Fig. 7
From: Promoter-proximal pausing mediated by the exon junction complex regulates splicing
The role of Mago on promoter proximal pausing is conserved in human. a Metagene profiles based on averaged enrichment over input from two independent biological replicates of total Pol II occupancies in control and Magoh-depleted HeLa cells with standard error of the mean for all the Pol II bound genes. Log2 fold changes against input control are shown on Y-axis while X-axis depicts scaled genomic coordinates. b Metagene profiles of averaged enrichment from two independent biological replicates of total Pol II occupancies in control and Magoh-depleted HeLa cells with standard error of mean for all the Pol II bound genes, centered at the TSS in a ±1 Kb window. Log2 fold changes against input control are shown on Y-axis while X-axis depicts genomic coordinates. c The ECDF plot of PRR in HeLa cells treated with either control or Magoh siRNA. p-value is derived from two-sample Kolmogorov–Smirnov test. d Track examples of total Pol II ChIP-Seq from HeLa cells extracts, after either control or Magoh knockdown. The tracks are the average of two independent biological replicates after input normalization. e Western blots performed using antibodies against total Pol II, Ser2P, Ser5P, and Magoh, in HeLa cells treated with either control or Magoh siRNA. Similar to Drosophila, the loss of Magoh leads to elevated level of Ser2P without affecting Ser5P. f–h Co-immunoprecipitation of Magoh from HeLa cells extracts, using antibody directed against Magoh. Western blots using Pol II, Ser5P and Ser2P antibody reveal RNA-dependent interaction of Magoh with Pol II and Ser5P. There was no detectable interaction of Magoh with Ser2P. The specific bands for Pol II are highlighted by arrowheads in f. The lane labeled with “Lad” indicates ladder. i Co-immunoprecipitation of Cdk9 using anti-Cdk9 antibody from HeLa cells extracts treated with either control or Magoh siRNA. Western blot was performed with anti-Ser5P antibody. The quantification of the intensity was performed with ImageJ. j Model: The pre-EJC stabilizes Pol II pausing by restricting P-TEFb binding at promoter, and possibly by sequestering Cdk12. This activity is required for proper recognition of exons
