Biology subjects

Henry, I. M.

Publications and source records attributed to Henry, I. M..

11 recordsLinked to original sources

bioRxiv2025

Tobacco twin and single spots result from genome instability, chromosome breaks, and translocations

Somatic recombination can have profound consequences including clonal evolution. In plants, however, the contribution of homologous recombination (HR) to genomic instability remains unclear. The semidominant sulfur mutant of tobacco, named for its yellow foliage resulting from a chlorophyll defect, spontaneously develops green and white twin spots that have been attributed to mitotic crossovers. To test this hypothesis, we sequenced DNA from twin spots and surrounding tissues in su/+ F1 hybrids, expecting somatic crossovers to result in reciprocal chromosome arm exchange in the paired sectors. Contrary to expectations, 20 of 22 twin spots exhibited aneuploidy or reciprocal translocations between non-homologous chromosomes. One twin spot showed a pattern consistent with a reciprocal crossover. Single white or green spots were [~]10X more frequent than twin spots. Among 26 single spots, 18 resulted from terminal arm deletion, 2 from aneuploidy, and 4 were associated with translocations. Deletion and reciprocal exchange breakpoints clustered near the centromere. Together, these findings indicate that nearly all spots arise from deletions, translocations, and other rearrangements likely mediated by chromosome missegregation or nonhomologous repair rather than by HR. These rearrangements altered the ratio of mutant to wild-type alleles and consequently chlorophyll levels. We conclude that mitotic recombination between homologous chromosomes is rare, and that genome instability is the dominant driver of somatic chimerism in this system.

genetics↗

bioRxiv2023

A K-mer based Bulked Segregant Analysis approach to map seed traits in unphased heterozygous potato genomes

High-throughput sequencing-based methods for bulked segregant analysis (BSA) allow for the rapid identification of genetic markers associated with traits of interest. BSA studies have successfully identified qualitative (binary) and quantitative trait loci using QTL mapping. However, most traditional QTL mapping approaches require a reference genome. Here, we combine high throughput short read sequencing with bulk segregant analysis of k-mers (BSA-k-mer). This method can be applied to any organism and is particularly useful for species with genomes too different from the closest sequenced genome. It is also instrumental when dealing with highly heterozygous and polyploid genomes without phased haplotype assemblies and for which a single haplotype can control a trait. Finally, it is flexible in terms of population structure. Here, we present the application of the BSA-k-mer method for the rapid identification of candidate regions related to seed spot and seed size in diploid potato. While conventional QTL mapping of parental genotypes did not generate any signal, candidate loci were identified for each trait using the BSA-k-mer approach. The seed spot loci match with previously identified loci associated with pigmentation in potato. The loci associated with seed size are novel. Both sets of loci are potentially instrumental in future breeding towards true seeds in potato. Article SummaryIdentifying genes linked to agronomic traits in potatoes is challenging because potato genomes are complex and variable. We are investigating the genetic basis of seed size and color. Seeds were categorized as large or small, or spotted or not, based on simple visual observation. Next, DNA sequences from each individual were mined for association between random short sequences (k-mers) and those two traits. This more flexible method allowed us to identify regions of the potato genome associated with both traits.

plant biology↗

bioRxiv2023

Differential mutation rates in plant meristematic layers

The upper plant body is formed by the continued growth of shoot apical meristems. In angiosperms, meristems are organized in three cell layers that tend to remain clonally isolated. Somatic variants emerge when mutant cells overtake part or all of a meristem. During sexual reproduction, only one layer, the L2, contributes to the next generation by forming gametes. The L2 is known to frequently invade and replace the L3, while L1-L2 separation is persistent. The role of different meristem layers in mutation accumulation is unknown. We discovered a potato periclinal chimera in which the L2 and L3, but not the L1, are marked by a chromosomal translocation. This enabled the identification of plants regenerated from leaf protoplasts originating from either the L1 or L2+L3. Leveraging these layer-specific clones, we identified and compared mutations that accumulated in the layers in the clonal parent for several decades. Here we show that the L1 accumulates mutations at 1.9 times the rate of the L2, indicating that plants might protect the germline by mechanisms that reduce the rate of mutation accumulation in the L2. In contrast to these layer-specific mutations, we found no evidence of somatic mutation fixation in all three meristem layers. Our findings highlight how periclinal chimeras are formed by independent mutational processes in which a mutation-prone epidermal layer could increase clonal variation.

