2-Aminopurine Selectively Inhibits Splicing of Tumor Necrosis Factor Alpha mRNA

Abstract

2-Aminopurine (2-AP) inhibits specific kinases that phosphorylate the α subunit of eukaryotic translation initiation factor 2. One of these, PKR, is also involved in signal transduction. We show here that 2-AP selectively inhibits expression of tumor necrosis factor alpha (TNF-α) mRNA in primary human lymphoid cells. 2-AP does not inhibit transcription of the human TNF-α gene, nor does it affect mRNA stability. Instead, the flow of short-lived precursor transcripts into mature TNF-α mRNA is blocked. When 2-AP is present during induction, unspliced TNF-α precursor transcripts accumulate at the expense of mRNA. Using RNase protection analysis with genomic probes for different exon-intron junctions, we show that 2-AP blocks splicing of TNF-α mRNA. Neither the TNF-β nor the interleukin-1β gene shows such regulation. 2-AP also inhibits splicing of precursor RNA transcribed from an exogenous human TNF-α gene. Sequences within this gene thus confer sensitivity to 2-AP. Yet, control is not exerted at a specific splice site. Our results reveal the involvement of a 2-AP-sensitive component, expressed in functional form before induction, in the splicing of TNF-α mRNA.