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Response to oxidative stress is universal in almost all organisms and the mitochondrial membrane protein, BbOhmm, negatively affects oxidative stress responses and virulence in the insect fungal pathogen,Beauveria bassiana. Nothing further, however, is known concerning howBbOhmmand this phenomenon is regulated.

Threeoxidativestressresponse regulating Zn₂Cys₆transcription factors (BbOsrR1, 2, and 3) were identified and verified via chromatin immunoprecipitation (ChIP)-qPCR analysis as binding to theBbOhmmpromoter region, with BbOsrR2 showing the strongest binding. Targeted gene knockout ofBbOsrR1orBbOsrR3led to decreasedBbOhmmexpression and consequently increased tolerances to free radical generating compounds (H₂O₂and menadione), whereas the ΔBbOsrR2strain showed increasedBbOhmmexpression with concomitant decreased tolerances to these compounds. RNA and ChIP sequencing analysis revealed that BbOsrR1 directly regulated a wide range of antioxidation and transcription-associated genes, negatively affecting the expression of theBbClp1cyclin andBbOsrR2. BbClp1 was shown to localize to the cell nucleus and negatively mediate oxidative stress responses. BbOsrR2 and BbOsrR3 were shown to feed into the Fus3-MAPK pathway in addition to regulating antioxidation and detoxification genes. Binding motifs for the three transcription factors were found to partially overlap in the promoter region ofBbOhmmand other target genes. Whereas BbOsrR1 appeared to function independently, co-immunoprecipitation revealed complex formation between BbClp1, BbOsrR2, and BbOsrR3, with BbClp1 partially regulating BbOsrR2 phosphorylation.

These findings reveal a regulatory network mediated by BbOsrR1 and the formation of a BbClp1-BbOsrR2-BbOsrR3 complex that orchestrates fungal oxidative stress responses.