plant biology↗

bioRxiv2022

Chromoanagenesis in the asy1 meiotic mutant of Arabidopsis

Chromoanagenesis is a catastrophic event that involves localized chromosomal shattering and reorganization. In this study, we report a case of chromoanagenesis resulting from defective meiosis in the MEIOTIC ASYNAPTIC MUTANT 1 (asy1) background in Arabidopsis thaliana. We provide a detailed characterization of the genomic structure of this individual with a severely shattered segment of chromosome 1. We reveal more than 300 candidate novel DNA junctions in the affected region, confirming that asy1-related defective meiosis is a potential trigger for chromoanagenesis. This is the first example of chromoanagenesis associated with female meiosis and indicates the potential for genome evolution during oogenesis.

genomics↗

bioRxiv2022

The Arabidopsis Hop1 homolog ASY1 mediates cross-over assurance and interference

The chromosome axis plays a crucial role in meiotic recombination. Here, we study the function of ASY1, the Arabidopsis homolog of the yeast chromosome axis associated component Hop1. Specifically, we characterized cross-over (CO) distribution in female and male meiosis by deep sequencing of the progeny of an allelic series of asy1 mutants. Combining data from nearly 1000 individual plants, we find that reduced ASY1 activity leads to genomic instability and sometimes drastic genomic rearrangements. We further observed that COs are less frequent and appear in more distal chromosomal regions in plants with no or reduced ASY1 activity, consistent with previous analyses. However, our sequencing approach revealed that the reduction in CO number is not as dramatic as suggested by cytological analyses. Analysis of double mutants of asy1 with mutants with three other CO factors, MUS81, MSH4 and MSH5 as well as the determination of foci number of the CO regulator MLH1 demonstrates that the majority of the COs in asy1, similar to the situation in the wildtype, largely belong to the class I, which are subject to interference. However, these COs are redistributed in asy1 mutants and typically appear much closer than in the wildtype. Hence, ASY1 plays a key role in CO interference that spaces COs along a chromosome. Conversely, since a large proportion of chromosomes do not receive any CO, we conclude that CO assurance, the process that ensures the obligatory assignment of one CO per chromosome, is also affected in asy1 mutants. Significant statementThe regulation of the number and placement of cross-overs (COs) during meiosis is critical to ensure meiotic fidelity and promote new genetic combinations. Here, we investigated the function of one of the proteins of the chromosome axis, which plays a key role in CO formation: ASY1. Our results show that COs in asy1 mutants are positioned closer to each other than in the wildtype and that, despite a roughly similar number of COs, not every chromosome receives a CO. With this, our results shed light on the mechanisms regulating two important but still poorly understood aspects of meiosis: CO assurance, which safeguards at least one CO per chromosome pair, and CO interference, which prevents two COs from occurring close to each other.

cell biology↗

bioRxiv2021

LD-CNV: rapid and simple discovery of chromosomal translocations using linkage disequilibrium between copy number variable loci

Large scale structural variations, such as chromosomal translocations, can have profound effects on fitness and phenotype, but are difficult to identify and characterize. Here, we describe a simple and effective method aimed at identifying translocations using only the dosage of sequence reads mapped on the reference genome. We binned reads on genomic segments sized according to sequencing coverage and identified instances when copy number segregated in populations. For each dosage-polymorphic 1Mb bin, we tested linkage disequilibrium with other variable bins. In nine potato (Solanum tuberosum) dihaploid families translocations affecting pericentromeric regions were common and in two cases were due to genomic misassembly. In two populations, we found evidence for translocation affecting euchromatic arms. In cv. PI 310467, a non-reciprocal translocation between chromosome 7 and 8 resulted in a 5-3 copy number change affecting several Mb at the respective chromosome tips. In cv. "Alca Tarma", the terminal arm of chromosome 4 translocated to the tip of chromosome 1. Using oligonucleotide-based fluorescent in situ hybridization painting probes (oligo-FISH), we tested and confirmed the predicted arrangement in PI 310467. In 192 natural accessions of Arabidopsis thaliana, dosage haplotypes tended to vary continuously and resulted in higher noise, but we identified pericentromeric LD suggesting the effect of repeats. This method should be useful in species where translocations are suspected because it tests linkage without the need for genotyping.

