Yeast Dun1 Kinase Regulates Ribonucleotide Reductase Inhibitor Sml1 in Response to Iron Deficiency

Sanvisens, Nerea; Romero, Antonia María; An, Xiuxiang; Zhang, Caiguo; Llanos, Rosa de; Martínez‐Pastor, María Teresa; Bañó, M. Carmen; Huang, Mingxia; Puig, Sergi

doi:10.1128/mcb.00472-14

Cited by 36 publications

(51 citation statements)

References 79 publications

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“…1, D and E, and 2, B and C). This was not a consequence of a reduction in Dun1-D328A protein abundance because its expression levels under low iron conditions were similar to those of wild-type Dun1 protein (47). These results reveal that Dun1 kinase activity is required for efficient nucleus-to-cytoplasm redistribution of RNR small subunit in response to iron deprivation and DNA damage.…”

Section: Dun1 Kinase Activity Is Essential For Subcellular Redistribumentioning

confidence: 58%

“…Then, Dun1 phosphorylates Sml1 protein, which is ubiquitylated and degraded to facilitate RNR activation under low iron conditions (47). We have also shown that the Rnr2-Rnr4 heterodimer is redistributed to the cytoplasm when iron availability is low (46).…”

Section: Dun1 Promotes Rnr2 and Rnr4mentioning

confidence: 71%

“…2, D and E). We have previously shown that Dun1-T380A protein levels diminish to 60% in iron-deficient conditions (47). However, this decrease in protein abundance can only partially, but not fully, explain the strong Rnr2-Rnr4 nuclear retention displayed by Dun1-T380A expressing cells.…”

Section: Contribution Of Dun1 Phosphorylation Sites To Rnr2 and Rnr4mentioning

confidence: 95%

“…The resulting decrease in Wtm1 protein abundance promotes Rnr2-Rnr4 relocalization to the cytoplasm and dNTP synthesis (46). Furthermore, we have reported that, in response to iron deficiency, Dun1 checkpoint kinase induces degradation of the Rnr1 inhibitor protein Sml1, promoting RNR activity (47).…”

mentioning

confidence: 97%

“…Redistribution to the Cytoplasm in Response to Iron Deficiency-We have previously shown that iron deficiency activates Dun1 protein kinase (47). Then, Dun1 phosphorylates Sml1 protein, which is ubiquitylated and degraded to facilitate RNR activation under low iron conditions (47).…”

Section: Dun1 Promotes Rnr2 and Rnr4mentioning

confidence: 99%

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Yeast Dun1 Kinase Regulates Ribonucleotide Reductase Small Subunit Localization in Response to Iron Deficiency

Sanvisens

Romero

Zhang

et al. 2016

Journal of Biological Chemistry

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Ribonucleotide reductase (RNR) is an essential iron-dependent enzyme that catalyzes deoxyribonucleotide synthesis in eukaryotes. Living organisms have developed multiple strategies to tightly modulate RNR function to avoid inadequate or unbalanced deoxyribonucleotide pools that cause DNA damage and genome instability. Yeast cells activate RNR in response to genotoxic stress and iron deficiency by facilitating redistribution of its small heterodimeric subunit Rnr2-Rnr4 from the nucleus to the cytoplasm, where it forms an active holoenzyme with large Rnr1 subunit. Dif1 protein inhibits RNR by promoting nuclear import of Rnr2-Rnr4. Upon DNA damage, Dif1 phosphorylation by the Dun1 checkpoint kinase and its subsequent degradation enhances RNR function. In this report, we demonstrate that Dun1 kinase triggers Rnr2-Rnr4 redistribution to the cytoplasm in response to iron deficiency. We show that Rnr2-Rnr4 relocalization by low iron requires Dun1 kinase activity and phosphorylation site Thr-380 in the Dun1 activation loop, but not the Dun1 forkhead-associated domain. By using different Dif1 mutant proteins, we uncover that Dun1 phosphorylates Dif1 Ser-104 and Thr-105 residues upon iron scarcity. We observe that the Dif1 phosphorylation pattern differs depending on the stimuli, which suggests different Dun1 activating pathways. Importantly, the Dif1-S104A/T105A mutant exhibits defects in nucleus-to-cytoplasm redistribution of Rnr2-Rnr4 by iron limitation. Taken together, these results reveal that, in response to iron starvation, Dun1 kinase phosphorylates Dif1 to stimulate Rnr2-Rnr4 relocalization to the cytoplasm and promote RNR function. Ribonucleotide reductase (RNR)5 catalyzes the rate-limiting step in the de novo deoxyribonucleotide (dNTP) synthesis by converting ribonucleoside diphosphates to the corresponding deoxy forms. In eukaryotes, the RNR holoenzyme is composed of a large or R1 subunit that contains the catalytic and allosteric sites, and a small or R2 subunit that harbors a di-iron center, which is responsible for generating and keeping a tyrosyl radical required for catalysis (reviewed in Refs.

