Fig. 7

Excess pronuclear localization of G9a impedes 5mC oxidation in paternal pronuclei. A Tet3 staining under TP conditions of control and DMOG-treated zygotes at the PN4-5 stage (10 hpf). B Quantification is represented as Tet3 signal intensity or a ratio between parental signals (pat./mat.). (n = 16 for control, n = 18 for iTet3.). C G9a staining of control, and DMOG-treated zygotes at the PN4-5 stage (10 hpf). D Quantification is represented as G9a signal intensity or a ratio (pat/mat). (n = 13 for control, n = 19 for iTet3). E Tet3 staining under TP conditions of control, and BIX-01294-treated zygotes at the PN4-5 stage (10 hpf). F Quantification is represented as Tet3 signal intensity or a ratio (pat./mat.) (n = 18 for control, n = 19 for iG9a.) G Competitive inhibition of G9a activity by BIX-01294 affects the accumulation of paternal 5hmC ERK1/2 inhibition (50 μM GDC-0994) produced as assessed by 5mC and 5hmC staining at the PN5 stage (11 hpf). H Quantification of 5mC and 5hmC is represented as signal intensity in paternal pronuclei. (n = 23 for control, n = 20 for iERK1/2, n = 17 for iERK1/2 + iG9a; n = 16 for iG9a.) I Tet3 inhibition by DMOG promotes the deposition of paternal H3K9me2 as assessed by H3K9me2 staining at PN4-5 stage (10hpf). J Quantification of the parental H3K9me2 signal intensity. Number of zygotes analysed for each group: n = 14 for control, n = 13 for iERK1/2, n = 19 for iERK1/2 + iTet3; n = 17 for iTet3. Statistical analysis was carried out using Student’s t-test (two-sided). P values are indicated. Error bars indicate SD. ♀, maternal pronucleus; ♂, paternal pronucleus. PB, polar body. Scale bar, 20 µm