Mechanisms of Chromosome Scale Signal Propagation

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Project summary The overall goal of this project is to determine how cells communicate chromosome break formation and repair across large chromosomal distances. DNA double-strand breaks (DSBs) are dangerous insults to genome integrity because of their potential to cause chromosome rearrangements and chromosome instability, both of which are strongly associated with cancer progression as well as birth defects. Remarkably, meiotic cells are able to efficiently orchestrate the formation and repair of hundreds of concurrent DSBs across their genome during meiotic recombination, a process that is essential for proper gamete formation and fertility. A key feature of meiotic DSB formation and repair is its coordination at the chromosomal level. In the previous funding period we provided evidence that the synaptonemal complex, a conserved protein lattice that forms between aligned homologous chromosomes in late meiotic prophase, communicates repair decisions along meiotic chromosomes in S. cerevisiae. We showed that this communication resulted in reduced DSB formation as well as simplified repair, and we identified several factors involved in this process. We now discovered the existence of privileged genomic regions near the ends of all chromosomes that appear resistant to regulation by the synaptonemal complex. These end-adjacent regions (EARs) cover large genomic distances (~100kb, which is nearly half the length of the shortest chromosome) and continue to form and repair DSBs well after DSB formation has stopped in the rest of the genome. Similar regions of elevated meiotic recombination are also observed in birds, chimps, and humans. The goal of this project is to define the chromosomal signal that generates these regions and to test if EARs help inheritance of short chromosomes. Our preliminary analyses suggest several roles of the nuclear envelope, both in the establishment of the EARs and in the suppression of DSBs in the rest of the genome. The dynamics of chromosomal signaling and its interaction with the nuclear envelope will be analyzed by genome-wide binding studies and super-resolution microscopy, taking advantage of a conditional nuclear depletion approach that we recently introduced into meiotic cells that allows stage-specific knock-downs of pleiotropic nuclear factors. In addition, signal integration will be analyzed using genetic epistasis analyses, cytology, and physical analysis of DSB formation. As EARs cover a proportionally much larger fraction of short chromosomes, the proposal will also use tetrad sequencing to test if these regions drive the widely observed increase in recombination rates on short chromosomes. Fluorescent marker segregation will be used to determine if EARs differentially improve the meiotic segregation fidelity of short chromosomes. Together, these analyses will provide key insights into the mechanisms of chromosomal signal propagation, and open new avenues for understanding the origins of birth defects such as Down syndrome (trisomy 21) and Edwards syndrome (trisomy 18), which are caused by meiotic missegregation of short chromosomes.