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Section: Dun1 Kinase Activity Is Essential For Subcellular Redistribumentioning

confidence: 58%

Section: Dun1 Promotes Rnr2 and Rnr4mentioning

confidence: 71%

Section: Contribution Of Dun1 Phosphorylation Sites To Rnr2 and Rnr4mentioning

confidence: 95%

mentioning

confidence: 97%

Section: Dun1 Promotes Rnr2 and Rnr4mentioning

confidence: 99%

See 3 more Smart Citations

Yeast Dun1 Kinase Regulates Ribonucleotide Reductase Small Subunit Localization in Response to Iron Deficiency

Sanvisens

Romero

Zhang

et al. 2016

Journal of Biological Chemistry

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Functional role of metalloproteins in genome stability

Zhang

Zhou

et al. 2016

Front. Biol.

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Functional Analysis of Ribonucleotide Reductase from Cordyceps militaris Expressed in Escherichia coli

Kato

Suparmin

Park

2017

Appl Biochem Biotechnol

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Cordyceps militaris produces cordycepin (3'-deoxyadenosine), which has various activities, including anti-oxidant, anti-tumoral, anti-viral, and anti-inflammatory. Ribonucleotide reductase (RNR) seems to be a candidate to produce cordycepin in C. militaris because RNR catalyzes the reduction of nucleotides to 2'-deoxynucleotides, whose structures are similar to that of cordycepin. However, the role of RNR has not been confirmed yet. In this study, complementary DNAs (cDNAs) of C. militaris RNR (CmRNR) large and small subunits (CmR1 and CmR2) were cloned from C. militaris NBRC9787 to investigate the function of CmRNR for its cordycepin production. C. militaris NBRC9787 began to produce cordycepin when grown in a liquid surface culture in medium composed of glucose and yeast extract for 15 days. CmR1 cDNA and CmR2 cDNA were obtained from its genomic DNA and from total RNA extracted from its mycelia after cultivation for 21 days, respectively. Recombinant CmR1 and CmR2 were expressed individually in Escherichia coli and purified. Purified recombinant CmR1 and CmR2 showed RNR activity toward adenosine diphosphate (ADP) only when two subunits were mixed but only show the reduction of ADP to 2'-deoxyADP. These results indicate that the pathway from ADP to 3'deoxyADP via CmRNR does not exist in C. militaris and cordycepin production in C. militaris may be mediated by other enzymes.

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Yeast Dun1 Kinase Regulates Ribonucleotide Reductase Inhibitor Sml1 in Response to Iron Deficiency

Cited by 36 publications

References 79 publications

Yeast Dun1 Kinase Regulates Ribonucleotide Reductase Small Subunit Localization in Response to Iron Deficiency

Yeast Dun1 Kinase Regulates Ribonucleotide Reductase Small Subunit Localization in Response to Iron Deficiency

Functional role of metalloproteins in genome stability

Functional Analysis of Ribonucleotide Reductase from Cordyceps militaris Expressed in Escherichia coli

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