POINT MUTATIONS IN THE STEM-LOOP AT THE 3' END OF MOUSE HISTONE MESSENGER-RNA REDUCE EXPRESSION BY REDUCING THE EFFICIENCY OF 3' END FORMATION

Mammalian histone mRNAs end in a highly conserved stem-loop structure, with a six-base stem and a four-base loop. We have examined the effec t of mutating the stem-loop on the expression of the histone mRNA in v ivo by introducing the mutated histone genes into CHO cells by stable transfection. Point mutations have been introduced into the loop seque nce and into the UA base pair at the top of the stem. Changing either the first or the third base of the conserved UYUN sequence in the loop to a purine greatly reduced expression, while changing both U's to pu rines abolished expression. A number of alterations in the stem sequen ce, including reversing the stem sequence, reversing the two base pair s at the base of the stem, or destroying the UA base pair at the top o f the stem, also abolished expression. Changing the UA base pair to a CG or a UG base pair also reduced expression. The loss of expression i s due to inefficient processing of the pre-mRNA, as judged by the effi ciency of processing in vitro. Addition of a polyadenylation site or t he wild-type histone processing signal downstream of a mutant stem-loo p resulted in rescuing the processing of the mutant pre-histone mRNA. These results suggest that if the histone pre-mRNA is not rapidly proc essed, then it is degraded.