Global analysis of genomic instability caused by DNA replication stress in Saccharomyces cerevisiae

One important source of genomic instability associated with tumor cells is DNA replication stress. In the current study, replication stress was induced in yeast by a 10-fold reduction in the level of the replicative DNA polymerase δ. By DNA microarray analysis and high-throughput DNA sequencing, we showed that this stress resulted in very high rates of both large (aneuploidy, mitotic recombination, deletions and duplications, and translocations) and small (point mutations and small insertion/deletions) genetic alterations. Some of these changes resulted in a selective growth advantage of the cells, demonstrating the role of elevated genetic instability in the rapid evolution of cells in challenging growth conditions

New azaphilone, isocoumarin and α-pyrone derivatives from the marine-derived gut fungus Paraphaeosphaeria sp. XZD2-1

Six new compounds including two azaphilones, lunatoic acids D–E (1, 2), three isocoumarins, lunatinins B–D (3-5), and one α-pyrone derivative, lunatinin E (6), as well as four known ones, lunatoic acid A (7), lunatinin (8), penicipyran D (9) and chaetoquadrin F (10) were isolated from the rice medium of the gut fungus Paraphaeosphaeria sp. XZD2-1. Their planar structures were elucidated by analysis of 1D and 2D NMR as well as HRESIMS spectroscopic data. The absolute configurations of compounds 1-3 were assigned by experimental and calculated ECD data and an ester hydrolysis reaction. Compounds 4 and 5 were a pair of enantiomeric excess mixture. Structure of compound 6 was further confirmed by comparing optical rotation with known compounds according to literature. The antimicrobial activity and cytotoxicity of compounds 1-10 were examined

Global analysis of genomic instability caused by DNA replication stress in Saccharomyces cerevisiae

DNA replication stress (DRS)-induced genomic instability is an important factor driving cancer development. To understand the mechanisms of DRS-associated genomic instability, we measured the rates of genomic alterations throughout the genome in a yeast strain with lowered expression of the replicative DNA polymerase δ. By a genetic test, we showed that most recombinogenic DNA lesions were introduced during S or G(2) phase, presumably as a consequence of broken replication forks. We observed a high rate of chromosome loss, likely reflecting a reduced capacity of the low-polymerase strains to repair double-stranded DNA breaks (DSBs). We also observed a high frequency of deletion events within tandemly repeated genes such as the ribosomal RNA genes. By whole-genome sequencing, we found that low levels of DNA polymerase δ elevated mutation rates, both single-base mutations and small insertions/deletions. Finally, we showed that cells with low levels of DNA polymerase δ tended to accumulate small promoter mutations that increased the expression of this polymerase. These deletions conferred a selective growth advantage to cells, demonstrating that DRS can be one factor driving phenotypic evolution

Genome sequencing and genetic breeding of a bioethanol <it>Saccharomyces cerevisiae</it> strain YJS329

Abstract Background Environmental stresses and inhibitors encountered by Saccharomyces cerevisiae strains are the main limiting factors in bioethanol fermentation. Strains with different genetic backgrounds usually show diverse stress tolerance responses. An understanding of the mechanisms underlying these phenotypic diversities within S. cerevisiae populations could guide the construction of strains with desired traits. Results We explored the genetic characteristics of the bioethanol S. cerevisiae strain YJS329 and elucidated how genetic variations in its genome were correlated with specified traits compared to similar traits in the S288c-derived strain, BYZ1. Karyotypic electrophoresis combined with array-comparative genomic hybridization indicated that YJS329 was a diploid strain with a relatively constant genome as a result of the fewer Ty elements and lack of structural polymorphisms between homologous chromosomes that it contained. By comparing the sequence with the S288c genome, a total of 64,998 SNPs, 7,093 indels and 11 unique genes were identified in the genome of YJS329-derived haploid strain YJSH1 through whole-genome sequencing. Transcription comparison using RNA-Seq identified which of the differentially expressed genes were the main contributors to the phenotypic differences between YJS329 and BYZ1. By combining the results obtained from the genome sequences and the transcriptions, we predicted how the SNPs, indels and chromosomal copy number variations may affect the mRNA expression profiles and phenotypes of the yeast strains. Furthermore, some genetic breeding strategies to improve the adaptabilities of YJS329 were designed and experimentally verified. Conclusions Through comparative functional genomic analysis, we have provided some insights into the mechanisms underlying the specific traits of the bioenthanol strain YJS329. The work reported here has not only enriched the available genetic resources of yeast but has also indicated how functional genomic studies can be used to improve genetic breeding in yeast.</p

