Regulation of meiotic prophase checkpoint function by the AAA+ ATPase Pch2 and the HORMA protein Hop1

Meiosis is a specialized cell division program utilized by sexually reproducing organisms to produce gametes (e.g. egg and sperm). As cell progress through meiotic prophase, the formation of physical linkages between the initially unpaired homologous chromosomes is established. Without the linkage between homologs, the probability of chromosome missegregation, and the associated rate of aneuploidy, increases manifold during chromosomal segregation events. Recombination, where programmed DNA double-strand breaks (DSBs) are repaired via homologous chromosomes, generates physical linkages between homologous chromosomes. This process is facilitated by synapsis, where two homologs come in close proximity to each other by virtue of the polymerization of synaptonemal complex (SC) at the interface of two homologs. Checkpoints operate during meiotic prophase to ensure the timely execution of both recombination and synapsis. Hop1, a HORMA domain-containing protein plays a central role in checkpoint signaling cascade emanating because of defects in either recombination or synapsis. On the other hand, Pch2, an AAA+ ATPase, is believed to play a role exclusively in relaying checkpoint function in response to defects in synapsis. Although Pch2 is known to execute almost all of its functions via its action on a single client protein, Hop1, it is not fully understood why Pch2 plays a role specifically in only one checkpoint signaling operating during the meiotic prophase. In addition, how does Pch2 function in checkpoint regulation during meiotic prophase has remained elusive.

Chromosomal Hop1 is essential for the proper assembly of SC, and Pch2 regulates the chromosomal abundance of Hop1. Deletion of PCH2 leads to increased colocalization of Hop1 and Zip1, the central component of SC on the chromosomes. In this doctoral work, by exploiting budding yeast genetics, I provide a framework of the feedback mechanism between Pch2, Hop1, and Zip1 responsible for regulating the SC assembly. I show that Pch2 restricts the chromosomal recruitment of Zip1 by modulating the chromosomal abundance of Hop1. I also revisited the idea of ​​multiple checkpoints operating during meiotic prophase and the role of Pch2 in the signaling of these checkpoints. I show that, in contrast to the current understanding, Pch2 plays a role in both recombination and synapsis checkpoint signaling. I propose that both these checkpoints follow the same signaling logic in which Pch2 and Hop1 play a combined role in optimal checkpoint signaling. I also define and explain the dual role of Pch2 in checkpoint signaling whereby Pch2, depending on the cellular context, can play the role of an agonist as well as an antagonist in checkpoint signaling. I show that, under conditions where Pch2 cannot be recruited to the chromosomes, Pch2 helps in facilitating the incorporation of Hop1 in chromosome-based checkpoint signaling. On the other hand, under conditions where Pch2 can be recruited to the chromosomes, Pch2 fuels checkpoint silencing by removing Hop1 from chromosomes. Pch2 is able to perform this checkpoint silencing activity even in the face of unrepaired DSBs. Although multiple factors control chromosomal recruitment of Pch2, I show that, under unperturbed conditions, by determining the cellular localization of Pch2, polymerization of SC dictates switch-like checkpoint behavior of Pch2. Collectively, this study shows that Pch2 and Hop1 establishes a dynamic AAA + HORMA module that regulates the meiotic checkpoint function in a manner analogous to the way the AAA + HORMA module of Pch2 / TRIP13 and Mad2 acts during checkpoint that monitors chromosome segregation during mitosis and meiosis. Polymerization of SC dictates switch-like checkpoint behavior of Pch2. Collectively, this study shows that Pch2 and Hop1 establishes a dynamic AAA + HORMA module that regulates the meiotic checkpoint function in a manner analogous to the way the AAA + HORMA module of Pch2 / TRIP13 and Mad2 acts during checkpoint that monitors chromosome segregation during mitosis and meiosis. Polymerization of SC dictates switch-like checkpoint behavior of Pch2. Collectively, this study shows that Pch2 and Hop1 establishes a dynamic AAA + HORMA module that regulates the meiotic checkpoint function in a manner analogous to the way the AAA + HORMA module of Pch2 / TRIP13 and Mad2 acts during checkpoint that monitors chromosome segregation during mitosis and meiosis.

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Raina, V.B., 2020. Regulation of meiotic prophase checkpoint function by the AAA+ ATPase Pch2 and the HORMA protein Hop1. https://doi.org/10.17185/duepublico/73479
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