Anna Kalinka

Cytomixis-like chromosomes/chromatin elimination from pollen mother cells (PMCs) in wheat-rye allopolyploids

Polyploidy is an important event and major force in plant speciation. Amongst the polyploids, allopolyploids have attracted special attention to investigate genetic and epigenetic mechanisms. Also, they are the means for the development of new genotypes and genomic combinations to facilitate genetic enhancement and agricultural productivity. Whereas natural allopolyploids are genetically stable and well adapted, the newly synthesized ones are highly unstable. This instability is manifested into alterations at genomic and/or phenotypic level. Here we present the phenomenon of direct chromosome/chromatin elimination from pollen mother cells (PMCs) in wheat-rye hybrids as one aspect of instability leading to irregular meiosis and disturbances in meiotic process. One of the prominent irregularities noticed is peripherally separated uncondensed or pycnotic masses of chromatin in all meiotic stages. We have observed that this chromatin undergoes elimination by budding-like way, whereby a “mini-cell” is created. It was also found that nucleoli are the first to be eliminated along with a small mass of chromatin. By means of GISH we have shown that both rye and wheat chromatin might be eliminated. In the separated groups of chromosomes/chromatin neither DNaseI nor DNase II activity was detected. Immunolocalization of tubulin allowed for differentiation between chromatin elimination from microspores and elimination from earlier stages of meiosis. It was noticeable, that in microspores special cytoskeleton structure pushing micronuclei out from the cells was created. Elimination occurred before and after meiosis as well as in each stage of meiotic division, but its intensity varied, depending on the PMC. The basis of the elimination mechanism might be the same as in cytomixis, because both phenomena share common symptoms, although cytomixis per se was rare in the analyzed hybrids.

Molecular phylogenetics, seed morphometrics, chromosome number evolution and systematics of European Elatine L. (Elatinaceae) species

The genus Elatine contains ca 25 species, all of which are small, herbaceous annuals distributed in ephemeral waters on both hemispheres. However, due to a high degree of morphological variability (as a consequence of their amphibious life-style), the taxonomy of this genus remains controversial. Thus, to fill this gap in knowledge, we present a detailed molecular phylogenetic study of this genus based on nuclear (rITS) and plastid (accD-psaI, psbJ-petA, ycf6-psbM-trnD) sequences using 27 samples from 13 species. On the basis of this phylogenetic analysis, we provide a solid phylogenetic background for the modern taxonomy of the European members of the genus. Traditionally accepted sections of this tree (i.e., Crypta and Elatinella) were found to be monophyletic; only E. borchoni—found to be a basal member of the genus—has to be excluded from the latter lineage to achieve monophyly. A number of taxonomic conclusions can also be drawn: E. hexandra, a high-ploid species, is most likely a stabilised hybrid between the main sections; E. campylosperma merits full species status based on both molecular and morphological evidence; E. gussonei is a more widespread and genetically diverse species with two main lineages; and the presence of the Asian E. ambigua in the European flora is questionable. The main lineages recovered in this analysis are also supported by a number of synapomorphic morphological characters as well as uniform chromosome counts. Based on all the evidence presented here, two new subsections within Elatinella are described: subsection Hydropipera consisting of the temperate species of the section, and subsection Macropodae including the Mediterranean species of the section.

Chromosome number of Elatine gussonei (Sommier) Brullo (Elatinaceae) and its distribution on the Maltese islands

Abstract Elatine gussonei (Sommier) Brullo is an endemic species, with a distribution restricted to the central part of the Mediterranean Basin (Maltese islands, Lampedusa, southern part of Sicily). This hydrophyte grows in rainwater pools and cavities in karstic limestone. Although the morphology has been well studied, no karyological study has been carried out, and hence this work brings the first chromosome data for the Maltese-pelago endemic E. gussonei.We have found a diploid number of 54 chromosomes in E. gussonei, which differs from the chromosome number of most of Elatine species (2n = 36). Additionally, this account gives a recent distribution of the species on the Maltese islands.

