FIG. 7.

UV cross-linking analysis of human and mouse mutants with HeLa nuclear extract. Each RNA was labeled with [α-³²P]UTP and then incubated with HeLa nuclear extract before being subjected to UV cross-linking and digestion with RNase. Samples were then run on an SDS-11% PAGE gel and exposed to BioMax autoradiographic film. The electrophoretic mobility of prestained molecular size markers (Broad Range; New England Biolabs) is shown on the left (A). IP was then performed with the same amount of each UV cross-linked sample shown in panel A with specific MAbs against different SR proteins: SF2/ASF (B, MAb 96), the phosphorylated RS domain (C, MAb 1H4), and SC35 (D, anti-SC35). The mobility of the SR proteins is indicated on the left. The SF2/ASF antibody immunoprecipitates more than one protein band owing to the presence of differently phosphorylated forms, as specified by the manufacturer.