Chromosome translocations drive divergence of wild grass subspecies

Aegilops comosa, an annual diploid grass with the MM genome constitution, grows mainly in coastal and inland Greece, with scattered populations in Albania, the former Yugoslavia, Turkey, Cyprus and Bulgaria. Two morphologically distinct subspecies are recognized within the species: subsp. comosa and subsp. heldreichii. Some taxonomists treat them as two distinct species, while others regard them as varieties. The two forms often grow together in mixed stands on roadsides, grasslands and hillsides, sometimes in cultivated fields. Except for Greece, Ae. comosa is uncommon to rare throughout its range. Despite decades of botanical study, the genetic and karyotypic bases of their divergence were not fully understood.

A team of researchers from the N. I. Vavilov All-Russian Institute of Plant Genetic Resources in St. Petersburg, the Leibniz Institute of Plant Genetics and Crop Plant Research in Gatersleben, and collaborating institutions set out to characterize the genome and karyotype of Ae. comosa on a broad sample of accessions. They analyzed thirty-six accessions, including twenty of comosa and sixteen of heldreichii, collected from different regions of Greece and Turkey and maintained in genetic collections. The goal was to assess the level of genetic diversity and uncover the mechanisms leading to radiation of the subspecies. The study was published in the journal Comparative Cytogenetics.

The researchers employed fluorescence in situ hybridization, or FISH, with fifteen repetitive DNA probes, together with electrophoretic analysis of seed storage proteins known as gliadins. FISH allows specific DNA sequences to be visualized directly on metaphase chromosomes, revealing the amount and distribution of microsatellite and satellite DNA families, the number and position of rDNA clusters, and structural differences between chromosomes. Gliadin electrophoresis, a relatively cheap but informative marker for polymorphism analysis, separates seed storage proteins by electrophoretic mobility, producing spectra traditionally divided into four zones, alpha, beta, gamma and omega, controlled by genes on different homoeologous chromosome groups.

The results show that the two subspecies have similar overall karyotype structures, comprising metacentric, submetacentric and subacrocentric chromosomes, indicating that they belong to one biological species. However, they differ clearly in the size and morphology of chromosomes 3M and 6M. Chromosome 3M of comosa is larger and more asymmetric, submetacentric, than the metacentric 3M of heldreichii, due to the loss of a distal part of the long arm. The satellite on chromosome 6M is much larger in heldreichii, occupying nearly half of the arm length, compared with approximately one third to one quarter in comosa. The researchers hypothesize that these differences are due to a reciprocal translocation between chromosomes 3M and 6M, identified earlier by meiotic analysis of F1 hybrids.

The subspecies also differ in the amount and distribution of microsatellite and satellite DNA sequences. Hybridization with the GAA microsatellite probe produces C-banding-like patterns, with comosa chromosomes carrying numerous GAA signals of variable size in subterminal, interstitial and pericentromeric regions, while heldreichii possesses a lesser number of sites, most with low or moderate intensity. Both subspecies exhibit extremely broad polymorphism of GAA labeling patterns, with differences between accessions, between individual plants within an accession, and even between homologous chromosomes. Satellite DNA probes pTa-713, pSc119.2, pAs1, pTa-k566, oligo-44 and oligo-45 generate clear and reproducible patterns, several of which are subspecies-specific, allowing precise discrimination of comosa from heldreichii accessions.

The number and position of minor nucleolar organizer regions, especially on chromosomes 3M and 6M, also discriminate the subspecies. The pTa71 probe visualizes all major and minor rDNA loci, while a newly designed oligo-18S probe fails to reveal major NORs and Ae. comosa-specific terminal minor NORs on chromosomes 2M, 3M and 5M, but is more efficient in detecting minor NORs in interstitial and pericentromeric regions of many M-genome chromosomes. Several rDNA sites are common for both subspecies, while others are specific: a locus of variable intensity in the proximal part of chromosome 3M is observed in nearly all comosa genotypes but not in heldreichii, and minor pericentromeric sites on chromosomes 1M and 4M differ in position between the subspecies.

Gliadin electrophoresis of twenty-six accessions reveals extremely broad intraspecific variation: each accession shows a unique profile. Spectra of comosa accessions are usually more enriched in components compared to heldreichii, with a larger number of protein bands and higher intensities, especially in the alpha-zone controlled by chromosomes of homoeologous group 6. All heldreichii accessions carrying the large satellite on chromosome 6M showed relatively poor gliadin profiles. The researchers suggest that the 3M-6M translocation can change the functioning of gliadin loci on chromosome 6M of heldreichii, resulting in depletion of gliadin spectra in the alpha-zone. Based on common blocks of polypeptide bands, six gliadin families were discriminated among the accessions, with families including accessions mainly from one subspecies.

The study also identified frequent hybrids, which the researchers attribute to open pollination, overlapping heading and flowering times, and the partial fertility of hybrids between the subspecies. Accession K-1601 shows the highest heterogeneity, with each of seven genotypes examined by FISH showing unique patterns, and some genotypes sharing karyotypic features of both subspecies. Three accessions assigned to comosa based on botanical characters combine chromosomal features of both subspecies, suggesting hybrid origin followed by karyotype stabilization. One accession, K-3809, contains genotypes representing hybrids of Ae. comosa with an unknown tetraploid wheat species. The lack of geographic or genetic barriers between the subspecies, along with genetic heterogeneity of accessions, may contribute to the extremely broad intraspecific variation of GAA patterns and gliadin profiles, which is usually not observed in endemic plant species.

The researchers justify the classification of chromosomes 2M and 5M, which are difficult to discriminate owing to similar morphology and DNA content, by using group-specific FISH markers: the 5S rDNA locus occurs only on group 1 and 5 chromosomes, and its signal in the middle of the short arm confirms that the chromosome belongs to genetic group 5. They also suggest a set of DNA probes for precise identification of each of the seven M-genome chromosomes. Aegilops comosa possesses agronomically valuable traits such as pest and disease resistance and salt tolerance, which can potentially be used for wheat improvement. Resistance to yellow rust was introduced into wheat cultivar Compair from Ae. comosa in the late 1960s by genetically induced homoeologous recombination. However, modifications of the M-genome chromosomes over the course of evolution, in particular species-specific translocations, prevent direct utilization of the Ae. comosa gene pool in wheat breeding. A deeper understanding of the genome of this species, its chromosomal structure and the range of polymorphism is required for manipulation of its genetic material.

Subject of Research: Intraspecific divergence of the diploid grass Aegilops comosa is associated with structural chromosome changes, including a subspecies-specific reciprocal translocation between chromosomes 3M and 6M, revealed by FISH and gliadin electrophoresis.

Article Title: Intraspecific divergence of diploid grass Aegilops comosa is associated with structural chromosome changes

Article References: Badaeva, E. D., Kotseruba, V. V., Fisenko, A. V., Chikida, N. N., Belousova, M. K., Zhurbenko, P. M., Surzhikov, S. A., & Dragovich, A. Y. (2023). Intraspecific divergence of diploid grass Aegilops comosa is associated with structural chromosome changes. Comparative Cytogenetics, 17, 75-112. https://doi.org/10.3897/compcytogen.17.101008

Image Credits: AI Generated

DOI: 10.3897/CompCytogen.17.101008

Keywords: Aegilops comosa, chromosome translocation, FISH, gliadin electrophoresis, karyotype, subspecies divergence, wheat improvement, M genome, Intraspecific, divergence, diploid, grass