Figure 2. NusG stimulates intrinsic termination via its NGN domain.

(A) IGV screenshot of a genomic window centered around the yetJ terminator. Top track is the 3’ end identified by Term-seq. Bottom tracks are the RNA-seq coverage data for the nusA_dep ΔnusG (-), ΔnusG (+NusA), nusA_dep (+NusG), and wild-type (WT) (+NusA +NusG) strains. %T in each strain is shown on the right of each track. Transcription proceeds from left to right. (B) YetJ terminator showing the point of termination identified by Term-seq in vivo (POT_Term-seq). Disruptions in the U-rich tract are shown in red. The upstream A tract is also shown. (C) Single-round in vitro termination assay with the yetJ terminator. Experiments were performed in the absence (–) or presence of NusA (A), NusG (G), WT NGN domain, and/or mutant NGN domain as indicated (mutant NGN domain is signified as NGN**). Positions of terminated (POT) and run-off (RO) transcripts are marked. The arrowhead marks the most distal POT. %T is shown below each lane. (D–F) Identical to panels (A–C) except that it is the ktrD terminator.

Figure 2—figure supplement 1. Template design for in vitro transcription.

(A) Schematic representation of templates used for all in vitro transcription experiments except for those involving convergent transcription. Template features are described below. (B) Schematic representation of templates used for in vitro transcription experiments that involve convergent transcription. Template features are described below including locations of restriction sites and primer annealing for the generation of templates.