Functional repetitive sequences and fragile sites in chromosomes of Lolium perenne L.

Abstract

Lolium perenne is considered a high-quality forage widely used in temperate regions to meet the shortage of forage during the winter. In this species, some peculiarities related to cytogenetic aspects have already been described, as the variability in number and position of 45S ribosomal DNA (rDNA) sites and the expression of fragile sites, which require further studies to support the understanding of their causes and consequences. In this way, this study aimed to evaluate the relationship between the expression of fragile sites and functional repetitive sequences (rDNA and telomeric) in chromosomes of diploid and polyploid cultivars of L. perenne. The techniques of FISH, Ag-NOR and fluorescence banding were used to assess the distribution of sites of 45S rDNA, 5S, telomeric sequences, and the transcriptional activity of the 45S ribosomal genes and the distribution of AT- and/or GC-rich sequences in L. perenne, respectively. There was variability in the number and location of 45S rDNA sites, which was not observed for 5S rDNA sites. One of the genotypes showed two 45S rDNA sites on the same chromosome, located in different chromosome arms. Breaks and gaps were found in 45S rDNA sites in most metaphases evaluated for both cultivars. Telomeric sequences were not detected at the end of the chromosomal fragments corresponding to the location of breaks at 45S sites. Apparently, the transcriptional activity was modified in fragile sites. Variation in the number and size of nucleoli, nucleolar fusions and dissociations were observed. All CMA(+) bands were colocalized with the 45S sites.

Dynamics of 45S rDNA sites in the cell cycle: fragile sites and chromosomal stability in Lolium and Festuca

“…This suggests that there is no loss of chromosome fragments in these genotypes. Rocha et al (2016) suggested that the chromatin fiber in FSs of Lolium, Festuca, and Festulolium hybrids have a high decondensation process related to an increased genic activity, thus, supporting the findings by Huang et al (2012), Bustamante et al (2014), and Rocha et al (2015).…”

“…In interphasic G1 nuclei, the number of 45S signals measured was similar to the descriptions for mitotic chromosomes with six, seven, and four/five sites for L. multiflorum, L. perenne, and F. arundinacea, respectively (Thomas et al, 1996(Thomas et al, , 1997Bustamante et al, 2014;Rocha et al, 2015). In S/G2 nuclei, the number of 45S signals was higher for the three species studied here owing to the occurrence of FSs.…”

“…LOL 161) and L. perenne ('Ellet'), both 2n = 2x = 14 (Pereira et al, 2014;Rocha et al, 2015), in addition to two F. arundinacea cultivars ('Rizomat' and 'Quantum II') 2n = 4x = 42. The Lolium species came from Banco Ativo de Germoplasma de Plantas Forrageiras (Active Germplasm Bank) and Embrapa Recursos Genéticos e Biotecnologia.…”

“…and Lolium perenne L., FSs are randomly distributed in 45S rDNA sites and promote changes in the number and position of fluorescence in situ hybridization (FISH) signals. The predominant number of described 45S rDNA sites is six and seven for L. multiflorum and L. perenne, respectively (Thomas et al, 1996;Rocha et al, 2015). However, in C-metaphases with FSs, the number of 45S FISH signals range from 6 to 13 in L. multiflorum and from 7 to 13 in L. perenne.…”

“…In intact chromosomes, invariably, the signals are located at interstitial chromosome positions, whereas in chromosomes with FSs, the variable-sized signals are located in the terminal regions. Initially, it was proposed that the occurrence of breakages or gaps in these regions resulted in the formation of chromosomal fragments (Huang et al, 2008;Bustamante et al, 2014;Rocha et al, 2015).…”