Targeted induction of meiotic double-strand breaks reveals chromosomal domain-dependent regulation of Spo11 and interactions among potential sites of meiotic recombination

Abstract

Meiotic recombination is initiated by programmed DNA double-strand break (DSB) formation mediated by Spo11. DSBs occur with frequency in chromosomal regions called hot domains but are seldom seen in cold domains. To obtain insights into the determinants of the distribution of meiotic DSBs, we examined the effects of inducing targeted DSBs during yeast meiosis using a UAS-directed form of Spo11 (Gal4BD-Spo11) and a meiosis-specific endonuclease, VDE (PI-SceI). Gal4BD-Spo11 cleaved its target sequence (UAS) integrated in hot domains but rarely in cold domains. However, Gal4BD-Spo11 did bind to UAS and VDE efficiently cleaved its recognition sequence in either context, suggesting that a cold domain is not a region of inaccessible or uncleavable chromosome structure. Importantly, self-association of Spo11 occurred at UAS in a hot domain but not in a cold domain, raising the possibility that Spo11 remains in an inactive intermediate state in cold domains. Integration of UAS adjacent to known DSB hotspots allowed us to detect competitive interactions among hotspots for activation. Moreover, the presence of VDE-introduced DSB repressed proximal hotspot activity, implicating DSBs themselves in interactions among hotspots. Thus, potential sites for Spo11-mediated DSB are subject to domain-specific and local competitive regulations during and after DSB formation.

Numerical and spatial patterning of yeast meiotic DNA breaks by Tel1

“…Tel1-dependent DSB interference operates over a similar size scale (see Introduction) (Garcia et al 2015), so interference can, in principle, explain the local effect of strong artificial hotspots, assuming that interference includes both sister chromatids. Site-specific endonucleolytic DSBs (i.e., not made by Spo11) also suppress Spo11-generated DSBs nearby (Fukuda et al 2008), consistent with an inhibitory signal that spreads after DSB formation. Importantly, however, it remains unknown whether the effect of artificial Spo11 hotspot insertion is the same as DSB interference: To meet the formal definition of interference, occurrence of multiple events on the same chromosome must be shown to occur less often than expected from the product of the single-event frequencies, but this has not been tested.…”

Domain-Specific Regulation of Recombination inCaenorhabditis elegansin Response to Temperature, Age and Sex

“…Regulation of double crossovers differs between the sexes: Domain specific regulation of crossovers has been proposed to explain recombination maps in multiple systems (International Hapmap Consortium 2007; Coop et al 2008;Fukuda et al 2008). One of the governing principles for crossover regulation in most systems is CI.…”

“…In addition, telomeres and centromeres establish recombinationally repressed regions (Stern 1926;Lambie and Roeder 1986;Blitzblau et al 2007). How these crossover domains are established and regulated remains an outstanding question (Dorman et al 2007;Fukuda et al 2008).…”

Rec8 Guides Canonical Spo11 Distribution along Yeast Meiotic Chromosomes

“…In addition to that, DSBs were formed in cold domains (in rad50S) located near pericentromeric regions (Blitzblau et al, 2007;Buhler et al, 2007). Moreover, the formation of the meiosis-specific Spo11 multimer, which is essential to DSBs and requires Rec102 and Rec104 functions, has not been detected in the domains Fukuda et al, 2008).…”

“…However, a regional difference in DSB competency does not simply reflect the regulation of Spo11 binding to the chromosomes, because the targeting of Spo11 fused with Gal4 DNA binding domain (Gal4BD-Spo11) to cold domains such as yeast centromeric regions cannot induce DNA cleavage during meiosis (Robine et al, 2007;Fukuda et al, 2008). To uncover the regional regulation of Spo11, it is necessary to determine the chromosome-wide dynamic distribution of Spo11 in wild-type cells.…”