To gain a deep understanding of yeast-cell response to heat stress, multiple laboratory strains have been intensively studied via genome-wide expression analysis for the mechanistic dissection of classical heat-shock response (HSR). However, robust industrial strains of Saccharomyces cerevisiae have hardly been explored in global analysis for elucidation of the mechanism of thermotolerant response (TR) during fermentation. Herein, we employed data-independent acquisition and sequential window acquisition of all theoretical mass spectra based proteomic workflows to characterize proteome remodeling of an industrial strain, ScY01, responding to prolonged thermal stress or transient heat shock. By comparing the proteomic signatures of ScY01 in TR versus HSR as well as the HSR of the industrial strain versus a laboratory strain, our study revealed disparate response mechanisms of ScY01 during thermotolerant growth or under heat shock. In addition, through proteomics data-mining for decoding transcription factor interaction networks followed by validation experiments, we uncovered the functions of two novel transcription factors, Mig1 and Srb2, in enhancing the thermotolerance of the industrial strain. This study has demonstrated that accurate and high-throughput quantitative proteomics not only provides new insights into the molecular basis for complex microbial phenotypes but also pinpoints upstream regulators that can be targeted for improving the desired traits of industrial microorganisms.
Phycocyanin (PC) is a pigment-protein complex. It has been reported that PC exerts anti-colorectal cancer activities, although the underlying mechanism has not been fully elucidated. In the present study, azoxymethane (AOM)/dextran sulfate sodium (DSS)-induced mice were orally administrated with PC, followed by microbiota and transcriptomic analyses to investigate the effects of PC on colitis-associated cancer (CAC). Our results indicated that PC ameliorated AOM/DSS induced inflammation. PC treatment significantly reduced the number of colorectal tumors and inhibited proliferation of epithelial cell in CAC mice. Moreover, PC reduced the relative abundance of Firmicutes, Deferribacteres, Proteobacteria and Epsilonbacteraeota at phylum level. Transcriptomic analysis showed that the expression of genes involved in the intestinal barrier were altered upon PC administration, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed the IL-17 signaling pathway was affected by PC treatment. The study demonstrated the protective therapeutic action of PC on CAC.
Sugarcane smut, caused by Sporisorium scitamineum, is one of the most devastating
fungal diseases affecting sugarcane
worldwide. To develop a potent sugarcane smut fungicide, secondary
metabolites of marine-derived Bacillus siamensis were isolated and screened for inhibitory activities, which led
to the discovery of five new 24-membered macrolactins, bamemacrolactins
A-E (1–5), with 3 being
the most potent inhibitor. The antifungal mechanism of 3 was studied by assessing its effects on mycelial morphology and
the cell wall. Differential proteomics were used to analyze proteins
in S. scitamineum upon treatment with
bamemacrolactin C and to elucidate its antifungal mechanism. A total
of 533 differentially expressed proteins were found. After the Gene
Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses,
eight target proteins were selected, and their functions were discussed.
Six of the eight proteins were reported as antifungal targets. The
target proteins are involved in the oxidative phosphorylation pathway.
Therefore, the potent inhibition of S. scitamineum by compound 3 is most likely through oxidative phosphorylation
and targeting a series of enzymes.
TALENs-assisted multiplex editing (TAME) toolbox was previously established and used to successfully enhance ethanol stress tolerance of Saccharomyces cerevisiae laboratory strain. Here, the TAME toolbox was harnessed to improve and elucidate stress tolerances of S. cerevisiae industrial strain. One osmotolerant strain and one thermotolerant strain were selected from the mutant library generated by TAME at corresponding stress conditions, and exhibited 1.2-fold to 1.3-fold increases of fermentation capacities, respectively. Genome resequencing uncovered genomic alterations in the selected stress-tolerant strains, suggesting that cell wall and membrane-related proteins might be major factors behind improved tolerances of yeast to different stresses. Furthermore, amplified mitochondrial DNA might also have an important impact on increased stress tolerance. Unexpectedly, none of predesigned target potential TALENs modification sites showed any genomic variants in sequenced genomes of the selected strains, implicating that the improved stress tolerances might be due to indirect impacts of genome editing via TALENs rather than introducing genomic variants at potential target sites. Our findings not only confirmed TAME could be a useful tool to accelerate the breeding of industrial strain with multiple stress tolerance, but also supported the previous understandings of the complicated mechanisms of multiple stress tolerance in yeast.
Background
Macrolactins, a type of macrolide antibiotic, are toxic to the producer strains. As such, its level is usually maintained below the lethal concentration during the fermentation process. To improve the production of macrolactins, we applied adaptive laboratory evolution technology to engineer a saline-resistant mutant strain. The hypothesis that strains with saline resistance show improved macrolactins production was investigated.
Results
Using saline stress as a selective pressure, we engineered a mutant strain with saline resistance coupled with enhanced macrolactins production within 60 days using a self-made device. As compared with the parental strain, the evolved strain produced macrolactins with 11.93% improvement in non-saline stress fermentation medium containing 50 g/L glucose, when the glucose concentration increased to 70 g/L, the evolved strain produced macrolactins with 71.04% improvement. RNA sequencing and metabolomics results revealed that amino acid metabolism was involved in the production of macrolactins in the evolved strain. Furthermore, genome sequencing of the evolved strain revealed a candidate mutation, hisDD41Y, that was causal for the improved MLNs production, it was 3.42 times higher than the control in the overexpression hisDD41Y strain. Results revealed that saline resistance protected the producer strain from feedback inhibition of end-product (macrolide antibiotic), resulting in enhanced MLNs production.
Conclusions
In the present work, we successfully engineered a mutant strain with enhanced macrolactins production by adaptive laboratory evolution using saline stress as a selective pressure. Based on physiological, transcriptomic and genetic analysis, amino acid metabolism was found to benefit macrolactins production improvement. Our strategy might be applicable to improve the production of other kinds of macrolide antibiotics and other toxic compounds. The identification of the hisD mutation will allow for the deduction of metabolic engineering strategies in future research.
Yeast cells suffer from continuous and long-term thermal stress during high-temperature ethanol fermentation. Understanding the mechanism of yeast thermotolerance is important not only for studying microbial stress biology in basic research but also for developing thermotolerant strains for industrial application. Here, we compared the effects of 23 transcription factor (TF) deletions on high-temperature ethanol fermentation and cell survival after heat shock treatment and identified three core TFs, Sin3p, Srb2p and Mig1p, that are involved in regulating the response to long-term thermotolerance. Further analyses of comparative transcriptome profiling of the core TF deletions and transcription regulatory associations revealed a hierarchical transcriptional regulatory network centered on these three TFs. This global transcriptional regulatory network provided a better understanding of the regulatory mechanism behind long-term thermal stress tolerance as well as potential targets for transcriptome engineering to improve the performance of high-temperature ethanol fermentation by an industrial Saccharomyces cerevisiae strain.
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