The methylated H3 histone and heterochromatin protein 1 (HP1) are markers of heterochromatin in several eukaryotes possessing monocentric chromosomes. In order to confirm that these epigenetic markers of heterochromatin are evolutionary conserved, the distribution of methylated H3 histones and HP1 homologues on the holocentric chromosomes of the cabbage moth Mamestra brassicae (Lepidoptera) were studied. In particular, PCR experiments with degenerated primers identified a HP1 homologue (called MbHP1) in the M. brassicae genome. Sequencing showed that the MbHP1 gene is 737 bp long including a 102 bp 5'UTR and a 635 bp coding portion (comprising an 80 bp intron). The MbHP1 peptide consisted of 184 amino acids, had a 20 kDa molecular mass and a net negative charge. At the structural level, it showed an N terminal chromo-domain and a chromo-shadow-domain at the C terminus linked by a short hinge region. At the cytogenetic level, MbHP1 was located exclusively in the heterochromatic regions of the chromosomes. The same heterochromatic regions became labelled after immuno-staining with antibodies against H3 histone methylated at lysine 9, reinforcing the hypothesis that this modified histone is essential for HP1 binding. Our data, as a whole, confirm that heterochromatic components and markers are evolutionary conserved both in mono- and holocentric chromosomes despite the difference in the distribution of heterochromatin on chromosomes.
The structure of the holocentric chromosomes of the rosy apple aphid, Dysaphis plantaginea (2n = 12), and pear-grass aphid, Melanaphis pyraria (2n = 8), was studied using C-banding, NOR, Giemsa and fluorochrome staining, and fluorescent in situ hybridization (FISH). Contrary to the equilocal distribution of heterochromatin typical of monocentric chromosomes, in both species C-banding evidenced a tendency of highly repetitive DNAs to be restricted to the X chromosomes. Silver staining and FISH, using a 28S rDNA probe, located rDNA genes on one telomere of each X chromosome, the only brightly fluorescent C-positive sites revealed by CMA3 staining, whereas all other heterochromatic C-bands were DAPI positive. Both species showed a noticeable amount of rDNA heteromorphism. Mitotic recombination is proposed as a possible mechanism responsible for the variation in size of rDNA.