Figure 4.

Interaction among the subunits. (A) Coexpression of the four conserved subunits. Expression of the four subunits, Rpb1H, Rpb2, GST–Rpb3 and Rpb11, were detected by western blotting of the lysate with antisera against each subunit. The coexpressed proteins are indicated on the top: G, GST; G3, GST–Rpb3; 1H, Rpb1H; 2, Rpb2; 11, Rpb11, and the detected subunits are indicated on the left. As a control, GST was expressed in place of GST–Rpb3 (lane 1). Portions of the filters corresponding to the migration positions of each subunit in the SDS–PAGE are arrayed. (B) Analysis of the assemblies of four subunits. The extracts of the coinfected cells in various combinations were subjected to GSH-affinity chromatography and the eluted fractions were analyzed by western blotting as in (A). (C) Coexpression of the five small subunits. Expression of the five subunits, GST–Rpb3, Rpb5, Rpb7, Rpb8 and Rpb11, in the coinfected cells was detected by western blotting with antisera against each subunit. The proteins coexpressed are indicated on the top: GUS, β-glucuronidase as a control for antisera cross-reaction; G, GST; G3, GST–Rpb3; 5, Rpb5; 7, Rpb7; 8, Rpb8; 11, Rpb11, and the detected subunits are indicated on the left. Portions of the filters are arrayed as in (A). (D) Analysis of the assemblies of five small subunits. The extracts of the coinfected cells were subjected to GSH-affinity chromatography and the eluted fractions were analyzed by western blotting as in (C).