CMG helicase disassembly is essential and driven by two pathways in budding yeast.

IF 9.4 1区 生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY EMBO Journal Pub Date : 2024-09-01 Epub Date: 2024-07-22 DOI:10.1038/s44318-024-00161-x
Cristian Polo Rivera, Tom D Deegan, Karim P M Labib
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Abstract

The CMG helicase is the stable core of the eukaryotic replisome and is ubiquitylated and disassembled during DNA replication termination. Fungi and animals use different enzymes to ubiquitylate the Mcm7 subunit of CMG, suggesting that CMG ubiquitylation arose repeatedly during eukaryotic evolution. Until now, it was unclear whether cells also have ubiquitin-independent pathways for helicase disassembly and whether CMG disassembly is essential for cell viability. Using reconstituted assays with budding yeast CMG, we generated the mcm7-10R allele that compromises ubiquitylation by SCFDia2. mcm7-10R delays helicase disassembly in vivo, driving genome instability in the next cell cycle. These data indicate that defective CMG ubiquitylation explains the major phenotypes of cells lacking Dia2. Notably, the viability of mcm7-10R and dia2∆ is dependent upon the related Rrm3 and Pif1 DNA helicases that have orthologues in all eukaryotes. We show that Rrm3 acts during S-phase to disassemble old CMG complexes from the previous cell cycle. These findings indicate that CMG disassembly is essential in yeast cells and suggest that Pif1-family helicases might have mediated CMG disassembly in ancestral eukaryotes.

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在芽殖酵母中,CMG 螺旋酶的分解是必不可少的,并由两种途径驱动。
CMG螺旋酶是真核生物复制体的稳定核心,在DNA复制终止时被泛素化和分解。真菌和动物使用不同的酶对CMG的Mcm7亚基进行泛素化,这表明CMG泛素化在真核生物进化过程中反复出现。直到现在,人们还不清楚细胞是否也有不依赖泛素的螺旋酶分解途径,也不清楚CMG分解是否是细胞存活的必要条件。我们利用芽殖酵母 CMG 的重组实验,产生了 mcm7-10R 等位基因,它损害了 SCFDia2 的泛素化作用。mcm7-10R 在体内延迟了螺旋酶的分解,导致下一个细胞周期中基因组的不稳定性。这些数据表明,CMG泛素化缺陷解释了缺乏Dia2的细胞的主要表型。值得注意的是,mcm7-10R 和 dia2∆ 的活力依赖于相关的 Rrm3 和 Pif1 DNA 螺旋酶,它们在所有真核生物中都有直向同源物。我们发现,Rrm3 在 S 期的作用是分解上一个细胞周期的旧 CMG 复合物。这些发现表明,CMG的分解在酵母细胞中是必不可少的,并表明Pif1家族螺旋酶可能在真核生物的祖先中介导了CMG的分解。
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来源期刊
EMBO Journal
EMBO Journal 生物-生化与分子生物学
CiteScore
18.90
自引率
0.90%
发文量
246
审稿时长
1.5 months
期刊介绍: The EMBO Journal has stood as EMBO's flagship publication since its inception in 1982. Renowned for its international reputation in quality and originality, the journal spans all facets of molecular biology. It serves as a platform for papers elucidating original research of broad general interest in molecular and cell biology, with a distinct focus on molecular mechanisms and physiological relevance. With a commitment to promoting articles reporting novel findings of broad biological significance, The EMBO Journal stands as a key contributor to advancing the field of molecular biology.
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