Non‐Canonical Replication Initiation: You’re Fired!

The division of prokaryotic and eukaryotic cells produces two cells that inherit a perfect copy of the genetic material originally derived from the mother cell. The initiation of canonical DNA replication must be coordinated to the cell cycle to ensure the accuracy of genome duplication. Controlled replication initiation depends on a complex interplay of cis‐acting DNA sequences, the so‐called origins of replication (ori), with trans‐acting factors involved in the onset of DNA synthesis. The interplay of cis‐acting elements and trans‐acting factors ensures that cells initiate replication at sequence‐specific sites only once, and in a timely order, to avoid chromosomal endoreplication. However, chromosome breakage and excessive RNA:DNA hybrid formation can cause breakinduced (BIR) or transcription‐initiated replication (TIR), respectively. These non‐canonical replication events are expected to affect eukaryotic genome function and maintenance, and could be important for genome evolution and disease development. In this review, we describe the difference between canonical and non‐canonical DNA replication, and focus on mechanistic differences and common features between BIR and TIR. Finally, we discuss open issues on the factors and molecular mechanisms involved in TIR.España, MINECO BFU2015-69183-

Saccharomyces paradoxus Transcriptional Alterations in Cells of Distinct Phenotype and Viral dsRNA Content

Killer yeasts are attractive antifungal agents with great potential applications in the food industry. Natural Saccharomyces paradoxus isolates provide new dsRNA-based killer systems available for investigation. The presence of viral dsRNA may alter transcriptional profile of S. paradoxus. To test this possibility, a high-throughput RNA sequencing was employed to compare the transcriptomes of S. paradoxus AML 15-66 K66 killer strains after curing them of either M-66 alone or both M-66 and L-A-66 dsRNA viruses. The S. paradoxus cells cured of viral dsRNA(s) showed respiration deficient or altered sporulation patterns. We have identified numerous changes in the transcription profile of genes including those linked to ribosomes and amino acid biosynthesis, as well as mitochondrial function. Our work advance studies of transcriptional adaptations of Saccharomyces spp. induced by changes in phenotype and set of dsRNA viruses, reported for the first time.This article belongs to the Special Issue Recent Advances in the Yeast Killer Systems ResearchThis research was partly funded by European Regional Funds (CZ.02.1.01/16_019/0000759)

Saccharomyces genties mielių dgRNR virusų funkcionavimo tyrimai

Yeasts are fungal microorganisms distributed in various natural environments. Yeasts are found in atmospheric, aquatic, and terrestrial habitats and are the most abundant in a nutrient-rich environment, such as fruits, berries, stems, and flowers (Starmer and Lachance, 2011). Symbiotic relationships between yeasts and viruses, bacteria, fungi, plants, and animals are common. Yeast ability to produce killer toxins is an important factor, investigated in biocontrol, clinical, ecological, and evolutionary studies (Drinnenberg et al., 2011; Freimoser et al., 2019; Mannazzu et al., 2019; Schaffrath et al., 2018)

Conjugation of phosphonoacetic acid to nucleobase promotes a mechanism-based inhibition

Small molecule inhibitors have a powerful blocking action on viral polymerases. The bioavailability of the inhibitor, nevertheless, often raise a significant selectivity constraint and may substantially limit the efficacy of therapy. Phosphonoacetic acid has long been known to possess a restricted potential to block DNA biosynthesis. In order to achieve a better affinity, this compound has been linked with natural nucleotide at different positions. The structural context of the resulted conjugates has been found to be crucial for the acquisition by DNA polymerases. We show that nucleobase-conjugated phosphonoacetic acid is being accepted, but this alters the processivity of DNA polymerases. The data presented here not only provide a mechanistic rationale for a switch in the mode of DNA synthesis, but also highlight the nucleobase-targeted nucleotide functionalization as a route for enhancing the specificity of small molecule inhibitors

Adaptive response of Saccharomyces hosts to Totiviridae L-A dsRNA viruses Is achieved through intrinsically balanced action of targeted transcription factors

