Figure 5

Relationship between ZMYM3 and LSD1 and epigenetic modifications. (a–b) LSD1 is expressed in c-KIT⁺ cells at a higher level than in GFRα1⁺ cells. Whole-mount co-immunostainings of LSD1 with GFRα1 and c-KIT were conducted on seminiferous tubules from P6 and P8 mice, respectively. (c) Co-immunostaining of LSD1 with GFRα1 in cultured SSCs. Note the inverse relationship in the signals of LSD1 and GFRα1. The signal of LSD1 is stronger in SSCs which are weaker in GFRα1 staining (arrow heads) but weaker in SSCs which are stronger in GFRα1staining (arrows). (d) KO of Lsd1 in cultured inducible Cas9-SSCs with Doxycyclin suppresses the proliferation of SSCs (n =3). (e) The larger isoform but not the smaller one of ZMYM3 co-immunoprecipitates with LSD1 using an antibody against LSD1. (f–g) Western blot of LSD1 expression in WT and KO SSCs (n=3). (g) is the quantitative densitometry of the Western blot result in (f). (h–i) Western blot of H3K4me1/2 and H3K9me2 in WT and KO SSCs (n=3). (i) is the quantitative densitometry of the Western blot result in (h). (j) Immunostaining analysis of H3K4me2 and H3K9me2 in spermatocyte spreads of WT and KO mice