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doi: 10.1242/10.1242/jcs.00172

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Differential acetylation of histones H3 and H4 in paternal and maternal germline chromosomes during development of sciarid flies

Departamento de Biología Celular y del Desarrollo, Centro de Investigaciones Biológicas, CSIC, 28006 Madrid, Spain

View larger version (28K): [in a new window]   Fig. 1. Scheme summarizing the most relevant chromosomal events of germline nuclei at the resting stage and male meiosis in sciarids. Maternally derived chromosomes are represented in blue and paternally derived chromosomes are represented in red. Early embryonic germ nuclei contain three maternal and paternal autosomes (A) plus one maternal and two paternal X-chromosomes (X). E refers to the occurrence of chromosome elimination. Note that the parental origin of meiotic chromosome products in the sperm nucleus and egg are represented as they will be recognized after fertilization (paternal and maternal, respectively).

View larger version (84K): [in a new window]   Fig. 2. Distribution of acetylated histone H4 at lysine 8 in S. ocellaris and S. coprophila embryonic germ nuclei at the resting stage. (A-D) Chromosome DAPI staining, (A'-D') Indirect immunolabeling with H4Ac8 antibody and (A''-D'') superimposed images. where the antibody staining is in red. (A) S. ocellaris germ nuclei prior to, (B) during and (C) after the occurrence of Xp chromosome elimination. (D) A S. coprophila nucleus before the elimination of Xp and L chromosomes. In all cases only four chromosomes of the complement are highly H4Ac8-labelled (A'-D' and A''-D''). (B-B'') a S. ocellaris nucleus undergoing elimination where it is seen that the Xp chromosome expelled from the nucleus (asterisk in B) is not stained (asterisks in B' and B''); a labeled somatic nucleus is also shown for comparison (arrows). (C'-C'') A post-elimination S. ocellaris nucleus where it is possible to observe that only one of the chromosomes corresponding to the longest autosomes pair is labeled (arrows). (D-D'') The four S. coprophila chromosomes that are H4Ac8-labelled (short arrows in D') correspond to the ones that are slightly less stained with DAPI (short arrows in D); the H4Ac8-unstained eight chromosomes contain three L chromosomes (possibly the ones indicated by long arrows in D''). Bar, 10 µm.

View larger version (48K): [in a new window]   Fig. 3. Colocalization of acetylated histone H3 and acetylated histone H4 at lysines 8 and 12 in S. ocellaris embryonic germ nuclei. (A-D) DAPI staining, (A'-D') indirect immunolabeling with different antibody combinations that are specified in the top of the figure and (A''-D'') superimposed images where antibody staining is in red. In all cases the antibodies labeling is restricted to the same four chromosomes of the complement. Bar, 10 µm.

View larger version (76K): [in a new window]   Fig. 4. Distribution of acetylated histone H4 at lysine 8 in germ cells of S. ocellaris male larvae at one day (1d), five days (5d) and 10 days (10d) from hatching. (A-C) DAPI staining, (A'-C') indirect immunolabeling with H4Ac8 antibody and (A''-C'') superimposed images where antibody staining is in red. (A) Maternal chromosomes appear condensed (long arrows) whereas paternal chromosomes, less stained with DAPI, initiate decondensation (short arrows); (A',A'') H4Ac8-staining is restricted to the paternal chromosome set. (B) Maternal chromosomes remain condensed (long arrows) whereas paternal ones appear totally decondensed (short arrows); (B',B'') H4Ac8-labeling is restricted to the chromatin corresponding to the paternal chromosome set. (C) All chromosomes appear equally condensed and exhibit H4Ac8-staining (C',C''). Bar, 10 µm.

View larger version (71K): [in a new window]   Fig. 5. Distribution of acetylated histone H4 at lysine 8 in germ cells of S. coprophila male larvae at 3 days (3d) and 10 days (10d) from hatching. (A,B) DAPI staining, (A',B') indirect immunolabeling with H4Ac8 antibody and (A'',B'') superimposed images where antibody staining is in red. (A) Maternal chromosomes (long arrows) and L chromosomes (arrowheads) appear condensed and clustered whereas paternal chromosomes are fully decondensed (short arrows); (A',A'') Paternal chromatin exhibits H4Ac8-staining whereas maternal and L chromosomes are devoid of staining. (B-B'') All chromosomes are condensed and show H4Ac8-staining except the L chromosomes (arrowheads in B and B''). Bar, 10 µm.

View larger version (62K): [in a new window]   Fig. 6. Distribution of acetylated histones in S. ocellaris (A-C) and S. coprophila (D) spermatocytes undergoing paternal chromosomes elimination at first meiotic division. (A-D) DAPI staining of anaphase-like stage figures where m refers to the maternal group of chromosomes, p refers to paternal chromosomes and * marks the position of the single spindle pole. (A'-D') Indirect immunolabeling with the antibody specified in the top of the figure and (A''-D''') superimposed images where antibody staining is in red. In all cases (A''-D''), the antibody labeling is restricted to the four maternal chromosomes that face the pole whereas the opposed paternal group of chromosomes is devoid of staining. Arrows in D indicate S. coprophila L chromosomes that migrate together with the maternal group of chromosomes and do not exhibit antibody labeling (D'-D''). Bar, 10 µm.

View larger version (98K): [in a new window]   Fig. 7. Distribution of histone H4 acetylation at lysine 12 during S. ocellaris spermiogenesis (A-C) and in an embryo at early fertilization stage (D). DAPI staining of elongated spermatids (A), late spermatids (B), mature spermatozoa (C) and the whole embryo (D). (A',B',D') Indirect immunolabeling with H4Ac12 antibody and (A'',B'',D'') superimposed images where antibody staining is in red. In A',A'' H4Ac12 distributes on the nuclear chromatin whereas in B',B'' the labeling is restricted to a few nuclear sites. In C no labeling is observed. In D the short arrow points to the sperm nucleus and long arrows to the haploid maternal meiotic products. (D',D'') Enlarged images of the sperm nucleus and maternal chromosomes where it is seen that H4Ac12-labelling localizes to the decondensing sperm nucleus chromatin and not to the maternal chromatin. Bar, 10 µm.

View larger version (93K): [in a new window]   Fig. 8. Distribution of acetylated histones in Sciara embryos at first somatic division. DAPI staining of S. ocellaris first metaphase chromosomes (A-C) and of S. coprophila first anaphase chromosomes (D). (A'-D') Indirect immunolabeling with the antibodies specified in the top of the figure. (A''-D'') Superimposed images where antibody staining is in red. Bar, 5 µm.

View larger version (27K): [in a new window]   Fig. 9. Scheme summarizing the distribution of acetylated histones H4 (Lys8 and Lys12) and H3 (Lys9 and Lys14) in early germline nuclei and male meiosis of S. ocellaris. Maternally derived chromosomes are represented in blue and paternally derived chromosomes are represented in red. Chromosomes containing highly acetylated histones H4 and H3 are marked with yellow color, whereas chromosomes containing under-acetylated histones H4 and H3 are unmarked. E refers to chromosomal elimination events.