ABSTRACT
A synthesis program for structurally complex macrocycles is very challenging. Herein, we propose a biosynthesis pathway of the pyranylated cyclodepsipeptide verucopeptin to make enough supply and to diversify verucopeptin by genetic manipulation and one-step semisynthesis. The synthesis relies on the intrinsic reactivity of the interchangeable hemiketal pyrane and opened keto along with adjacent alkene. Biological evaluation of verucopeptin-oriented analogs delivers a potent AMP-activated protein kinase (AMPK) agonist, antibacterial agent, and selective NFκB modulator.
Subject(s)
AMP-Activated Protein Kinases/metabolism , Anti-Bacterial Agents/pharmacology , Depsipeptides/pharmacology , NF-kappa B/antagonists & inhibitors , Protein Kinase Inhibitors/pharmacology , Staphylococcus aureus/drug effects , Actinomadura/chemistry , Anti-Bacterial Agents/biosynthesis , Anti-Bacterial Agents/chemistry , Depsipeptides/biosynthesis , Depsipeptides/chemistry , HEK293 Cells , Humans , Microbial Sensitivity Tests , Molecular Structure , NF-kappa B/metabolism , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/metabolism , Signal Transduction/drug effectsABSTRACT
Candida glabrata is a promising microorganism for the production of organic acids. Here, we report deletion and quantitative-expression approaches to elucidate the role of C. glabrata Med3AB (CgMed3AB), a subunit of the mediator transcriptional coactivator, in regulating cell growth. Deletion of CgMed3AB caused an 8.6% decrease in final biomass based on growth curve plots and 10.5% lower cell viability. Based on transcriptomics data, the reason for this growth defect was attributable to changes in expression of genes involved in pyruvate and acetyl-coenzyme A (CoA)-related metabolism in a Cgmed3abΔ strain. Furthermore, the mRNA level of acetyl-CoA synthetase was downregulated after deleting Cgmed3ab, resulting in 22.8% and 21% lower activity of acetyl-CoA synthetase and cellular acetyl-CoA, respectively. Additionally, the mRNA level of CgCln3, whose expression depends on acetyl-CoA, was 34% lower in this strain. As a consequence, the cell size and budding index in the Cgmed3abΔ strain were both reduced. Conversely, overexpression of Cgmed3ab led to 16.8% more acetyl-CoA and 120% higher CgCln3 mRNA levels, as well as 19.1% larger cell size and a 13.3% higher budding index than in wild-type cells. Taken together, these results suggest that CgMed3AB regulates cell growth in C. glabrata by coordinating homeostasis between cellular acetyl-CoA and CgCln3.IMPORTANCE This study demonstrates that CgMed3AB can regulate cell growth in C. glabrata by coordinating the homeostasis of cellular acetyl-CoA metabolism and the cell cycle cyclin CgCln3. Specifically, we report that CgMed3AB regulates the cellular acetyl-CoA level, which induces the transcription of Cgcln3, finally resulting in alterations to the cell size and budding index. In conclusion, we report that CgMed3AB functions as a wheel responsible for driving cellular acetyl-CoA metabolism, indirectly inducing the transcription of Cgcln3 and coordinating cell growth. We propose that Mediator subunits may represent a vital regulatory target modulating cell growth in C. glabrata.
