FIG. 1.

Preferential binding of Pnn to spliced mRNA in vitro. (A) Immunoprecipitation was performed with ³⁵S-labeled HeLa cell lysate using anti-mouse immunoglobulin G (lane 1, mock) or anti-Pnn antibodies (lane 2). Anti-Pnn antibodies specifically precipitated a protein of about 130 kDa (arrowhead). Protein size markers in kilodaltons are indicated at left. (B) In vitro splicing reaction was carried out at 30°C for 45 min with PIP85A (lanes 1 to 3) or its intron-lacking version (Δi RNA; lanes 4 to 6) as substrate. Immunoprecipitation was then performed with mock (lanes 2 and 5) or anti-Pnn (lanes 3 and 6) antibodies. Lanes 1 and 4 show 10% of the splicing reaction mixture used in immunoprecipitation. The Δi RNA (arrowhead) contains an extra 83-nt sequence derived from the vector at its 5′ end, thus being larger than the spliced mRNA. Size markers are radiolabeled MspI-digested pUC19 as indicated at left.