A NOVEL TRANSFORMING GROWTH FACTOR-BETA(1) RESPONSIVE CYTOPLASMIC TRANS-ACTING FACTOR BINDS SELECTIVELY TO THE 3'-UNTRANSLATED REGION OF MAMMALIAN RIBONUCLEOTIDE REDUCTASE-R2 MESSENGER-RNA - ROLE IN MESSAGE STABILITY

Ribonucleotide reductase is a highly regulated enzyme that provides th e four deoxyribonucleotides required for DNA synthesis. Our studies sh owed that TGF-beta1 treatment of BALB/c 3T3 mouse fibroblasts markedly elevated ribonucleotide reductase R2 mRNA levels, and also increased the half-life of R2 message by 4-fold from 1.5 h in untreated cells to 6 h in treated cells. We describe a novel 75 Kd sequence-specific cyt oplasmic factor (p75) that binds selectively to a 83-nucleotide 3'-unt ranslated region of R2 mRNA and did not bind to the 5'UTR, the coding region of the R2 message or to the 3'UTRs of other mRNAs (from c-myc, GM-CSF and the iron responsive element from the transferrin receptor m RNA), or to the homopolymer poly(A) sequence. p75-RNA binding activity , which requires new protein synthesis, is not present in untreated ce lls, but is induced following TGF-beta1 stimulation. The in vivo kinet ics of appearance of p75 binding activity paralleled the accumulation of R2 mRNA. Insertion of the 3'-untranslated region into the chloramph enicol acetyltransferase (CAT) message confers TGF-beta1 induced stabi lity of RNA in stably transfected cells, while the same insert carryin g a deletion of the 83-nucleotide fragment had little affect on RNA le vels. Furthermore, in vitro decay reactions that contained the 83-nucl eotide RNA or deletion of this fragment caused a significant decrease in TGF-beta1 stabilization of R2 message. A model is presented of R2 m essage regulation in which TGF-beta1 mediated stabilization of R2 mess age involves a specific interaction of a p75-trans-acting factor with a cis-element(s) stability determinant within the 83-nucleotide sequen ce which is linked to a reduction in the rate of R2 mRNA degradation.