The DNA methylation level against the background of the genome size and t-heterochromatin content in some species of the genus Secale L

Methylation of cytosine in DNA is one of the most important epigenetic modifications in eukaryotes and plays a crucial role in the regulation of gene activity and the maintenance of genomic integrity. DNA methylation and other epigenetic mechanisms affect the development, differentiation or the response of plants to biotic and abiotic stress. This study compared the level of methylation of cytosines on a global (ELISA) and genomic scale (MSAP) between the species of the genus Secale. We analyzed whether the interspecific variation of cytosine methylation was associated with the size of the genome (C-value) and the content of telomeric heterochromatin. MSAP analysis showed that S. sylvestre was the most distinct species among the studied rye taxa; however, the results clearly indicated that these differences were not statistically significant. The total methylation level of the studied loci was very similar in all taxa and ranged from 60% in S. strictum ssp. africanum to 66% in S. cereale ssp. segetale, which confirmed the lack of significant differences in the sequence methylation pattern between the pairs of rye taxa. The level of global cytosine methylation in the DNA was not significantly associated with the content of t-heterochromatin and did not overlap with the existing taxonomic rye relationships. The highest content of 5-methylcytosine was found in S. cereale ssp. segetale (83%), while very low in S. strictum ssp. strictum (53%), which was significantly different from the methylation state of all taxa, except for S. sylvestre. The other studied taxa of rye had a similar level of methylated cytosine ranging from 66.42% (S. vavilovii) to 74.41% in (S. cereale ssp. afghanicum). The results obtained in this study are evidence that the percentage of methylated cytosine cannot be inferred solely based on the genome size or t-heterochromatin. This is a significantly more complex issue.

Reorganization of wheat and rye genomes in octoploid triticale (× Triticosecale)

AbstractMain conclusionThe analysis of early generations of triticale showed numerous rearrangements of the genome. Complexed transformation included loss of chromosomes, t-heterochromatin content changes and the emergence of retrotransposons in new locations. This study investigated certain aspects of genomic transformations in the early generations (F5 and F8) of the primary octoploid triticale derived from the cross of hexaploid wheat with the diploid rye. Most of the plants tested were hypoploid; among eliminated chromosomes were rye chromosomes 4R and 5R and variable number of wheat chromosomes. Wheat chromosomes were eliminated to a higher extent. The lower content of telomeric heterochromatin was also found in rye chromosomes in comparison with parental rye. Studying the location of selected retrotransposons from Ty1-copia and Ty3-gypsy families using fluorescence in situ hybridization revealed additional locations of these retrotransposons that were not present in chromosomes of parental species. ISSR, IRAP and REMAP analyses showed significant changes at the level of specific DNA nucleotide sequences. In most cases, the disappearance of certain types of bands was observed, less frequently new types of bands appeared, not present in parental species. This demonstrates the scale of genome rearrangement and, above all, the elimination of wheat and rye sequences, largely due to the reduction of chromosome number. With regard to the proportion of wheat to rye genome, the rye genome was more affected by the changes, thus this study was focused more on the rye genome. Observations suggest that genome reorganization is not finished in the F5 generation but is still ongoing in the F8 generation.

Possible ancient origin of heterochromatic JNK sequences in chromosomes 2R of Secale vavilovii Grossh.

Employing FISH analysis as well as BLAST and CUSTAL W (1.82) programs, we investigated types of DNA nucleotide sequences building an additional heterochromatic band in 2R chromosomes of 3 lines ofSecale vavilovii Grossh. The probes used in FISH analysis were designed based on the reverse transcriptase sequence of Ty1-copia and Ty3-gypsy retrotransposons and the 5S rRNA gene sequence. No hybridization signals from the reverse transcriptase probes were observed in the chromosome region where the additional band occurs. On the other hand, signals were observed after hybridization with the 5S rDNA probe, clearly suggesting the presence of that type of sequences in the analyzed heterochromatin band. Using BLAST and CUSTAL W programs, we revealed high similarity of the JNK1 sequence to the 5S rRNA gene fromHordeum chilense (HCH1016, HCH1018, 88%) and to a fragment of the 5S rRNA sequence ofH. marinum (HMAR003, 97%). In addition, the same fragment of JNK1 was shown to be very similar to the part of theAngela retrotransposon (92%) as well as to theSNAC 426K20-1 transposon (89%) belonging to CACTA family, both fromTriticum monococcum, and toZingeria biebersteiniana pericentromeric sequences (78%). The similarity of JNK1 to those sequences may be accidental or the JNK1 may represent an ancient mobile genetic element that caught the 5S rRNA sequence. During the evolution those sequences might have been accumulated in the particular region on the 2R chromosome. Our results suggest that the additional heterochromatin band in chromosomes 2R ofS. vavilovii is a collection of defective genes and/or mobile genetic elements.