Totiviridae L-A virus is a widespread yeast dsRNA virus. The persistence of the L-A virus alone appears to be symptomless, but the concomitant presence of a satellite M virus provides a killer trait for the host cell. The presence of L-A dsRNA is common in laboratory, industrial, and wild yeasts, but little is known about the impact of the L-A virus on the host’s gene expression. In this work, based on high-throughput RNA sequencing data analysis, the impact of the L-A virus on whole-genome expression in three different Saccharomyces paradoxus and S. cerevisiae host strains was analyzed. In the presence of the L-A virus, moderate alterations in gene expression were detected, with the least impact on respiration-deficient cells. Remarkably, the transcriptional adaptation of essential genes was limited to genes involved in ribosome biogenesis. Transcriptional responses to L-A maintenance were, nevertheless, similar to those induced upon stress or nutrient availability. Based on these data, we further dissected yeast transcriptional regulators that, in turn, modulate the cellular L-A dsRNA levels. Our findings point to totivirus-driven fine-tuning of the transcriptional landscape in yeasts and uncover signaling pathways employed by dsRNA viruses to establish the stable, yet allegedly profitless, viral infection of fungi

Essential oils of Mentha arvensis and Cinnamomum cassia exhibit distinct antibacterial activity at different temperatures in vitro and on chicken skin

The bacterial contamination of meat is a global concern, especially for the risk of Salmonella infection that can lead to health issues. Artificial antibacterial compounds used to preserve fresh meat can have negative health effects. We investigated the potential of natural essential oils (EOs), namely Mentha arvensis (mint) and Cinnamomum cassia (cinnamon) EOs, to prevent contamination of the food pathogen, Salmonella enterica subsp. enterica serotype Typhimurium, in vitro and on chicken skin. The gas chromatography–mass spectrometry (GC-MS) technique was used to determine the compositions of mint EO (MEO) and cinnamon EO (CEO); the most abundant compound in MEO was menthol (68.61%), and the most abundant compound was cinnamaldehyde (83.32%) in CEO. The antibacterial activity of MEO and CEO were examined in vapor and direct contact with S. typhimurium at temperatures of 4 °C, 25 °C, and 37 °C. The minimal inhibitory concentration at 37 °C for MEO and CEO reached 20.83 µL/mL, and the minimal bactericidal concentration of CEO was the same, while for MEO, it was two-fold higher. We report that in most tested conditions in experiments performed in vitro and on chicken skin, CEO exhibits a stronger antibacterial effect than MEO. In the vapor phase, MEO was more effective against S. typhimurium than CEO at 4 °C. In direct contact, the growth of S. typhimurium was inhibited more efficiently by MEO than CEO at small concentrations and a longer exposure time at 37 °C. The exploration of CEO and MEO employment for the inhibition of Salmonella bacteria at different temperatures and conditions expands the possibilities of developing more environment- and consumer-friendly antibacterial protection for raw meat

Different Metabolic Pathways Are Involved in Response of Saccharomyces cerevisiae to L-A and M Viruses

Competitive and naturally occurring yeast killer phenotype is governed by coinfection with dsRNA viruses. Long-term relationship between the host cell and viruses appear to be beneficial and co-adaptive; however, the impact of viral dsRNA on the host gene expression has barely been investigated. Here, we determined the transcriptomic profiles of the host Saccharomyces cerevisiae upon the loss of the M-2 dsRNA alone and the M-2 along with the L-A-lus dsRNAs. We provide a comprehensive study based on the high-throughput RNA-Seq data, Gene Ontology and the analysis of the interaction networks. We identified 486 genes differentially expressed after curing yeast cells of the M-2 dsRNA and 715 genes affected by the elimination of both M-2 and L-A-lus dsRNAs. We report that most of the transcriptional responses induced by viral dsRNAs are moderate. Differently expressed genes are related to ribosome biogenesis, mitochondrial functions, stress response, biosynthesis of lipids and amino acids. Our study also provided insight into the virus–host and virus–virus interplays