2 GENES DIFFERENTIALLY REGULATED IN THE CELL-CYCLE AND BY DNA-DAMAGING AGENTS ENCODE ALTERNATIVE REGULATORY SUBUNITS OF RIBONUCLEOTIDE REDUCTASE
2 GENES DIFFERENTIALLY REGULATED IN THE CELL-CYCLE AND BY DNA-DAMAGING AGENTS ENCODE ALTERNATIVE REGULATORY SUBUNITS OF RIBONUCLEOTIDE REDUCTASE
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DOI：
10.1101/gad.4.5.740
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发表时间：
1990-05-01
影响因子：
10.5
通讯作者：
DAVIS, RW
中科院分区：
文献类型：
--
作者：
ELLEDGE, SJ;DAVIS, RW
核糖核苷酸还原酶活性对细胞周期的进展至关重要，催化DNA合成所需的脱氧核糖核苷酸生产的限速步骤。该酶的酶活性在细胞周期中波动，在S期具有最大活性。我们已经确定并表征了两个酿酒酵母基因编码的核糖核苷酸还原酶，RNR 1和RNR 3的调节亚基。它们彼此共有约80%的氨基酸同一性，与哺乳动物同源物M1共有60%的氨基酸同一性。遗传破坏表明，RNR 1基因是必不可少的有丝分裂的活力，而RNR 3基因是不是必不可少的。RNR 3的高拷贝数克隆能够抑制rnr 1突变的致死性。在细胞周期同步培养的mRNA水平的分析表明，RNR 1 mRNA的细胞周期密切调控，波动15至30倍，并协调调节与POL 1 mRNA，被表达在晚期G1和S期的细胞周期。从α-因子诱导的G1阻断RNR 1 mRNA的诱导被放线菌酮阻断，进一步确定了G1-S-相变中蛋白质合成的需要。RNR 1和RNR 3转录物都可以通过损伤DNA的处理诱导，如4-硝基喹啉-1-氧化物甲磺酸甲酯，或阻断DNA复制，如羟基脲。RNR 1可诱导3- 5倍，RNR 3可诱导> 100倍。当MATa细胞在G1期被α-SMA阻滞时，因子，RNR 1和RNR 3 mRNA仍然可被DNA损伤诱导，表明所观察到的诱导可以发生在S期之外。通过羟基脲处理抑制核糖核苷酸还原酶活性导致细胞周期停滞在S期，如大的出芽的单核细胞。这种特定的细胞周期停滞不依赖于RAD 9基因，定义了DNA合成和细胞周期进程协调的单独途径。
Ribonucleotide reductase activity is essential for progression through the cell cycle, catalyzing the rate-limiting step for the production of deoxyribonucleotides needed for DNA synthesis. The enzymatic activity of the enzyme fluctuates in the cell cycle with an activity maximum in S phase. We have identified and characterized two Saccharomyces cerevisiae genes encoding the regulatory subunit of ribonucleotide reductase, RNR1 and RNR3. They share .apprx.80% amino acid identity with each other and 60% with the mammalian homolog, M1. Genetic disruption reveals that the RNR1 gene is essential for mitotic viability, whereas the RNR3 gene is not essential. A high-copy-number clone of RNR3 is able to suppress the lethality of rnr1 mutations. Analysis of mRNA levels in cell-cycle-synchronized cultures reveals that the RNR1 mRNA is tightly cell-cycle regulated, fluctuating 15- to 30-fold, and is coordinately regulated with the POL1 mRNA, being expressed in the late G1 and S phases of the cell cycle. Progression from the .alpha.-factor-induced G1 block to induction of RNR1 mRNA is blocked by cycloheximide, further defining the requirement for protein synthesis in the G1- to S-phase transition. Both RNR1 and RNR3 transcripts are inducible by treatments that damage DNA, such as 4-nitroquinoline-1-oxide an methylmethanesulfonate, or block DNA replication, such as hydroxyurea. RNR1 is inducible 3- to 5-fold, and RNR3 is inducible > 100-fold. When MATa cells are arrested in G1 by .alpha.-factor, RNR1 and RNR3 mRNA is still inducible by DNA damage, indicating that the observed induction can occur outside of S phase. Inhibition of ribonucleotide reductase activity by hydroxyurea treatment results in arrest of the cell cycle in S phase as large budded, uninucleate cells. This specific cell-cycle arrest is independent of the RAD9 gene, defining a separate pathway for the coordination of DNA synthesis and cell-cycle progression.