Fig. 1

CGGBP1 regulates cytosine methylation in a GC-content and cytosine context dependent manner. a Chromosome-wise distribution of cytosines exhibiting methylation changes (GoM and LoM) or no methylation changes (RoM and RuN). The similar distribution of the four methylation change states on all the chromosomes showed no major chromosomal preference for CGGBP1-regulation of cytosine methylation. b, c Measurement of intra-chromosomal variabilities in methylation states in S1 and S2 shows preference for GoM in G-rich R-bands (Giemsa-negative) and LoM in G-bands (Giemsa positive 100). In R-bands, the paired GoM and LoM events had closely related values like RoM and RuN events but GoM was significantly more than LoM (paired t-test p value = 2.239e−012) (b). However, at G-bands the methylation state was reversed and LoM was higher than GoM (p value = 0) (c). Similarly, while RuN was significantly higher in R-bands (paired t-test p value = 0) (b), RoM was higher in G-bands (paired t-test p value = 0.000636). For paired t-test, n = 780 R bands in B and n = 81 G bands in c. d–k The methylated and unmethylated fractions of cytosines in all three contexts have differential susceptibility to methylation change in absence of CGGBP1 function. d–f The context-wise distribution of methylated cytosines in S1 (d) underwent LoM wherein the proportion of CpG was lower and that of CHG and CHH were more than expected (e). From the same pool of cytosines methylated in S1 (d) that retained methylation upon CGGBP1 loss-of-function had a highly enriched CpG fraction and lower CHG and CHH fractions (f). g–i The context-wise distribution of unmethylated cytosines in S1 (g) that underwent GoM (h) also displayed an unexpected and disproportionate increase in CpG context. The context distribution amongst the cytosines that remained unmethylated upon CGGBP1 loss-of-function displayed a reduction in CpG context (i). A comparison of d and g clearly shows that the major fraction of CpG context was methylated in presence of CGGBP1 whereas CHH and CHG together comprised the most of unmethylated fractions. Comparison of e and h show that despite differences in the absolute numbers as well as relative abundance of the three contexts in methylated and unmethylated pools in S1 (d, g), the GoM (e) and LoM (h) cytosines were unexpectedly similar in magnitude with near identical context composition. If LoM and GoM were occurring randomly in the methylated and unmethylated pools of cytosines, then the magnitude and context distributions observed in d and g were proportionately expected in e and h respectively. Obs/Exp analyses of e and h against d and g revealed a highly significant unexpected composition of e and h (refer to Additional file 1: Table S6). j Plotting of the number of cytosines sequenced in S1 that underwent methylation change upon CGGBP1 depletion shows a disproportionate change in methylation states such that the GoM and LoM are quantitatively coincidental. k Conversely to j, the number of cytosines sequenced in S1 that resisted methylation change upon CGGBP1 depletion are disproportionately different and non-coincidental. All graphs are generated using GraphPad Prism7