Nitric oxide (˙NO) is a free radical that induces nitrosative stress, which can jeopardize cell viability. Yeasts have evolved diverse detoxification mechanisms to effectively counteract ˙NO-mediated cytotoxicity. One mechanism relies on the flavohemoglobin Yhb1, whereas a second one requires the S-nitrosoglutathione reductase Fmd2. To investigate heme-dependent activation of Yhb1 in response to ˙NO, we use hem1Δ-derivative Schizosaccharomyces pombe strains lacking the initial enzyme in heme biosynthesis, forcing cells to assimilate heme from external sources. Under these conditions, yhb1⁺ mRNA levels are repressed in the presence of iron through a mechanism involving the GATA-type transcriptional repressor Fep1. In contrast, when iron levels are low, the transcription of yhb1⁺ is derepressed and further induced in the presence of the ˙NO donor DETANONOate. Cells lacking Yhb1 or expressing inactive forms of Yhb1 fail to grow in a hemin-dependent manner when exposed to DETANONOate. Similarly, the loss of function of the heme transporter Str3 phenocopies the effects of Yhb1 disruption by causing hypersensitivity to DETANONOate under hemin-dependent culture conditions. Coimmunoprecipitation and bimolecular fluorescence complementation assays demonstrate the interaction between Yhb1 and the heme transporter Str3. Collectively, our findings unveil a novel pathway for activating Yhb1, fortifying yeast cells against nitrosative stress.

中文翻译：

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黄素血红蛋白 Yhb1 是血红素转运蛋白 Str3 的新相互作用伙伴


一氧化氮 (NO) 是一种自由基，可引起亚硝化应激，从而危及细胞活力。酵母已经进化出多种解毒机制来有效抵消一氧化氮介导的细胞毒性。一种机制依赖于黄素血红蛋白 Yhb1，而第二种机制则需要 S-亚硝基谷胱甘肽还原酶 Fmd2。为了研究 Yhb1 响应 ˙NO 的血红素依赖性激活，我们使用了 hem1Δ 衍生的粟酒裂殖酵母菌株，该菌株缺乏血红素生物合成中的初始酶，迫使细胞从外部来源同化血红素。在这些条件下，yhb1 ⁺ mRNA 水平在铁存在下通过涉及 GATA 型转录抑制子 Fep1 的机制受到抑制。相反，当铁水平较低时，yhb1 ⁺ 的转录受到抑制，并在 ˙NO 供体 DETANONOate 的存在下进一步被诱导。当暴露于 DETANONOate 时，缺乏 Yhb1 或表达非活性形式 Yhb1 的细胞无法以血红素依赖性方式生长。同样，血红素转运蛋白 Str3 功能的丧失通过在血红素依赖性培养条件下引起对 DETANONOate 的超敏反应来表现 Yhb1 破坏的影响。免疫共沉淀和双分子荧光互补测定证明了 Yhb1 和血红素转运蛋白 Str3 之间的相互作用。总的来说，我们的研究结果揭示了一种激活 Yhb1 的新途径，可增强酵母细胞抵抗亚硝化应激的能力。