Alternative Oxidase Transcription Factors AOD2 and AOD5 ofNeurospora crassaControl the Expression of Genes Involved in Energy Production and Metabolism

Abstract

In Neurospora crassa, blocking the function of the standard mitochondrial electron transport chain results in the induction of an alternative oxidase (AOX). AOX transfers electrons directly from ubiquinol to molecular oxygen. AOX serves as a model of retrograde regulation since it is encoded by a nuclear gene that is regulated in response to signals from mitochondria. The N. crassa transcription factors AOD2 and AOD5 are necessary for the expression of the AOX gene. To gain insight into the mechanism by which these factors function, and to determine if they have roles in the expression of additional genes in N. crassa, we constructed strains expressing only tagged versions of the proteins. Cell fractionation experiments showed that both proteins are localized to the nucleus under both AOX inducing and noninducing conditions. Furthermore, chromatin immunoprecipitation and high throughput sequencing (ChIP-seq) analysis revealed that the proteins are bound to the promoter region of the AOX gene under both conditions. ChIP-seq also showed that the transcription factors bind to the upstream regions of a number of genes that are involved in energy production and metabolism. Dependence on AOD2 and AOD5 for the expression of several of these genes was verified by quantitative PCR. The majority of ChIP-seq peaks observed were enriched for both AOD2 and AOD5. However, we also observed occasional sites where one factor appeared to bind preferentially. The most striking of these was a conserved sequence that bound large amounts of AOD2 but little AOD5. This sequence was found within a 310 bp repeat unit that occurs at several locations in the genome.

Characterization of Single Gene Deletion Mutants Affecting Alternative Oxidase Production in Neurospora crassa: Role of the yvh1 Gene

“…The production of the AOD1 protein in N. crassa is known to be controlled at the level of transcription of the aod-1 gene [ 19 , 30 , 31 , 33 , 37 ] and at the level of translation of the transcript [ 19 , 37 ]. To distinguish between the two possible mechanisms preventing expression of the protein in Δ yvh1 cells, we analyzed the levels of aod-1 transcript in the deletion mutant.…”

“…A number of genes that affect aod-1 expression have been identified [ 23 , 28 , 29 ]. Two zinc cluster transcription factors, encoded by the aod-2 and aod-5 genes [ 23 , 30 ], bind constitutively as a heterodimer in the upstream region of the aod-1 gene and are required for the increased transcription of aod-1 that occurs under inducing conditions [ 19 , 30 , 31 , 32 , 33 ]. Similarly, the expression of AOX in Aspergillus nidulans and Podospora anserina requires the orthologues of N. crassa AOD2 and AOD5 [ 34 , 35 , 36 ].…”

“…Similarly, the expression of AOX in Aspergillus nidulans and Podospora anserina requires the orthologues of N. crassa AOD2 and AOD5 [ 34 , 35 , 36 ]. In all three species, these transcription factors play additional roles in the expression of certain genes involved in gluconeogenesis and other metabolic processes [ 33 , 35 , 36 ].…”

Involvement of a G Protein Regulatory Circuit in Alternative Oxidase Production inNeurospora crassa

“…Gene rescue experiments, sequencing, and bioinformatics revealed that aod-2 and aod-5 encode zinc cluster transcription factors (Chae et al 2007b). These factors bind constitutively as a heterodimer to an alternative oxidase induction motif (AIM) that exists in the promoter region of the aod-1 gene (Chae et al 2007a; Qi et al 2017). Transcription of aod-1 is induced in response to an unknown signal(s) generated when the standard electron transport chain is disrupted.…”

“…Transcription of aod-1 is induced in response to an unknown signal(s) generated when the standard electron transport chain is disrupted. In the absence of either aod-2 or aod-5 , the induction of aod-1 is essentially eliminated (Tanton et al 2003; Chae et al 2007a; Chae et al 2007b; Chae and Nargang 2009; Qi et al 2017). Orthologs of the AOD2 and AOD5 proteins exist in other fungi and have been shown to be required for AOX induction in Aspergillus nidulans (Suzuki et al 2012) and Podospora anserina (Bovier et al 2014).…”