Nonlethal Furfural Exposure Causes Genomic Alterations and Adaptability Evolution in Saccharomyces cerevisiae

ABSTRACT Furfural is a major inhibitor found in lignocellulosic hydrolysate, a promising feedstock for the biofermentation industry. In this study, we aimed to investigate the potential impact of this furan-derived chemical on yeast genome integrity and phenotypic evolution by using genetic screening systems and high-throughput analyses. Our results showed that the rates of aneuploidy, chromosomal rearrangements (including large deletions and duplications), and loss of heterozygosity (LOH) increased by 50-fold, 23-fold, and 4-fold, respectively, when yeast cells were cultured in medium containing a nonlethal dose of furfural (0.6 g/L). We observed significantly different ratios of genetic events between untreated and furfural-exposed cells, indicating that furfural exposure induced a unique pattern of genomic instability. Furfural exposure also increased the proportion of CG-to-TA and CG-to-AT base substitutions among point mutations, which was correlated with DNA oxidative damage. Interestingly, although monosomy of chromosomes often results in the slower growth of yeast under spontaneous conditions, we found that monosomic chromosome IX contributed to the enhanced furfural tolerance. Additionally, terminal LOH events on the right arm of chromosome IV, which led to homozygosity of the SSD1 allele, were associated with furfural resistance. This study sheds light on the mechanisms underlying the influence of furfural on yeast genome integrity and adaptability evolution. IMPORTANCE Industrial microorganisms are often exposed to multiple environmental stressors and inhibitors during their application. This study demonstrates that nonlethal concentrations of furfural in the culture medium can significantly induce genome instability in the yeast Saccharomyces cerevisiae. Notably, furfural-exposed yeast cells displayed frequent chromosome aberrations, indicating the potent teratogenicity of this inhibitor. We identified specific genomic alterations, including monosomic chromosome IX and loss of heterozygosity of the right arm of chromosome IV, that confer furfural tolerance to a diploid S. cerevisiae strain. These findings enhance our understanding of how microorganisms evolve and adapt to stressful environments and offer insights for developing strategies to improve their performance in industrial applications

Summary of changes in chromosome number in MD741 and MD704 isolates.

(XLSX)</p

Nanopore analysis of chromosome rearrangements in MD704-2h-1.

The I-DEL (coordinates 489–514 kb) on chromosome IV, evident by sequencing, has the same Ty-associated breakpoints as in MD741-5 (S9B Fig). The Ty elements that are at the breakpoints of the YJM789-derived homolog, but are absent in S288c are shown as a gap. (TIF)</p

Patterns of LOH and I-DELs observed in isolate MD704-4h-6P.

(A) Chromosome III has a duplication of the sequences between Ty elements located in FS1 and FS2 in the W303-1A homolog. (B) Chromosome VII has a T-DEL with a breakpoint in a Ty located near coordinate 536 kb. (C) Chromosome XIII has a complex pattern of alterations (breakpoints of each transition in parentheses): 1. Triplication of W303-1A-derived sequences (from left end of chromosome to 184 kb), 2. One copy of W303-1A and YJM789 sequences (184–280 kb), 3. Two copies of YJM789-derived sequences and no copies of W303-1A-derived sequences (280–372 kb), and 4. One copy of YJM789-derived sequences and no copies of W303-1A-derived sequences (372 kb to the right end of chromosome). (TIF)</p