Xu, Jie; Li, Huijie; Zhang, Yongyong; Ma, Zheng; Bechthold, Andreas; Yu, Xiaoping

doi:10.1002/jobm.202100622

Cited by 2 publications

(2 citation statements)

References 34 publications

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“…(38), cytoskeletons from mNG::PKAC1 expressing cells were incubated with cAMP or toyocamycin prior to imaging and quantification of axonemal fluorescence (Figure 4). In addition to these, we tested the effect of inosine, considering that it is similar in structure to toyocamycin (which derives from Streptomyces (68)) but perhaps more biologically relevant since it is present endogenously as part of purine metabolism pathways (69).…”

Section: Resultsmentioning

confidence: 99%

Divergent Protein Kinase A contributes to the regulation of flagellar waveform of the motile flagellum ofLeishmania mexicana

Fochler,

Walker,

Wheeler

et al. 2023

Preprint

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Protein kinase A (PKA) is one of the most studied eukaryotic signal transducers and a key target of the second messenger cAMP. Kinetoplastids, a branch of early diverging eukaryotes, possess homologues of catalytic and regulatory PKA subunits that, unusually, are insensitive to cAMP with uncertainty about the endogenous activating ligand. While previous evidence has implicated both cAMP and PKA in the regulation of flagellar motility in kinetoplastids, the specific contributions of the two regulatory (PKAR) and three catalytic (PKAC) subunits of PKA remain unclear. Here we conducted a systematic study combining reverse genetics and microscopy analysis to investigate PKAC and PKAR subcellular localisations, pairing preferences, and the effect of PKA gene deletions on swimming speed and flagellar waveforms inLeishmania mexicanapromastigotes.LmxPKAC1,LmxPKAC2 andLmxPKAR1 are enriched in the flagellum and upon detergent extraction, theLmxPKAR1 andLmxPKAC1 signals remained associated with the cytoskeleton. The flagellarLmxPKAC1 andLmxPKAC2 signals were, however, greatly diminished by removal ofLmxPKAR1, consistent with an anchoring function for the R-subunit. Previous work identified theStreptomycesantimetabolite toyocamycin, activated the divergent PKA ofTrypanosoma brucei. The patterns of sequence divergence within the phosphate binding cassettes ofLmxPKAR1 andLmxPKAR3 suggest both are divergent fromT. brucei. We therefore tested the effect of toyocamycin and a similarly structured compound of the purine metabolism pathway, inosine, on the localisation ofLmxPKAC1 and found that theLmxPKAC1 mNG signal was lost from the cytoskeletal fraction following treatment with either toyocamycin or inosine.LmxPKAC3 andLmxPKAR3 were localised to the cell cortex. Upon removal ofLmxPKAR3,LmxPKAC3 was lost from the cortex and became distributed throughout the cell, including an increased presence in the flagellum. Assessing motility phenotypes, we found this increase ofLmxPKAC3 signal in the flagellum correlated with an increase in swimming speed. Whereas the deletion ofLmxPKAC3 resulted in flagella beating at significantly reduced frequencies and a resulting decrease in population swimming speed. Moreover,LmxPKAC1 null mutants showed a significantly reduced swimming speed and cells were unable to produce symmetric flagellar waves. Taken together these data indicate that the divergent PKA pathway inLeishmaniamodulates flagellar motility.Author SummaryMany cells use whip-like cellular appendages, called flagella, for swimming. Regulating the rhythm and strength of the flagellar beat is important to determine the speed at which cells swim and in what direction they move. How this regulation is accomplished is not fully understood. Here we examined the role of protein kinase A (PKA) in flagellar motility of the unicellular parasite Leishmania. Leishmania have three catalytic PKA proteins, which transduce upstream signals through protein phosphorylation, and two regulatory PKA proteins that serve to anchor the catalytic subunits to specific locations within the cell. We show that two catalytic and one regulatory subunit are part of the flagellar cytoskeleton, while the other regulatory subunit docks its catalytic subunit to the cytoskeleton surrounding the cell body. Removal of individual PKA proteins perturbed the flagellar beat in different ways. We found that cells that lacked the catalytic subunit PKAC1 could only beat their flagella in an uncoordinated manner and as a consequence swam more slowly. These findings suggest that Leishmania PKA proteins are part of a pathway that regulates flagellar beating.

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Section: Resultsmentioning

confidence: 99%

Divergent Protein Kinase A contributes to the regulation of flagellar waveform of the motile flagellum ofLeishmania mexicana

Fochler,

Walker,

Wheeler

et al. 2023

Preprint

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“…S. diastatochromogenes 1628 produces guanosine triphosphate (GTP) as a precursor for nucleoside antibiotic toyocamycin (TM) with potent activity against a wide range of plant pathogenic fungi. The differentially expressed protein X0NBV6 for ribose-phosphate pyrophosphokinase (RHP) is involved in GTP biosynthesis and was expressed at a higher expression level in mutant 1628-T15, and the yield of TM produced by 1628-RHP (340.2 mg/L) was 133.3% higher than that produced by S. diastatochromogenes 1628 (228%) [ 26 ]. Both lincomycin and melanin use l-tyrosine as precursor, and the regulator AdpA lin in S. lincolnensis is a pleiotropic transcriptional regulator.…”

Section: Effect Of Precursors On Antibiotic Biosynthesismentioning

confidence: 99%

Effect of Precursors and Their Regulators on the Biosynthesis of Antibiotics in Actinomycetes

Yan,

Dong,

et al. 2024

Molecules

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During the life activities of microorganisms, a variety of secondary metabolites are produced, including antimicrobials and antitumor drugs, which are widely used in clinical practice. In addition to exploring new antibiotics, this makes it one of the research priorities of Actinomycetes to effectively increase the yield of antibiotics in production strains by various means. Most antibiotic-producing strains have a variety of functional regulatory factors that regulate their growth, development, and secondary metabolite biosynthesis processes. Through the study of precursor substances in antibiotic biosynthesis, researchers have revealed the precursor biosynthesis process and the mechanism by which precursor synthesis regulators affect the biosynthesis of secondary metabolites, which can be used to obtain engineered strains with high antibiotic production. This paper summarizes the supply of antibiotic biosynthesis precursors and the progress of research on the role of regulators in the process of precursors in biosynthesis. This lays the foundation for the establishment of effective breeding methods to improve antibiotic yields through the manipulation of precursor synthesis genes and related regulators.

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Enhancement of toyocamycin production through increasing supply of precursor GTP in Streptomyces diastatochromogenes 1628

Cited by 2 publications

References 34 publications

Divergent Protein Kinase A contributes to the regulation of flagellar waveform of the motile flagellum ofLeishmania mexicana

Divergent Protein Kinase A contributes to the regulation of flagellar waveform of the motile flagellum ofLeishmania mexicana

Effect of Precursors and Their Regulators on the Biosynthesis of Antibiotics in Actinomycetes

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