Condensation patterns of prophase/prometaphase chromosome are correlated with H4K5 histone acetylation and genomic DNA contents in plants

Abstract

Mitotic prophase chromosome condensation plays an essential role in nuclear division being therefore regulated by highly conserved mechanisms. However, degrees of chromatin condensation in prophase-prometaphase cells may vary along the chromosomes resulting in specific condensation patterns. We examined different condensation patterns (CPs) of prophase and prometaphase chromosomes and investigated their relationship with genome size and distribution of histone H4 acetylated at lysine 5 (H4K5ac) in 17 plant species. Our results showed that most species with small genomes (2C < 5 pg) (Arachis pusilla, Bixa orellana, Costus spiralis, Eleutherine bulbosa, Indigofera campestris, Phaseolus lunatus, P. vulgaris, Poncirus trifoliata, and Solanum lycopersicum) displayed prophase chromosomes with late condensing terminal regions that were highly enriched in H4K5ac, and early condensing regions with apparently non-acetylated proximal chromatin. The species with large genomes (Allium cepa, Callisia repens, Araucaria angustifolia and Nothoscordum pulchellum) displayed uniformly condensed and acetylated prophase/prometaphase chromosomes. Three species with small genomes (Eleocharis geniculata, Rhynchospora pubera, and R. tenuis) displayed CP and H4K5ac labeling patterns similar to species with large genomes, whereas a forth species (Emilia sonchifolia) exhibited a gradual chromosome labeling, being more acetylated in the terminal regions and less acetylated in the proximal ones. The nucleolus organizer chromatin was the only chromosomal region that in prometaphase or metaphase could be hyperacetylated, hypoacetylated or non-acetylated, depending on the species. Our data indicate that the CP of a plant chromosome complement is influenced but not exclusively determined by nuclear and chromosomal DNA contents, whereas the CP of individual chromosomes is clearly correlated with H4K5ac distribution.

Fig 1. Distribution of H4K5ac in chromosomes of species with low DNA content.

a, Indigofera campestris; b, Phaseolus lunatus; c, P. vulgaris; d, Solanum lycopersicum. Images display, respectively, Giemsa (black and white), DAPI (gray), FITC-conjugated anti-H4K5ac (green), DAPI/FITC (gray/green), and CMA/DAPI (yellow/gray) staining. Insets show magnified chromosomes displaying high and low condensed regions (a1-d1), and CMA⁺ NORs unlabeled with anti-H4K5ac (b4, b5). Arrows in c and d point to hyperacetylated CMA⁺ NORs. Insets in d2 and d3 show magnified chromosome (left arrow). Arrowheads in c3 and c4 indicate hypoacetylated CMA⁺ chromocenters. Scale bar in a1 corresponds to 10 μm.

Fig 2. Distribution of H4K5ac in chromosomes of species with low DNA content.

a, Poncirus trifoliata; b, Arachis pusilla; c, Bixa orellana. Images display, respectively, Giemsa (black and white), DAPI (gray), FITC-conjugated anti-H4K5ac (green), DAPI/FITC (gray/green), CMA (yellow), and CMA/DAPI (yellow/gray) staining. Inset in a1 shows a chromosome with a proximal early condensing chromatin and a terminal heterochromatic block. Arrowheads in b point to the weakly stained A chromosome pair. Asterisks in a indicate chromosomes amplified in a4. Arrows in c point to the heterochromatic arm of chromosome pair 1 in prophase nuclei. Asterisks in c indicate the hypoacetylated CMA⁺ regions around the secondary constriction. Scale bar in a1 corresponds to 10 μm.

Fig 3

Distribution of H4K5ac in chromosomes of Emilia sonchifolia (a), Rhynchospora tenuis (b), R. pubera (c), and Eleocharis geniculata (d). Images display, respectively, Giemsa (black and white), DAPI (gray), FITC-conjugated anti-H4K5ac (green), DAPI/FITC (gray/green), and CMA/DAPI (yellow/gray) staining. Arrows in a5 and d2 point to NORs. Scale bar in a1 corresponds to 10 μm.

Fig 4. Distribution of H4K5ac in chromosomes of species with high DNA content.

a, Allium cepa; b, Callisia repens; c, Nothoscordum pulchellum; d, Araucaria angustifolia. Images display, respectively, Giemsa (black and white), DAPI (gray), FITC-conjugated anti-H4K5ac (green), and CMA/DAPI (yellow/gray) staining. Arrows point to the NORs. Scale bar in a1 corresponds to 10 μm.