Supplementary Figure 2: Gdown1 interaction sites overlap with TFIIF and TFIIB contact sites on Pol II.
From: Architecture of Pol II(G) and molecular mechanism of transcription regulation by Gdown1
a, Position of the TFIIF dimerization domain relative to Gdown1 density. K345 (shown in green) in the RPB2 lobe domain was cross-linked to K282 in the Gdown1 C-terminal region. b, Position of the TFIIB B-core domain relative to Gdown1 density. K427 (shown in green) in the RPB2 protrusion domain was cross-linked to the Gdown1 C-terminal region. c, Gdown1 C-terminal region interaction sites around the RPB1 clamp loops. Cross-linked lysines in the RPB1 clamp (K213) and rudder (K331) domains are shown in green. TFIIB is shown in yellow ribbon. d, Pol II binding assay. 35S-labeled TFIIB was incubated with Pol II or Pol II(G) and subjected to immunoprecipitation with anti-CTD antibody (8W16). Bound proteins were analyzed by SDS-PAGE and autoradiography. e, In vitro transcription assay with premelted promoter template. In vitro transcription assay was performed as described in Fig. 3d, except that the reaction time was reduced to 5 min. Note that the use of a premelted template bypasses the normal absolute TFIIF requirement for transcription, although transcription is still enhanced by TFIIF. The bands represent short transcripts resulting from specific initiation followed by Pol II pausing and/or premature termination.
