ALTERNATIVE FORMS OF THE HUMAN THIOREDOXIN MESSENGER-RNA - IDENTIFICATION AND CHARACTERIZATION

Thioredoxin (TRX) is an ubiquitous and relatively conserved oxidoreduc tant enzyme which is involved in a multitude of redox reactions throug h the formation of reversible disulfide bonds, A recent report indicat es the presence of novel isoforms of TRX proteins isolated from MP6 ce ll lines [Rosen et al., Int. Immunol. 7 (1995) 625-633], In these isof orms, as evidenced from amino acid sequencing, several Lys residues of the wild-type sequence were replaced by Arg, Although the human genom e contains several (isoformic) copies of the TRX gene, only one appear s to be transcriptionally active [Kaghad et al., Gene, 140 (1994) 273- 278]. As we characterized the isoforms of TRX mRNAs, we found that sev eral MP6 TRX cDNA clones were devoid of the characteristic poly(A) tai l. In order to increase the efficiency of isolating mRNAs without the poly(A) tail, we developed a novel procedure for exclusive capturing o f a specific mRNA by magnetic beads coated with biotinylated antisense oligodeoxyribonucleotide. Using this method on MP6 cell total RNA, we isolated an additional truncated version of the TRX mRNA. This latter form does not produce any variant TRX enzyme, as an inframe stop codo n truncates the product, This isoform was also present in mRNAs isolat ed from human placenta, leucocyte cells and Molt4 cells, the latter tw o being the progenitors of MP6 cells. From a thorough analysis of the sequence of the truncated TRX mRNA, we conclude that this variant orig inated from an event of altered splicing, as consensus splice sites we re present in the normal TRX cDNA at precise positions.