Hypoxia increases ins-PRS-protein binding activity. The 32P-labelled insulin mRNA-oligonucleotide, ins-PRS, was incubated with rat islet extract, from islets incubated for 1 hr in normoxic or hypoxic (5% O2) 2.8 mM glucose-containing medium. The reactions were performed with (11 mM) or without DTT. In (A), the samples were cross-linked by exposure to UV radiation (260 nm) for 10 min and then analyzed by boiling in SDS-sample buffer without beta-mercaptoethanol and separation on a SDS-PAGE gel for 1 hr at 160 V. In (B), the samples were directly separated with nondenaturating polyacrylamide gel electrophoresis. Results shown are representative for three separate observations.