Supplementary Figure 11: MiCEE is required for nucleolar structure
From: MiCEE is a ncRNA-protein complex that mediates epigenetic silencing and nucleolar organization
a, MLg cells were transfected with a control (Ctrl) or C1d-specific shRNA constructs. Transfected cells were analyzed by confocal microscopy after double immunostaining using FBL- and H3K27me3-specific antibodies. Arrows, perinucleolar region. DAPI, nuclear staining. Scale bars, 10µm. See Fig. 6a and S6g. b, MLg cells were treated with a control (Ctrl, DMSO) or EZH2 inhibitor (EZH2inh, UNC1999). Treated cell were analyzed by confocal microscopy after double immunostaining using FBL- and H3K27me3-specific antibodies. Arrows, perinucleolar region. DAPI, nuclear staining. Scale bars, 10µm. See Fig. 6a and S8g. c, Schematic representation of the Mirlet7d locus (left) explaining the strategy for depletion of the Mirlet7d using the CRISPR/Cas9 technology. Arrows show the direction of expression of Mirlet7. Arrowheads represent guide RNAs for Cas9 binding (magenta) and primers for screening of Mirlet7d -/- cells (blue). Representative results of genotyping of wildtype (+/+) and Mirlet7d depleted (-/-) MLg cells (middle). TaqMan assay based expression analysis of the indicated Mirlet7 family members in the wildtype (+/+) and Mirlet7d depleted (-/-) MLg cells (right). In a-c, representative results from 3 independent experiments (see Supplementary Data Sets 3 and 4)
