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Abstract Intracellular transfers of mitochondrial DNA continue to shape nuclear genomes. Chromosome 2 of the model plant Arabidopsis thaliana contains one of the largest known nuclear insertions of mitochondrial DNA (numts). Estimated at over 600 kb in size, this numt is larger than the entire Arabidopsis mitochondrial genome. The primary Arabidopsis nuclear reference genome contains less than half of the numt because of its structural complexity and repetitiveness. Recent data sets generated with improved long-read sequencing technologies (PacBio HiFi) provide an opportunity to finally determine the accurate sequence and structure of this numt. We performed a de novo assembly using sequencing data from recent initiatives to span the Arabidopsis centromeres, producing a gap-free sequence of the Chromosome 2 numt, which is 641 kb in length and has 99.933% nucleotide sequence identity with the actual mitochondrial genome. The numt assembly is consistent with the repetitive structure previously predicted from fiber-based fluorescent in situ hybridization. Nanopore sequencing data indicate that the numt has high levels of cytosine methylation, helping to explain its biased spectrum of nucleotide sequence divergence and supporting previous inferences that it is transcriptionally inactive. The original numt insertion appears to have involved multiple mitochondrial DNA copies with alternative structures that subsequently underwent an additional duplication event within the nuclear genome. This work provides insights into numt evolution, addresses one of the last unresolved regions of the Arabidopsis reference genome, and represents a resource for distinguishing between highly similar numt and mitochondrial sequences in studies of transcription, epigenetic modifications, and de novo mutations. 

Centromeres attach chromosomes to spindle microtubules during cell division and, despite this conserved role, show paradoxically rapid evolution and are typified by complex repeats. We used long-read sequencing to generate the Col-CEN Arabidopsis thaliana genome assembly that resolves all five centromeres. The centromeres consist of megabase-scale tandemly repeated satellite arrays, which support CENTROMERE SPECIFIC HISTONE H3 (CENH3) occupancy and are densely DNA methylated, with satellite variants private to each chromosome. CENH3 preferentially occupies satellites that show the least amount of divergence and occur in higher-order repeats. The centromeres are invaded by ATHILA retrotransposons, which disrupt genetic and epigenetic organization. Centromeric crossover recombination is suppressed, yet low levels of meiotic DNA double-strand breaks occur that are regulated by DNA methylation. We propose that Arabidopsis centromeres are evolving through cycles of satellite homogenization and retrotransposon-driven diversification. 

Meiotic crossovers facilitate chromosome segregation and create new combinations of alleles in gametes. Crossover frequency varies along chromosomes and crossover interference limits the coincidence of closely spaced crossovers. Crossovers can be measured by observing the inheritance of linked transgenes expressing different colors of fluorescent protein inArabidopsispollen tetrads. Here we establish DeepTetrad, a deep learning‐based image recognition package for pollen tetrad analysis that enables high‐throughput measurements of crossover frequency and interference in individual plants. DeepTetrad will accelerate the genetic dissection of mechanisms that control meiotic recombination.

 

During meiosis homologous chromosomes undergo crossover recombination. Sequence differences between homologs can locally inhibit crossovers. Despite this, nucleotide diversity and population-scaled recombination are positively correlated in eukaryote genomes. To investigate interactions between heterozygosity and recombination we crossed Arabidopsis lines carrying fluorescent crossover reporters to 32 diverse accessions and observed hybrids with significantly higher and lower crossovers than homozygotes. Using recombinant populations derived from these crosses we observed that heterozygous regions increase crossovers when juxtaposed with homozygous regions, which reciprocally decrease. Total crossovers measured by chiasmata were unchanged when heterozygosity was varied, consistent with homeostatic control. We tested the effects of heterozygosity in mutants where the balance of interfering and non-interfering crossover repair is altered. Crossover remodeling at homozygosity-heterozygosity junctions requires interference, and non-interfering repair is inefficient in heterozygous regions. As a consequence, heterozygous regions show stronger crossover interference. Our findings reveal how varying homolog polymorphism patterns can shape meiotic recombination.