The Cross-Regulation Between Set1, Clr4, and Lsd1/2 in Schizosaccharomyces pombe
Fig 2
Genetic and physical interactions between Lsd1/2, CLRC, and COMPASS.
(A) The genetic interactions between lsd1-ΔHMG or lsd2ΔC with the loss of the methyltransferases Clr4 and Set1. (B) A schematic illustration of the key members of the CLRC and COMPASS complexes, highlighting their functional interactions with Lsd1/2. The dashed lines indicate the protein interactions that were detected by Western blot. (C-F) Protein-protein interactions between Lsd1 and Swd2 (C) or Raf2 (D), and between Lsd2 and Swd2 (E) or Raf2 (F) were revealed through co-immunoprecipitation followed by western blotting. +/- signs indicate the presence or absence of the indicated genetic background. IgG Sepharose beads were used for protein immunoprecipitation. WCE: whole cell extract. PAP was used to detect FTP signals. Asterisks mark the expected size for Lsd1-FTP (C-F), while arrows indicate the expected sizes for Swd2-GFP-HA (C & E), Raf2-myc (D & F). (G-H) ChIP-Seq peak enrichments for Lsd1 (G) and Lsd2 (H) with indicated genetic background are mapped to genomic regions spanning 500 base pairs upstream of TSS to 500 base pairs downstream of the TES. Aligned Lsd1 or Lsd2 occupancy values for all selected genes are plotted as moving averages along their relative genomic positions. The gene sets and genomic regions correspond to those depicted in Fig 1C. Color bars represent different genetic backgrounds, and Log2 fold enrichment indicates the Log2 fold change of Lsd1 and Lsd2 occupancy at specific genome loci compared to the untagged negative control. (I) Analysis of WCE from cells expressing untagged proteins and FTP-tagged Lsd1 and Lsd2 through SDS-PAGE and western blotting. PAP was used to detect Lsd1/2 protein levels. “Untagged” denotes cells with no tagged protein, while “WT” indicates cells expressing wild-type Lsd1/2-FTP. Mlo3 levels were used as the loading control. Asterisks signify the expected full-length size for Lsd1-FTP and Lsd2-FTP. The denotations for “Untagged” and “WT” remain consistent in the rest of the figures.