Temperaturedependent splicing of b-globin pre-mRNA

A T®G mutation at nucleotide 705 of human b-globin intron 2 creates an aberrant 5 ¢ splice site and activates a cryptic 3 ¢ splice site upstream. In consequence, the pre-mRNA is spliced via aberrant splice sites, despite the presence of the still func-tional correct sites. Surprisingly, when IVS2-705 HeLa or K562 cells were cultured at temperatures below 30°C, aberrant splicing was inhibited and correct splicing was restored. Similar temperature effects were seen for another b-globin pre-mRNA, IVS2-745, and in a construct in which a b-globin intron was inserted into a coding sequence of EGFP. Temperature-induced alternative splicing was affected by the nature of the internal aberrant splice sites ¯anking the correct sites and by exonic sequences. The results indicate that in the context of thalassemic splicing mutations and possibly in other alternatively spliced pre-mRNAs, temperature is one of the parameters that affect splice site selection