biostudies-literatureYu PMOST | National Natural Science Foundation of China (NSFC)| Beijing Municipal Natural Science Foundation ()e0070324https://www.ebi.ac.uk/biostudies/studies/S-EPMC11774033biostudies-literatureUnknown10(1)The widespread use of azole antifungals in agriculture and clinical settings has led to serious drug resistance. Overexpression of the azole drug target 14α-demethylase ERG11 (CYP51) is the most common fungal resistance mechanism. However, the presence of additional regulatory proteins in the transcriptional response of <i>erg11</i> is not yet fully elucidated. In this study, leveraging the identified key promoter region of <i>erg11</i> that controls its response to azoles in <i>Neurospora crassa</i>, we pinpointed a protein, Crf4-3, which harbors a PWWP domain and exerts a positive regulatory influence on azole resistance, as determined by DNA pulldown assays. The removal of Crf4-3 results in heightened sensitivity to azoles while remaining unaffected by other stressors tested. AdditionalmSpherePWWP domain-containing protein Crf4-3 specifically modulates fungal azole susceptibility by regulating sterol C-14 demethylase ERG11.PMC11774033No.32370082No.32100056No.5242020Zhou MHu CLi SZhang ZYu PYe SZhang LSun XfalsePWWP domain-containing protein Crf4-3 specifically modulates fungal azole susceptibility by regulating sterol C-14 demethylase ERG11.The widespread use of azole antifungals in agriculture and clinical settings has led to serious drug resistance. Overexpression of the azole drug target 14α-demethylase ERG11 (CYP51) is the most common fungal resistance mechanism. However, the presence of additional regulatory proteins in the transcriptional response of <i>erg11</i> is not yet fully elucidated. In this study, leveraging the identified key promoter region of <i>erg11</i> that controls its response to azoles in <i>Neurospora crassa</i>, we pinpointed a protein, Crf4-3, which harbors a PWWP domain and exerts a positive regulatory influence on azole resistance, as determined by DNA pulldown assays. The removal of Crf4-3 results in heightened sensitivity to azoles while remaining unaffected by other stressors tested. Additional2025-01-01T00:00:00Z2025 Jan2026-06-02T10:10:58.498Z2025-04-03T23:26:44.54ZS-EPMC117740333967073010.1128/msphere.00703-24