Transcriptome

Biological duplicate cultures from the control strain and L-UBB⁺¹ strain were sampled during EX and D6 for microarray analysis. Cells were frozen immediately in liquid nitrogen for rapid quenching of mRNA turnover [86]. Cells were mechanically disrupted using a FastPrep homogenizer (MP Biomedicals, USA) and total RNA was extracted using the RNeasy Mini Kit (QIAGEN, Germany). Quality of total RNA was assessed using an RNA 6000 Nano LabChip Kit (Agilent Technologies, USA) with an Agilent 2100 Bioanalyzer (Agilent Technologies, USA). The labeled RNA was generated using the GeneChip^® 3′ IVT Plus Reagent Kit (Affymetrix, USA), which was hybridized to GeneChip^® Yeast Genome 2.0 Arrays (Affymetrix, USA). Staining and washing of the hybridized arrays were performed on the GeneChip^® Fluidics Station 450 (Affymetrix, USA). Further microarrays were scanned in GeneChip^® Scanner 7G (Affymetrix, USA). RNA labelling, array hybridization and scanning were performed by the Bioinformatics and Expression Analysis core facility at Karolinska Institute, Sweden. Microarray data are available at the Genome Expression Omnibus website (GEO, http://www.ncbi.nlm.nih.gov/geo/) with the accession numbers {"type":"entrez-geo","attrs":{"text":"GSE129688","term_id":"129688"}}GSE129688. The transcriptome data (CEL files) were analyzed using the R version 3.4.0 and the PIANO package (Platform for Integrative Analysis of Omics) with information from the Saccharomyces Genome Database (https://www.yeastgenome.org/) [87]. Gene set enrichment analysis (GSA) was performed to identify overrepresentation of functional annotation categories using the Database for Annotation, Visualization and Integrated Discovery (David, https://david.ncifcrf.gov/). The S288C yeast genome background was used to analyze the magnitude of fold enrichment. The differential gene expression (log₂-FC) and corresponding significance (adjusted p-value) were calculated by the Benjamini–Hochberg method. Heatmaps of significantly differentially expressed genes and gene sets were generated by pheatmap R package.