genomics↗

bioRxiv2020

Rare instances of haploid inducer DNA in potato dihaploids and ploidy-dependent genome instability

In cultivated tetraploid potato, reduction to diploidy (dihaploidy) allows hybridization to diploid germplasm, introgression breeding, and may facilitate the production of inbreds. Pollination with haploid inducers yields maternal dihaploids, as well as triploid and tetraploid hybrids. It is not known if dihaploids result from parthenogenesis, entailing development of embryos from unfertilized eggs, or genome elimination, entailing missegregation and loss of paternal chromosomes. A sign of genome elimination is the occasional persistence of haploid inducer DNA in some of the dihaploids. We characterized the genomes of 1,001 putative dihaploids and 134 hybrids produced by pollinating tetraploid clones with three haploid inducers, IVP35, IVP101, and PL4. We detected inheritance of full or partial chromosomes from the haploid inducer parent in 0.87% of the overall dihaploid progeny, irrespective of the combination of parental genotypes. Chromosomal breaks commonly affected the paternal genome in the dihaploid and tetraploid progeny, but not in the triploid progeny. Residual haploid inducer DNA is consistent with genome elimination as the mechanism of haploid induction. Further, the fact that paternal chromosome breaks are specific to dihaploids and tetraploid progeny suggests that they may be specific to 2x sperms, and supports the hypothesis that 2x sperms facilitate genome elimination.

plant biology↗

bioRxiv2019

Genomic Outcomes of Haploid Induction Crosses in Potato (Solanum tuberosum L.)

The challenges of breeding autotetraploid potato (Solanum tuberosum) have motivated the development of alternative breeding strategies. A common approach is to obtain uniparental dihaploids from a tetraploid of interest through pollination with S. tuberosum Andigenum Group (formerly S. phureja) cultivars. The mechanism underlying haploid formation of these crosses is unclear, and questions regarding the frequency of paternal DNA transmission remain. Previous reports described aneuploid and euploid progeny, which, in some cases, displayed genetic markers from the haploid inducer. Here, we surveyed a population of 167 presumed dihaploids for large-scale structural variation that would underlie chromosomal addition from the haploid inducer, and for small-scale introgression of genetic markers. In 19 progeny, we detected ten of the twelve possible trisomies and, in all cases, demonstrated the non-inducer parent origin of the additional chromosome. Deep sequencing indicated that occasional, short-tract signals appearing of haploid inducer origin were better explained as technical artifacts. Leveraging recurring CNV patterns, we documented sub-chromosomal dosage variation indicating segregation of polymorphic maternal haplotypes. Collectively, 52% of assayed chromosomal loci were classified as dosage variable. Our findings help elucidate the genomic consequences of potato haploid induction and suggest that most potato dihaploids will be free of residual pollinator DNA.

genomics↗

bioRxiv2019

The persimmon genome reveals clues to the evolution of a lineage-specific sex determination system in plants

