The Zn(II)2Cys6-Type Transcription Factor ADA-6 Regulates Conidiation, Sexual Development, and Oxidative Stress Response in Neurospora crassa

Abstract

Conidiation and sexual development are critical for reproduction, dispersal and better-adapted survival in many filamentous fungi. The <i>Neurospora crassa</i> gene <i>ada-6</i> encodes a Zn(II)2Cys6-type transcription factor, whose deletion resulted in reduced conidial production and female sterility. In this study, we confirmed the positive contribution of <i>ada-6</i> to conidiation and sexual development by detailed phenotypic characterization of its deletion mutant and the complemented mutant. To understand the regulatory mechanisms of ADA-6 in conidiation and sexual development, transcriptomic profiles generated by RNA-seq from the Δ<i>ada-6</i> mutant and wild type during conidiation and sexual development were compared. During conidial development, differential expressed genes (DEGs) between the Δ<i>ada-6</i> mutant and wild type are mainly involved in oxidation-reduction process and single-organism metabolic process. Several conidiation related genes are positively regulated by ADA-6, including genes that positively regulate conidiation (<i>fluffy</i> and <i>acon-3</i>), and genes preferentially expressed during conidial development (<i>eas</i>, <i>con-6</i>, <i>con-8</i>, <i>con-10</i>, <i>con-13</i>, <i>pcp-1</i>, and NCU9357), as the expression of these genes were lower in the Δ<i>ada-6</i> mutant compared to wild type during conidial development. Phenotypic observation of deletion mutants for other genes with unknown function down-regulated by <i>ada-6</i> deletion revealed that deletion mutants for four genes (NCU00929, NCU05260, NCU00116, and NCU04813) produced less conidia than wild type. Deletion of <i>ada-6</i> resulted in female sterility, which might be due to that ADA-6 affects oxidation-reduction process and transmembrane transport process, and positively regulates the transcription of <i>pre-2</i>, <i>poi-2</i>, and NCU05832, three key genes participating in sexual development. In both conidiation and the sexual development process, ADA-6 regulates the transcription of <i>cat-3</i> and other genes participating in reactive oxygen species production according to RNA-seq data, indicating a role of ADA-6 in oxidative stress response. This was further confirmed by the results that deletion of <i>ada-6</i> led to hypersensitivity to oxidants H₂O₂ and menadione. Together, these results proved that ADA-6, as a global regulator, plays a crucial role in conidiation, sexual development, and oxidative stress response of <i>N. crassa</i>.