INTRONS ARE CIS EFFECTORS OF THE NONSENSE-CODON-MEDIATED REDUCTION INNUCLEAR MESSENGER-RNA ABUNDANCE

The translation of human triosephosphate isomerase (TPI) mRNA normally terminates at codon 249 within exon 7, the final exon. Frameshift and nonsense mutations of the type that cause translation to terminate pr ematurely at or upstream of codon 189 within exon 6 reduce the level o f nuclear TPI mRNA to 20 to 30% of normal by a mechanism that is not a function of the distance of the nonsense codon from either the transl ation initiation or termination codon. In contrast, frameshift and non sense mutations of another type that cause translation to terminate pr ematurely at or downstream of codon 208, also within exon 6, have no e ffect on the level of nuclear TPI mRNA. In this work, quantitations of RNA that derived from TPI alleles in which nonsense codons had been g enerated between codons 189 and 208 revealed that the boundary between the two types of nonsense codons resides between codons 192 and 195. The analysis of TPI gene insertions and deletions indicated that the p ositional feature differentiating the two types of nonsense codons is the distance of the nonsense codon upstream of intron 6. For example, the movement of intron 6 to a position downstream of its normal locati on resulted in a concomitant downstream movement of the boundary betwe en the two types of nonsense codons. The analysis of intron 6 mutation s indicated that the intron 6 effect is stipulated by the 88 nucleotid es residing between the 5' and 3' splice sites. Since the deletion of intron 6 resulted in only partial abrogation of the nonsense codon-med iated reduction in the level of TPI mRNA, other sequences within TPI p re-mRNA must function in the effect. One of these sequences may be int ron 2, since the deletion of intron 2 also resulted in partial abrogat ion of the effect. In experiments that switched introns 2 and 6, the r eplacement of intron 6 with intron 2 was of no consequence to the effe ct of a nonsense codon within either exon 1 or exon 6. In contrast, th e replacement of intron 2 with intron 6 was inconsequential to the eff ect of a nonsense codon in exon 6 but resulted in partial abrogation o f a nonsense codon in exon 1.