Pobre, V. | Search bioRxiv

bioRxiv2023

RNase R Controls Membrane Fatty Acid Composition in Streptococcus pneumoniae

AO_SCPLOWBSTRACTC_SCPLOWPrevious studies on RNase R have highlighted significant effects of this ribonuclease in several processes of Streptococcus pneumoniae biology. In this work we have studied the global impact of RNase R by comparing the transcriptional landscape of a deleted RNase R mutant to that of the wild-type strain, and this led us investigate specific targets affected by RNase R. RNA-Seq showed that RNase R deletion affects transcripts from several different biological processes. Of particular interest, elimination of RNase R results in overexpression of most of the genes encoding the components of type II fatty acid biosynthesis (FAS-II) cluster. We demonstrate that RNase R governs the turnover of most of genes from this pathway, affecting the outcome of the whole FAS-II cluster, and leading to an unbalanced membrane fatty acid composition. Our results show that the membrane of the deleted strain contains a higher proportion of unsaturated and long-chained fatty acids than the wild type strain. This leads to a higher fluidity of the {Delta}rnr mutant membrane, which is probably related with the increased sensitivity to detergent observed in this strain. We demonstrate that RNase R expression is induced in cells challenged with H2O2, which is suggestive of a role for this ribonuclease on the regulation of membrane homeostasis under oxidative stress. Reprogramming of membrane fluidity is an adaptative cell response crucial for bacterial survival in constantly changing environmental conditions. The fact that RNase R controls the expression of several essential genes to the fatty acid synthesis unveils a new important function of this enzyme.

molecular biology↗

bioRxiv2021

Copper Acts Synergistically with Fluconazole in Candida glabrata by Compromising Drug Efflux, Sterol Metabolism, and Zinc Homeostasis

The synergistic combinations of drugs are promising strategies to boost the effectiveness of current antifungals and thus prevent the emergence of resistance. In this work, we show that copper and the antifungal fluconazole act synergistically against Candida glabrata, an opportunistic pathogenic yeast intrinsically tolerant to fluconazole. Analyses of the transcriptomic profile of C. glabrata after the combination of copper and fluconazole showed that the expression of the multidrug transporter gene CDR1 was decreased, suggesting that fluconazole efflux could be affected. In agreement, we observed that copper inhibits the transactivation of Pdr1, the transcription regulator of multidrug transporters and leads to the intracellular accumulation of fluconazole. Copper also decreases the transcriptional induction of ergosterol biosynthesis (ERG) genes by fluconazole, which culminates in the accumulation of toxic sterols. Co-treatment of cells with copper and fluconazole should affect the function of proteins located in the plasma membrane, as several ultrastructural alterations, including irregular cell wall and plasma membrane and loss of cell wall integrity, were observed. Finally, we show that the combination of copper and fluconazole downregulates the expression of the gene encoding the zinc-responsive transcription regulator Zap1, which possibly, together with the membrane transporters malfunction, generates zinc depletion. Supplementation with zinc reverts the toxic effect of combining copper with fluconazole, underscoring the importance of this metal in the observed synergistic effect. Overall, this work, while unveiling the molecular basis that supports the use of copper to enhance the effectiveness of fluconazole, paves the way for the development of new metal-based antifungal strategies.

microbiology↗