Molecular mechanism of enhanced ethanol tolerance associated with hfq overexpression in Zymomonas mobilis

Abstract

Zymomonas mobilis is a promising microorganism for industrial bioethanol production. However, ethanol produced during fermentation is toxic to Z. mobilis and affects its growth and bioethanol production. Although several reports demonstrated that the RNA-binding protein Hfq in Z. mobilis contributes to the tolerance against multiple lignocellulosic hydrolysate inhibitors, the role of Hfq on ethanol tolerance has not been investigated. In this study, hfq in Z. mobilis was either deleted or overexpressed and their effects on cell growth and ethanol tolerance were examined. Our results demonstrated that hfq overexpression improved ethanol tolerance of Z. mobilis, which is probably due to energy saving by downregulating flagellar biosynthesis and heat stress response proteins, as well as reducing the reactive oxygen species induced by ethanol stress via upregulating the sulfate assimilation and cysteine biosynthesis. To explore proteins potentially interacted with Hfq, the TEV protease mediated Yeast Endoplasmic Reticulum Sequestration Screening system (YESS) was established in Z. mobilis. YESS results suggested that Hfq may modulate the cytoplasmic heat shock response by interacting with the heat shock proteins DnaK and DnaJ to deal with the ethanol inhibition. This study thus not only revealed the underlying mechanism of enhanced ethanol tolerance by hfq overexpression, but also provided an alternative approach to investigate protein-protein interactions in Z. mobilis.

Analysis of heterologous expression of phaCBA promotes the acetoin stress response mechanism in Bacillus subtilis using transcriptomics and metabolomics approaches

“…Similarly, reduced chemotaxis activity helps the cell adapt to its environment and allocate resources more wisely under stressful conditions. This aligns with previous studies that have shown down-regulation of flagellar biosynthesis in response to stress, suggesting a conserved strategy among bacteria [ 31 , 32 ]. Up-regulation of transporter proteins: Several transporter protein genes, such as molybdate ( modA , modB ), osmoprotectant ( opuBC , opuBB , opuBA ), biotin ( bioY ), and zinc/manganese/iron (II), were significantly up-regulated in the ABC transporters system (Additional file 3 : Table S4).…”

“…This aligns with previous studies that have shown down-regulation of flagellar Fig. 7 Batch fermentation of recombinant strain BS03-phaCBA in a 7.5 L bioreactor for the production of acetoin biosynthesis in response to stress, suggesting a conserved strategy among bacteria [31,32]. Up-regulation of transporter proteins: Several transporter protein genes, such as molybdate (modA, modB), osmoprotectant (opuBC, opuBB, opuBA), biotin (bioY), and zinc/manganese/iron (II), were significantly up-regulated in the ABC transporters system (Additional file 3: Table S4).…”

In vitro characterization of a pAgo nuclease TtdAgo from Thermococcus thioreducens and evaluation of its effect in vivo

“…All plasmids were assembled by the Gibson Assembly method (Gibson et al, 2009). After purification, gene and vector fragments were ligated through the T5 exonuclease (NEB, WA, United States)dependent DNA assembly method as described previously (Tang et al, 2022). After transformation in E. coli, the correct colonies were selected by colony PCR and confirmed by Sanger sequencing (Tsingke Biotechnology, Beijing, China).…”

Comprehensive network of stress-induced responses in Zymomonas mobilis during bioethanol production: from physiological and molecular responses to the effects of system metabolic engineering

“…Hfq is an RNA-binding protein that regulates gene expression during post-transcriptional levels. Hfq plays various functions in the RNA metabolism of bacteria, including stabilizing small non-coding RNAs (sRNAs) and aiding in their interactions with messenger RNAs (mRNAs), which consequently impact the stability and translation of the specific targeted mRNAs [ 90 ]. This protein is homologous to Lms proteins found in eukaryotic cells in both functions and structures.…”

“…Moreover, overexpression of hfq resulted in tolerance of Z. mobilis to ethanol stress, which was attributed to the downregulation of genes related to flagellar biosynthesis and heat stress response proteins. Additionally, there was an upregulation of genes related to sulfate assimilation and cysteine biosynthesis, leading to a reduction in reactive oxygen species induced during ethanol stress [ 90 ].…”