HIGH CROSSOVER RATE1 encodes PROTEIN PHOSPHATASE X1 and restricts meiotic crossovers in Arabidopsis

Abstract

Meiotic crossovers are tightly restricted in most eukaryotes, despite an excess of initiating DNA double-strand breaks. The majority of plant crossovers are dependent on Class I interfering repair, with a minority formed via the Class II pathway. Class II repair is limited by anti-recombination pathways, however similar pathways repressing Class I crossovers are unknown. We performed a forward genetic screen in Arabidopsis using fluorescent crossover reporters, to identify mutants with increased or decreased recombination frequency. We identified HIGH CROSSOVER RATE1 ( HCR1 ) as repressing crossovers and encoding PROTEIN PHOSPHATASE X1. Genome-wide analysis showed that hcr1 crossovers are increased in the distal chromosome arms. MLH1 foci significantly increase in hcr1 and crossover interference decreases, demonstrating an effect on Class I repair. Consistently, yeast two-hybrid and in planta assays show interaction between HCR1 and Class I proteins, including HEI10, PTD, MSH5, and MLH1. We propose that HCR1 plays a major role in opposition to pro-recombination kinases to restrict crossovers in Arabidopsis.

Rewiring Meiosis for Crop Improvement

“…The regulation of HEI10 dosage is a promising avenue to increase CO number in crops by stabilizing the recombination events maturing into class I COs and reducing the strength of CO interference. Recent studies identified protein phosphatase X1 and ZYP1/ZEP1 as additional factors limiting class I CO formation suggesting that other strategies may be possible to increase class I CO rate ( Wang et al, 2010 , 2015 ; Capilla-Perez et al, 2021 ; France et al, 2021 ; Nageswaran et al, 2021 ).…”

Fast and Precise: How to Measure Meiotic Crossovers in Arabidopsis

“…Acid spreading is often used with MLH1/3 immunostaining to determine the number of class I COs at the diakinesis stage ( Fig. 2B ) ( Modliszewski et al, 2018 ; Nageswaran et al, 2021 ; Ziolkowski et al, 2017 ). Combining the detergent spreading method with confocal laser scanning microscopy (CLSM), MLH1/MLH3 proteins can be co-immunostained with REC8 or ZYP1 to visualize the fully synapsed chromosome axes or SC at the pachytene stage, thus enabling the number of class I COs to be counted ( Capilla-Pérez et al, 2021 ; France et al, 2021 ; Lloyd et al, 2018 ).…”

“…2C ). Images of 1,000-3,000 seeds per plant can be analyzed by CellProfiler, an image processing program that enables high-throughput measurement of sex-averaged CO frequency ( Nageswaran et al, 2021 ; Ziolkowski et al, 2015 ; 2017 ). The seed FTL system has five advantages: (1) it provides a sufficient number of seeds for each individual plant, resulting in robust CO frequency between individuals of the same genotype with little variation; (2) seeds can be stored for future large scale analysis; (3) hemizygous seeds ( FTL GR/++ ) can be pre-selected, saving time and space for plant growth; (4) a genome-wide set of seed FTLs is available for a landscape of CO frequencies along chromosomes; (5) sex-specific and average CO frequency can be measured ( Saini et al, 2020 ).…”

“…Additionally, use of the 420 line led to the identification of HEI10, TAF4B , and SNI1 as natural modifiers of CO frequency with different Arabidopsis accessions ( Lawrence et al, 2019 ; Zhu et al, 2021 ; Ziolkowski et al, 2017 ). Moreover, the 420 seed FTL high-throughput system enabled the forward genetic screening of low or high crossover rate ( lcr or hcr ) mutants using ethyl methanesulfonate mutagenesis ( Kim et al, 2021 ; Nageswaran et al, 2021 ). The seed FTL-based genetic screen revealed that HCR1 encodes a PPX1 phosphatase that limits class I COs by interacting with ZMM proteins, and that HCR2/HSBP is abundantly expressed in meiocytes and restricts the number of COs by repressing HEI10 transcription via attenuation of heat shock factor activity ( Kim et al, 2021 ; Nageswaran et al, 2021 ).…”

A Modified Meiotic Recombination in Brassica napus Largely Improves Its Breeding Efficiency

“…For instance, out of the 150 to 250 DSBs generated per meiosis, only~10 result in crossovers in Arabidopsis, the others giving rise to non-crossovers (NCOs) [5,6]. In the last decade, several proteins were highlighted to promote the repair of DSBs into NCOs in A. thaliana (e.g., FANCM, RECQ4, FIGL1, HCR1), thereby limiting the overall number of crossovers generated per meiosis [7][8][9][10]. Furthermore, crossovers are unevenly distributed along chromosomes [4], with a gradient from the telomere to the centromere, as exemplified in bread wheat [11], maize [12] or potato [13].…”