RESUMO
In the ciliated protozoan Paramecium tetraurelia, Piwi-associated small RNAs are generated upon the elimination of tens of thousands of short transposon-derived DNA segments as part of development. These RNAs then target complementary DNA for elimination in a positive feedback process, contributing to germline defense and genome stability. In this work, we investigate the formation of these RNAs, which we show to be transcribed directly from the short (length mode 27 bp) excised DNA segments. Our data support a mechanism whereby the concatenation and circularization of excised DNA segments provides a template for RNA production. This process allows the generation of a double-stranded RNA for Dicer-like protein cleavage to give rise to a population of small regulatory RNAs that precisely match the excised DNA sequences. VIDEO ABSTRACT.
Assuntos
DNA Concatenado , Paramecium tetraurellia/genética , Núcleo Celular/metabolismo , DNA Ligase Dependente de ATP/metabolismo , Elementos de DNA Transponíveis , Exodesoxirribonucleases/metabolismo , Paramecium tetraurellia/citologia , Paramecium tetraurellia/metabolismo , RNA/genética , Transcrição GênicaRESUMO
Distillery yeasts are poorly characterized physiological group among the Saccharomyces sensu stricto complex. As industrial yeasts are under constant environmental stress during fermentation processes and the nucleolus is a stress sensor, in the present study, nucleolus-related parameters were evaluated in 22 commercially available distillery yeast strains. Distillery yeasts were found to be a heterogeneous group with a variable content and length of rDNA and degree of nucleolus fragmentation. The levels of rDNA were negatively correlated with Nop1 (r = -0.59, p = 0.0038). Moreover, the protein levels of Sir transcriptional silencing complex and longevity regulators, namely Sir1, Sir2, Sir3 and Fob1, were studied and negative correlations between Sir2 and Nop1 (r = -0.45, p = 0.0332), and between Sir2 and Fob1 (r = -0.49, p = 0.0211) were revealed. In general, S. paradoxus group of distillery yeasts with higher rDNA pools and Sir2 level than S. bayanus group was found to be more tolerant to fermentation-associated stress stimuli, namely mild cold/heat stresses and KCl treatment. We postulate that rDNA state may be considered as a novel factor that may modulate a biotechnological process.
Assuntos
Bebidas Alcoólicas/microbiologia , DNA Ribossômico/genética , Proteínas Nucleares/genética , Ribonucleoproteínas Nucleolares Pequenas/genética , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Proteínas Reguladoras de Informação Silenciosa de Saccharomyces cerevisiae/genética , Sirtuína 2/genética , Proteínas de Ligação a DNA/genética , Região Organizadora do Nucléolo/genéticaRESUMO
Industrial yeast strains of economic importance used in winemaking and beer production are genomically diverse and subjected to harsh environmental conditions during fermentation. In the present study, we investigated wine yeast adaptation to chronic mild alcohol stress when cells were cultured for 100 generations in the presence of non-cytotoxic ethanol concentration. Ethanol-induced reactive oxygen species (ROS) and superoxide signals promoted growth rate during passages that was accompanied by increased expression of sirtuin proteins, Sir1, Sir2 and Sir3, and DNA-binding transcription regulator Rap1. Genome-wide array-CGH analysis revealed that yeast genome was shaped during passages. The gains of chromosomes I, III and VI and significant changes in the gene copy number in nine functional gene categories involved in metabolic processes and stress responses were observed. Ethanol-mediated gains of YRF1 and CUP1 genes were the most accented. Ethanol also induced nucleolus fragmentation that confirms that nucleolus is a stress sensor in yeasts. Taken together, we postulate that wine yeasts of different origin may adapt to mild alcohol stress by shifts in intracellular redox state promoting growth capacity, upregulation of key regulators of longevity, namely sirtuins and changes in the dosage of genes involved in the telomere maintenance and ion detoxification.
Assuntos
Adaptação Biológica/efeitos dos fármacos , Cromossomos Fúngicos/genética , Etanol/farmacologia , Fermentação/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Saccharomyces cerevisiae/fisiologia , Cerveja , Nucléolo Celular/efeitos dos fármacos , Cromossomos Fúngicos/efeitos dos fármacos , Hibridização Genômica Comparativa , Indústria Alimentícia , Dosagem de Genes , Oxirredução , Proteínas de Saccharomyces cerevisiae/metabolismo , Complexo Shelterina , Transdução de Sinais/efeitos dos fármacos , Proteínas Reguladoras de Informação Silenciosa de Saccharomyces cerevisiae/metabolismo , Sirtuína 2/metabolismo , Sirtuínas , Homeostase do Telômero/efeitos dos fármacos , Homeostase do Telômero/genética , Proteínas de Ligação a Telômeros/metabolismo , Fatores de Transcrição/metabolismo , VinhoRESUMO
The nucleolus is considered to be a stress sensor and rDNA-based regulation of cellular senescence and longevity has been proposed. However, the role of rDNA in the maintenance of genome integrity has not been investigated in detail. Using genomically diverse industrial yeasts as a model and array-based comparative genomic hybridization (aCGH), we show that chromosome level may be balanced during passages and as a response to alcohol stress that may be associated with changes in rDNA pools. Generation- and ethanol-mediated changes in genes responsible for protein and DNA/RNA metabolism were revealed using next-generation sequencing. Links between redox homeostasis, DNA stability, and telomere and nucleolus states were also established. These results suggest that yeast genome is dynamic and chromosome homeostasis may be controlled by rDNA.
