The modifying enzyme Tsr3 establishes the hierarchy of Rio kinase activity in 40S ribosome assembly

Ribosome assembly is an intricate process, which in eukaryotes is promoted by a large machinery comprised of over 200 assembly factors (AF) that enable the modification, folding, and processing of the ribosomal RNA (rRNA) and the binding of the 79 ribosomal proteins. While some early assembly steps occur via parallel pathways, the process overall is highly hierarchical, which allows for the integration of maturation steps with quality control processes that ensure only fully and correctly assembled subunits are released into the translating pool. How exactly this hierarchy is established, in particular given that there are many instances of RNA substrate “handover” from one highly related AF to another remains to be determined. Here we have investigated the role of Tsr3, which installs a universally conserved modification in the P-site of the small ribosomal subunit late in assembly. Our data demonstrate that Tsr3 separates the activities of the Rio kinases, Rio2 and Rio1, with whom it shares a binding site. By binding after Rio2 dissociation, Tsr3 prevents rebinding of Rio2, promoting forward assembly. After rRNA modification is complete, Tsr3 dissociates, thereby allowing for recruitment of Rio1. Inactive Tsr3 blocks Rio1, which can be rescued using mutants that bypass the requirement for Rio1 activity. Finally, yeast strains lacking Tsr3 randomize the binding of the two kinases, leading to the release of immature ribosomes into the translating pool. These data demonstrate a role for Tsr3 and its modification activity in establishing a hierarchy for the function of the Rio kinases.

Saccharomyces cerevisiae Cells Lacking the Zinc Vacuolar Transporter Zrt3 Display Improved Ethanol Productivity in Lignocellulosic Hydrolysates

Yeast-based bioethanol production from lignocellulosic hydrolysates (LH) is an attractive and sustainable alternative for biofuel production. However, the presence of acetic acid (AA) in LH is still a major problem. Indeed, above certain concentrations, AA inhibits yeast fermentation and triggers a regulated cell death (RCD) process mediated by the mitochondria and vacuole. Understanding the mechanisms involved in AA-induced RCD (AA-RCD) may thus help select robust fermentative yeast strains, providing novel insights to improve lignocellulosic ethanol (LE) production. Herein, we hypothesized that zinc vacuolar transporters are involved in vacuole-mediated AA-RCD, since zinc enhances ethanol production and zinc-dependent catalase and superoxide dismutase protect from AA-RCD. In this work, zinc limitation sensitized wild-type cells to AA-RCD, while zinc supplementation resulted in a small protective effect. Cells lacking the vacuolar zinc transporter Zrt3 were highly resistant to AA-RCD, exhibiting reduced vacuolar dysfunction. Moreover, zrt3Δ cells displayed higher ethanol productivity than their wild-type counterparts, both when cultivated in rich medium with AA (0.29 g L−1 h−1 versus 0.11 g L−1 h−1) and in an LH (0.73 g L−1 h−1 versus 0.55 g L−1 h−1). Overall, the deletion of ZRT3 emerges as a promising strategy to increase strain robustness in LE industrial production.

Whole-Genome Sequencing and Annotation of the Yeast Clavispora santaluciae Reveals Important Insights about Its Adaptation to the Vineyard Environment

Clavispora santaluciae was recently described as a novel non-Saccharomyces yeast species, isolated from grapes of Azores vineyards, a Portuguese archipelago with particular environmental conditions, and from Italian grapes infected with Drosophila suzukii. In the present work, the genome of five Clavispora santaluciae strains was sequenced, assembled, and annotated for the first time, using robust pipelines, and a combination of both long- and short-read sequencing platforms. Genome comparisons revealed specific differences between strains of Clavispora santaluciae reflecting their isolation in two separate ecological niches—Azorean and Italian vineyards—as well as mechanisms of adaptation to the intricate and arduous environmental features of the geographical location from which they were isolated. In particular, relevant differences were detected in the number of coding genes (shared and unique) and transposable elements, the amount and diversity of non-coding RNAs, and the enzymatic potential of each strain through the analysis of their CAZyome. A comparative study was also conducted between the Clavispora santaluciae genome and those of the remaining species of the Metschnikowiaceae family. Our phylogenetic and genomic analysis, comprising 126 yeast strains (alignment of 2362 common proteins) allowed the establishment of a robust phylogram of Metschnikowiaceae and detailed incongruencies to be clarified in the future.

The Influencing Factors on the Production of Alcoholic Drinking from Jackfruit (Artocarpus heterophyllus L.)

The large amount of jackfruit (Artocarpus heterophyllus Lam) harvested and their short use time caused many difficulties for the farmers. Fortunately, the high sugar content in jackfruit meat is a hopeful substance for wine production. This study aimed to consider the effect of yeast strains and their concentration on fermented jackfruit solution. Jackfruit juice with 14 °Brix is ​​fermented using 0.005 to 0.015% (w/v) Saccharomyces cerevisiae RV002, Mauri Instant Dry Yeast yeast under anaerobic conditions for 1 to 4 days at 30 °C. Survey samples were checked once a day to analyze the indicators. The functional report of the sugar in the fermentation time, shows that the higher incidence of yeast cultures and the initial sugar concentration inhibited yeast growth. The results showed that fermentation from jackfruit meat with 25 °Brix using Saccharomyces cerevisiae RV002 yeast with concentration of 0.01% for 3 days is the best to create a good quality with ethanol content 4,9% and characteristic aroma of jackfruit.

Evaluation of two brazilian native yeast strains (Pichia kudriavzevii) in craft beer

Beer production is an ancient biotechnological process and since yeast as discovered to be responsible for the transformation of barley wort into beer, studies have been carried out with the aim of understanding the behavior of these microorganisms. This work aimed to study the application of two strains of yeasts of the species Pichia kudriavzevii, isolated in the Brazilian Midwest for the production of craft beer and to analyze the occurrence of bioaromatization, with the production of volatile organic compounds (VOC) and to evaluate the sensory perception of the results with untrained end consumers through a quick sensory methodology called Check All That Apply (CATA). For this purpose, three batches of beer were produced and inoculated with commercial yeast (Saccharomyces cerevisiae, the control) and two strains of the same species, called Pichia kudriavzevii BB1 and Pichia kudriavzevii BB2. A total of 28 volatile organic compounds that differentiated the control of the Pichia BB1/BB2 group were detected, being 20 esters, 2 alcohols, 5 carboxylic acids and 1 hydrocarbon. There was no difference among the Pichia kudriavzevii BB1 and BB2 samples (p>0.05) in the sensory analysis using the CATA methodology. It was possible to distinguish two clusters between the tasters according to the habit of consuming craft beer, and those who consumed frequently, assigned a higher score in the hedonic test. It was concluded that Pichia kudriavzevii BB1 and BB2 influenced the beer bioaromatization, improving the acceptance test score with the tasters.