Pch2 is a meiosis-specific AAA+ protein that controls several important chromosomal processes. We previously demonstrated that Orc1, a subunit of the ORC, functionally interacts with budding yeast Pch2. The ORC (Orc1-6) AAA+ complex loads the AAA+ MCM helicase to origins of replication, but whether and how ORC collaborates with Pch2 remains unclear. Here, we show that a Pch2 hexamer directly associates with ORC during the meiotic G2/prophase. Biochemical analysis suggests that Pch2 uses its non-enzymatic NH₂-terminal domain and AAA+ core and likely engages the interface of ORC that also binds to Cdc6, a factor crucial for ORC-MCM binding. Canonical ORC function requires association with origins, but we show here that despite causing efficient removal of Orc1 from origins, nuclear depletion of Orc2 and Orc5 does not trigger Pch2/Orc1-like meiotic phenotypes. This suggests that the function for Orc1/Pch2 in meiosis can be executed without efficient association of ORC with origins of replication. In conclusion, we uncover distinct functionalities for Orc1/ORC that drive the establishment of a non-canonical, meiosis-specific AAA+ assembly with Pch2.

中文翻译：

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减数分裂特异性Pch2 / ORC AAA +装配体的生化和功能表征。

Pch2是减数分裂特异的AAA +蛋白，可控制几个重要的染色体过程。我们以前证明，Orc1，ORC的一个亚基，功能上与发芽的酵母Pch2相互作用。ORC（Orc1-6）AAA +复合体将AAA + MCM解旋酶加载到复制起点，但是ORC是否以及如何与Pch2合作尚不清楚。在这里，我们显示Pch2六聚体在减数分裂G2 /前期与ORC直接相关。生化分析表明Pch2使用其非酶促NH ₂-末端域和AAA +核心，并可能参与ORC的界面，该界面也与Cdc6结合，而Cdc6是ORC-MCM结合的关键因素。规范的ORC功能需要与起源相关联，但是我们在这里表明，尽管从起源中导致了Orc1的有效去除，但是Orc2和Orc5的核耗竭并不会触发类似Pch2 / Orc1的减数分裂表型。这表明Orc1 / Pch2在减数分裂中的功能可以在ORC与复制起点没有有效关联的情况下执行。总之，我们发现了Orc1 / ORC的不同功能，这些功能推动了Pch2的非经典减数分裂特定AAA +装配的建立。