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. 2017 Nov;23(11):1700–1711. doi: 10.1261/rna.061291.117

FIGURE 4.

FIGURE 4.

The HAdV-37 MLP-TSS-sRNAs are products of premature termination of RNA polymerase II transcription. (A) Schematic diagram of the different constructs used for the transfection assay. UnMLT contains the MLP fused to the first 200 nt downstream from the transcriptional start site, Trip contains the MLP fused to a cDNA copy of the spliced tripartite leader sequence, and RevTrip contains a cDNA copy of the tripartite leader where the second and third exons had been inserted in the reverse orientation. (B) Northern blot analysis of total cytoplasmic RNA isolated 24 h post transfection with the indicated plasmids. The same membrane was sequentially probed for the MLP-TSS-sRNA and tRNA lysine. (C) Northern blot analysis of total (Input), RNAPII-associated small RNA (RNAPII-IP), and Ago2-associated (Ago2-IP) RNA isolated from HAdV-37 infected 293-Flag-Ago2 cells at 30 hpi. The membrane was probed sequentially for the MLP-TSS-sRNA, 5′ mivaRNAII and tRNA lysine. (D) Western blot analysis of RNAPII, Ago2, and Actin in the fractions displayed in panel C.