Identification of proteins binding specifically to the 3′-untranslated region of granulocyte/macrophagecolony stimulating factor mRNA

The 3′-untranslated region of granulocyte/macrophage colony-stimulating factor (GM-CSF) mRNA contributes to the post-transcriptional regulation of gene expression. Degradation is partly mediated by adenosine-uridine-rich sequence elements (ARE), which serve as binding sites for specific proteins. Stabilization of RNA by phytohemagglutinin and concanavalin A treatment is dependent on regulatory sequence elements upstream of ARE. We have performed northwestern blot and filter binding assays using cell extracts and RNA sequences containing or lacking ARE. Murine and human T cell extracts (EL-4 and Jurkat) yielded two specific proteins of 93 and 94 kDa, respectively, that were binding to sequences upstream of ARE. Within this region, the human and murine RNA do not share any obvious sequence identity, yet both are target sites for the binding proteins. The smallest RNA fragments protected by the proteins from RNase A digestion, were 44 in the murine, and 38 ribonucleotides long in the human sequence. The binding activity of the 94 kDa protein derived from human Jurkat cells could be enhanced by phytohemagglutinin. The interaction with regulatory mRNA sequences and the responsiveness to phytohemagglutinin suggests that the proteins are involved in controlling GM-CSF mRNA turnove