Effect of Inhibition of the bc1 Complex on Gene Expression Profile in Yeast

Abstract

Because the respiratory chain is the major site of oxidation of the reduced equivalents and of energy production in aerobic cells, its inhibition has severe impact on the cells. Communication pathways from the respiratory chain are required to allow the cell to sense the defect and respond to it. In this work, we studied changes in gene expression induced by the treatment of yeast cells with myxothiazol, an inhibitor of the bc 1 complex, an enzyme of the respiratory chain. The pattern and time-course expression of the genes resemble those of the environmental stress response, a common gene expression program induced by sudden changes in the environment. In addition, the changes were, for most of the genes, mediated through the transcription factors Msn2/4, which play a central role in the cellular response to these stresses. By using a mutant with a myxothiazol-resistant bc 1 complex, we showed that the changes of expression of the majority of the genes was caused by the inhibition of the bc 1 complex but that other stresses might be involved. The expression pattern of CTT1, coding for a cytoplasmic catalase, was further studied. The expression of this gene was largely dependent on Msn2/4 and the inhibition of the cytochrome bc 1 . Addition of oxidants of NADH was found to decrease the expression of CTT1 induced by myxothiazol treatment, suggesting that the accumulation of NADH caused by the inhibition of the respiratory chain may be involved in the signaling pathway from the mitochondria to the transcription factor.The mitochondrial respiratory chain is the main site of oxidation of reducing equivalents and of energy production in aerobic cells, both of which are essential for metabolic pathways. Inhibition of the respiratory function is thus expected to have a severe impact on the cell and to cause re-adjustment of cellular metabolism. Herein, we report on the effect of respiratory chain dysfunction on the pattern of gene expression. We chose to study the inhibition of the respiratory chain at the level of the bc 1 complex, because several mutations in this complex have been reported in human diseases and, most interestingly, this enzyme is the target of fungicides or anti-parasite drugs used in agriculture and medicine. Therefore it seemed interesting to understand the cellular events occurring upon treatment.The mitochondrial bc 1 complex is a membrane-bound enzyme that catalyzes the transfer of electrons from ubiquinol to cytochrome c and couples this electron transfer to the vectorial translocation of protons across the inner mitochondrial membrane. The catalytic mechanism, called the Q-cycle, requires two distinct quinone-binding sites (Q o , quinol oxidation site, and Q i , quinone reduction site), which are located on opposite sides of the membrane and linked by a transmembrane electron-transfer pathway. The mitochondrially encoded cytochrome b subunit provides both the Q o and Q i pockets and the transmembrane electron pathway (via the hemes b). Q o -specific inhibitors (Q o

Metabolic-State-Dependent Remodeling of the Transcriptome in Response to Anoxia and Subsequent Reoxygenation in Saccharomyces cerevisiae

“…As indicated in Fig. 5, many genes in C g 13 and C g 14 are not differentially expressed in response to the O 2 shifts in glucose medium, suggesting that they respond to the abrupt cessation of respiration rather oxygen deprivation per se (10,61). In support of this, many of these genes are apparently involved in increasing cellular energy currency during the metabolic switch.…”

“…Those genes involved in mitochondrial functions that were transiently induced in C g 7 to C g 9 are not part of the environmental stress response and appear to respond directly to the inhibition of respiration. Although the initial event that triggers these changes is the abrupt decline in oxygen availability, the signal that is responsible for eliciting changes in gene network activity appears to be metabolic in origin and linked to the cessation of respiration, given that a similar response is evoked under normoxic conditions with treatment with either antimycin A (Lai et al, unpublished) or myxothiazol (10). Moreover, a similar response is not invoked in catabolite-repressed cells in which the respiratory capacity is repressed, and no change in growth rate is observed during the transition from aerobic to anaerobic conditions.…”

“…These results suggest that the "stress" encountered is the abrupt cessation of respiration and the associated energetic changes, not the withdrawal of oxygen per se. Indeed, studies in our laboratory (L.-C. Lai, M. T. Kissinger, and K. E. Kwast, unpublished data) and by others (10) have revealed that simply inhibiting the respiratory chain under aerobiosis produces a transcriptional response similar to that elicited by anaerobiosis when cells are grown in galactose.…”

Multiple Defects in the Respiratory Chain Lead to the Repression of Genes Encoding Components of the Respiratory Chain and TCA Cycle Enzymes

“…The protocol for preparation of the sample and the microarray analysis has been described in Ref. 37. These experiments were carried out using a complete Affymetrix GeneChip system yeast Genome S98 arrays.…”

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“…Notable among these are the global analysis of protein localization in yeast using tagged open reading frames [7,8], the proteomic analysis of purified mitochondria and mitochondrial substructures using mass spectroscopy-based methods [9-13], systematic analysis of the collections of yeast gene knock-outs for mitochondrial related phenotypes [14,15], and gene-expression profiling in conditions that require mitochondrial function or when mitochondrial oxidative phosphorylation is disrupted [16-23]. Several of these studies provided critical datasets used by Perocchi et al .…”