Most angiosperms bear hermaphroditic flowers, but a few species have evolved outcrossing strategies, such as dioecy, the presence of separate male and female individuals. We previously investigated the mechanisms underlying dioecy in diploid persimmon (D. lotus) and found that male flowers are specified by repression of the autosomal gene MeGI by its paralog, the Y-encoded pseudo-gene OGI. This mechanism is thought to be lineage-specific, but its evolutionary path remains unknown. Here, we developed a full draft of the diploid persimmon genome (D. lotus), which revealed a lineage-specific genome-wide paleoduplication event. Together with a subsequent persimmon-specific duplication(s), these events resulted in the presence of three paralogs, MeGI, OGI and newly identified Sister of MeGI (SiMeGI), from the single original gene. Evolutionary analysis suggested that MeGI underwent adaptive evolution after the paleoduplication event. Transformation of tobacco plants with MeGI and SiMeGI revealed that MeGI specifically acquired a new function as a repressor of male organ development, while SiMeGI presumably maintained the original function. Later, local duplication spawned MeGIs regulator OGI, completing the path leading to dioecy. These findings exemplify how duplication events can provide flexible genetic material available to help respond to varying environments and provide interesting parallels for our understanding of the mechanisms underlying the transition into dieocy in plants.\n\nAuthor summaryPlant sexuality has fascinated scientists for decades. Most plants can self-reproduce but not all. For example, a small subset of species have evolved a system called dioecy, with separate male and female individuals. Dioecy has evolved multiple times independently and, while we do not understand the molecular mechanisms underlying dioecy in many of these species yet, a picture is starting to emerge with recent progress in several dioecious species. Here, we focused on the evolutionary events leading to dioecy in persimmon. Our previous work had identified a pair of genes regulating sex in this species, called OGI and MeGI. We drafted the whole genome sequence of diploid persimmon to investigate their evolutionary history. We discovered a lineage-specific genome duplication event, and observed that MeGI underwent adaptive evolution after this duplication. Transgenic analyses validated that MeGI newly acquired a male-suppressor function, while the other copy of this gene, SiMeGI, did not. The regulator of MeGI, OGI, resulted from a second smaller-scale duplication event, finalizing the system. This study sheds light on the role of duplication as a mechanism that promote flexible genes functions, and how it can affect important biological functions, such as the establishment of a new sexual system.

evolutionary biology↗

bioRxiv2019

Two Y chromosome-encoded genes determine sex in kiwifruit

Dioecy, the presence of male and female individuals, has evolved independently in multiple flowering plant lineages. Although theoretical models for the evolution of dioecy, such as the \"two-mutation\" model, are well established, little is known about the specific genes determining sex and their evolutionary history. Kiwifruit, a major tree crop consumed worldwide, is a dioecious species. In kiwifruit, we had previously identified a Y-encoded sex-determinant candidate gene acting as the suppressor of feminization (SuF), named Shy Girl (SyGI). Here, we identified a second Y-encoded sex-determinant that we named Friendly boy (FrBy), which exhibits strong expression in tapetal cells. Gene-editing and complementation analyses in Arabidopsis thaliana and Nicotiana tabacum indicated that FrBy acts for the maintenance of male (M) functions, independently of SyGI, and that these functions are conserved across angiosperm species. We further characterized the genomic architecture of the small (< 1 Mb) male specific region of the Y-chromosome (MSY), which harbors only two genes significantly expressed in developing gynoecia and androecia, respectively: SyGI and FrBy. Resequencing of the genome of a natural hermaphrodite kiwifruit revealed that this individual is genetically male but carries deletion(s) of parts of the Y-chromosome, including SyGI. Additionally, expression of FrBy in female kiwifruit resulted in hermaphrodite plants. These results clearly indicate that Y-encoded SyGI and FrBy act independently as the SuF and M factors in kiwifruit, respectively, and provide insight into the evolutionary path leading to a two-factor sex determination system but also a new breeding approach for dioecious species.

plant biology↗

bioRxiv2018

Translocation and duplication from CRISPR-Cas9 editing in Arabidopsis thaliana

Cut DNA ends in plants may recombine to form novel molecules. We asked whether CRISPR-Cas9 expression in plants could induce nonhomologous recombination between diverse and heterologous broken DNA ends. We induced two breaks separated by 2.3 or by 8.5 kilobases leading to duplication of the intervening DNA and meiotic transmission of the 2.3kb duplication. Two or more dsDNA breaks in nonhomologous chromosomes led to ligation of breakpoints consistent with chromosome arm translocations. Screening 881 primary transformants we obtained 195 PCR products spanning independent, expected translocation junctions involving ends produced by cutting different loci. Sequencing indicated a true positive rate of 84/91 and demonstrated the occurrence of different junction alleles. A majority of the resulting structures would be deleterious and none were transmitted meiotically. Ligation of interchromosomal, heterologous dsDNA ends suggest that the CRISPR-Cas9 can be used to engineer plant genes and chromosomes in vivo.\n\nSignificance StatementWe explored how genome editing tools such as CRISPR-Cas9 could provide new ways to tailor novel genomic combinations and arrangements. We show that distant cut ends often precisely come together, that cuts in different chromosomes can result in translocations, and that two cuts within a chromosome often result in the duplication of the intervening segment. Formation of multiple structures with precise junctions will enable engineered rearrangements that can be predicted with accuracy.

genetics↗