Comparative Genomic and Transcriptomic Analysis Reveals Specific Features of Gene Regulation in Kluyveromyces marxianus

Abstract

Kluyveromyces marxianus is a promising host for producing bioethanol and heterologous proteins. It displays many superior traits to a conventional industrial yeast species, Saccharomyces cerevisiae, including fast growth, thermotolerance and the capacity to assimilate a wider variety of sugars. However, little is known about the mechanisms underlying the fast-growing feature of K. marxianus. In this study, we performed a comparative genomic analysis between K. marxianus and other Saccharomycetaceae species. Genes involved in flocculation, iron transport, and biotin biosynthesis have particularly high copies in K. marxianus. In addition, 60 K. marxianus specific genes were identified, 45% of which were upregulated during cultivation in rich medium and these genes may participate in glucose transport and mitochondrion related functions. Furthermore, the transcriptomic analysis revealed that under aerobic condition, normalized levels of genes participating in TCA cycles, respiration chain and ATP biosynthesis in the lag phase were higher in K. marxianus than those in S. cerevisiae. Levels of highly copied genes, genes involved in the respiratory chain and mitochondrion assembly, were upregulated in K. marxianus, but not in S. cerevisiae, in later time points during cultivation compared with those in the lag phase. Notably, during the fast-growing phase, genes involved in the respiratory chain, ATP synthesis and glucose transport were co-upregulated in K. marxianus. A few shared motifs in upstream sequences of relevant genes might result in the co-upregulation. Specific features in the co-regulations of gene expressions might contribute to the fast-growing phenotype of K. marxianus. Our study underscores the importance of genome-wide rewiring of the transcriptional network during evolution.

Characterization of essential elements for improved episomal expressions in Kluyveromyces marxianus

“…The transcription level of INU1 at different time points during culturing (0, 4, 6, 12, 24, 48, or 72 h) was 17~70 times that of OM45 in K. marxianus. [33] However, in a dual-reporter plasmid, the relative strength of a 1000-bp OM45 promoter was 1.3 times that of the 1136-bp INU1 promoter (Figure 2B).…”

“…RNA-seq data was extracted from a previous study. [33] In brief, wildtype FIM1 was cultured in the 50 mL YPD liquid medium (2% glucose, 2% polypeptone, 1% yeast extract) overnight and then transferred into 50 mL fresh YPD liquid medium with an optical density at 600 nm (OD 600 ) of 0. value by the average FPKM value of three housekeeping genes, including MPE1, TRK1, and SWC4. [33] In the wild-type FIM1 cells cultured in rich YPD medium, the stably expressed genes were defined as the coefficient of variations (standard deviation divided by its average FPKM value) <0.7 at 12, 24, and 72 h time points.…”

Characterization and optimization of the LAC4 upstream region for low‐leakage expression in Kluyveromyces marxianus

“…locating on the left end of Chromosome 3 encodes a full-length β-galactosidase and was systematically designated as FIM1_1709, while the LAC4 on the right end encodes a truncated β-galactosidase and was designated as FIM1_2437 (Yu et al, 2021). As shown in 1/11 of that in lactose medium and was 1/8 of that in galactose medium (Figure 1b).…”

Characterization and modulation of endoplasmic reticulum stress response target genes in Kluyveromyces marxianus to improve secretory expressions of heterologous proteins

“…FIM-1ΔU strain harbors 5202 genes in total [ 22 ]. After a 15-min treatment of DTT, 60 genes were significantly upregulated (Fold change > 2, q value < 0.1), among which genes involved in the glycolytic process were enriched (Fig.…”