Abnormally spliced messenger RNA in erythroid cells from patients with beta+ thalassemia and monkey cells expressing a cloned beta+-thalassemic gene.

The reduced beta-globin synthesis characterizing the beta+ thalassemia phenotype has been shown to be caused by anomalous processing within the small intervening sequence (IVS1) of the beta-globin mRNA precursor. The beta-globin gene from such patients contains a single base substitution within IVS1, located 22 bp from the 3' junction between IVS1 and exon 2, creating an alternative splice site within IVS1 and resulting in retention of the 3'-terminal 19 bases of IVS1. We have identified this abnormally spliced mRNA in the reticulocyte RNA of two patients with beta+ thalassemia, by S1 nuclease mapping and primer-extension analysis. Moreover, a cloned beta+-thalassemic gene preferentially generated the anomalously spliced RNA when expressed in monkey kidney cells. The anomalously spliced RNA constituted approximately 80%--90%, and normal beta RNA approximately 10%--20%, of the total beta mRNA. In contrast, the small amount of beta mRNA present in reticulocytes from such patients consisted predominantly of normal beta mRNA. These results suggest that the reduced amount of normally functioning beta mRNA present in such patients results from preferential processing at the alternative splice site, with subsequent instability, reduced nuclear processing and/or inadequate cytoplasmic transport of the abnormal RNA species.