Sanvisens Nerea, Romero Antonia M, An Xiuxiang, Zhang Caiguo, de Llanos Rosa, Martínez-Pastor María Teresa, Bañó M Carmen, Huang Mingxia, Puig Sergi
Departamento de Biotecnología, Instituto de Agroquímica y Tecnología de Alimentos (IATA), Consejo Superior de Investigaciones Científicas (CSIC), Paterna, Valencia, Spain.
Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, Denver, Colorado, USA.
Mol Cell Biol. 2014 Sep;34(17):3259-71. doi: 10.1128/MCB.00472-14. Epub 2014 Jun 23.
Iron is an essential micronutrient for all eukaryotic organisms because it participates as a redox-active cofactor in many biological processes, including DNA replication and repair. Eukaryotic ribonucleotide reductases (RNRs) are Fe-dependent enzymes that catalyze deoxyribonucleoside diphosphate (dNDP) synthesis. We show here that the levels of the Sml1 protein, a yeast RNR large-subunit inhibitor, specifically decrease in response to both nutritional and genetic Fe deficiencies in a Dun1-dependent but Mec1/Rad53- and Aft1-independent manner. The decline of Sml1 protein levels upon Fe starvation depends on Dun1 forkhead-associated and kinase domains, the 26S proteasome, and the vacuolar proteolytic pathway. Depletion of core components of the mitochondrial iron-sulfur cluster assembly leads to a Dun1-dependent diminution of Sml1 protein levels. The physiological relevance of Sml1 downregulation by Dun1 under low-Fe conditions is highlighted by the synthetic growth defect observed between dun1Δ and fet3Δ fet4Δ mutants, which is rescued by SML1 deletion. Consistent with an increase in RNR function, Rnr1 protein levels are upregulated upon Fe deficiency. Finally, dun1Δ mutants display defects in deoxyribonucleoside triphosphate (dNTP) biosynthesis under low-Fe conditions. Taken together, these results reveal that the Dun1 checkpoint kinase promotes RNR function in response to Fe starvation by stimulating Sml1 protein degradation.
铁是所有真核生物必需的微量营养素，因为它作为一种氧化还原活性辅因子参与许多生物过程，包括DNA复制和修复。真核核糖核苷酸还原酶（RNRs）是铁依赖性酶，催化二磷酸脱氧核糖核苷（dNDP）的合成。我们在此表明，Sml1蛋白（一种酵母RNR大亚基抑制剂）的水平在营养性和遗传性铁缺乏时会特异性降低，其降低方式依赖Dun1但不依赖Mec1/Rad53和Aft1。铁饥饿时Sml1蛋白水平的下降依赖于Dun1的叉头相关结构域和激酶结构域、26S蛋白酶体以及液泡蛋白水解途径。线粒体铁硫簇组装核心成分的缺失会导致Dun1依赖性的Sml1蛋白水平降低。在低铁条件下，Dun1对Sml1的下调作用的生理相关性通过dun1Δ和fet3Δ fet4Δ突变体之间观察到的合成生长缺陷得以体现，而SML1缺失可挽救该缺陷。与RNR功能增加一致，缺铁时Rnr1蛋白水平上调。最后，dun1Δ突变体在低铁条件下的三磷酸脱氧核糖核苷（dNTP）生物合成中表现出缺陷。综上所述，这些结果表明，Dun1检查点激酶通过刺激Sml1蛋白降解来促进铁饥饿时的RNR功能。
