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. Author manuscript; available in PMC: 2021 Oct 15.
Published in final edited form as: Mol Cell. 2020 Sep 22;80(2):345–358.e9. doi: 10.1016/j.molcel.2020.08.016

Figure 7. Function of INTS8 is conserved in human cells.

Figure 7.

(A) Western blot analysis of cell lysates derived from 293T cells transfected with control or INTS8 targeting siRNA. Lysates were harvested after 48 h and probed with antibodies against INTS8 or GAPDH as a loading control.

(B) 293T cells were transfected with control or INTS8 targeting siRNA. After 48 h, RNA was harvested for total RNA-seq (n=3). Normalized counts are shown for mRNA genes (N=12059). Affected genes are those with a fold-change > 2 and adjusted p-value <0.001.

(C) RNA-seq tracks are shown for ARC and GADD45B, genes upregulated upon INTS8 depletion.

(D) Schematic of the ARC and GADD45B reporter systems, wherein promoter and 5’ UTRs were cloned upstream of GFP. Top: In wild type cells, Integrator-PP2A drives Pol II termination and prevents GFP expression. Bottom: Upon loss of Integrator-PP2A association through INTS8 depletion or expression of INTS8-WFEF/A, Pol II productively elongates through the GFP ORF, yielding GFP expression.

(E) Western blot analysis of 293T cells transfected with the GFP reporters, along with control or IntS8 targeting siRNA, or IntS8 targeting siRNA plus RNAi-resistant INTS8-WT or INTS8-WFEF/A mutant cDNA.