26% or 19% of cuts (in apoptotic and MNase degradation, respectively) were located at intergenic sequences farther away from genes. (ii) In both cases, a preference was also found for interspersed repeats mainly from the CR1 families; while these repeats obviously could not be mapped, they are known to be preferentially located in GC-rich genome regions [15,16]. (iii) ‘‘Redundant’’ clones, namely clones mapping at identical chromosomal sites, were present in both cases, but they were more than twice as abundant when derived from apoptosis than from MNase (see Figures 3 and 4 and Table 2); incidentally, since ‘‘redundant’’ clones from repeated sequences could not be mapped, overall redundancies were underestimated. (iv) In the case of microchrosomes several of them (14, 19, 21, 25) were only cut by apoptosis, other ones (15, 22, 24, 26, 27) only by MNase (see Figure S2). (v) Minimal distances between cuts and their nearest neighbors ranged from 0.65 kb to 26,000 kb (with an average of 4,600 kb) in the case of apoptosis and from 1.80 kb to 36,000 kb (with an average of 5,700 kb) in that of MNase; in the first case, distances lower than 4 Mb were found for 65.4% of all fragments, vs only 47.6% in the second (see Figure 4). Interestingly minimal distances among autolytic cuts and MNase cuts ranged from 26 kb to 23,800 kb (with an average of 2,900 kb; and 77.6% of distances below 4 Mb; see Figure 4 and Table 2). The Apoptotic Attack on Chromatin distributions of distances in the 0–500 kb range are also shown in Figure 4, the short 50–100 kb range being more represented in the distances among different cuts (6.5%) than in the distances within the ‘‘same’’ cuts (1.7% for apoptotic or 2.8% MNase cuts). An interesting situation was the one in which the two cuts were within 100 kb (see Figure 4 and Table 2). A shown in Figure 5, out of ten such cases, four (1 to 4) were characterized by cuts in the same genes (in one of them, cuts were just outside the gene sequence which is known to lack the 59 end); two (5 and 6) had one cut in one gene and another cut in another gene (again in number 6 one cut was just outside the end of the gene), two (7 and 8) had a cut in one gene only, and two (9 and 10) had two cuts outside genes. In all likelihood, the two cuts concerned the same domain (or loop) of chromatin, being less than 50 Kb (in six cases) and less than 100 Kb (in four cases) apart. Sequence specificity of apoptotic endonucleases and MNase The preferential 59 and 39 terminal dinucleotides of the clones obtained from apoptotic degradation were CT/AG, GG/CC and TG/CA; Figure S3A), without any preference for purines or pyrimidines (Figure S3C). Figure 3. Mapping of apoptotic and MNase cuts on chicken chromosome 1. The GC level and the gene density in the isochores are shown for the telomeric 60 Mb of the p arm of chicken chromosome 3. The arrows indicate the cuts (in red for the autolysis and in black for MNase digestion). The down- and up- pointed arrows Q q indicate cuts in genes or in sequences within 50 kb (asterisks) or 100 kb from genes. Cut mapping on all other chromosomes is reported in Figure S1. doi:10.1371/journal.pone.0005010.g003 PLoS ONE | www.plosone.org 3 April 2009 | Volume 4 | Issue 4 | e5010