DIFFERENTIAL-EFFECTS OF SIMPLE REPEATING DNA-SEQUENCES ON GENE-EXPRESSION FROM THE SV40 EARLY PROMOTER

The influence of simple repeat sequences, cloned into different positi ons relative to the SV40 early promoter/enhancer, on the transient exp ression of the chloramphenicol acetyltransferase (CAT) gene was invest igated. Insertion of (G)(29).(C)(29) in either orientation into the 5' -untranslated region of the CAT gene reduced expression in CV-1 cells 50-100 fold when compared with controls with random sequence inserts. Analysis of CAT-specific mRNA levels demonstrated that the effect was due to a reduction of CAT mRNA production rather than to posttranscrip tional events. In contrast, insertion of the same insert in either ori entation upstream of the promoter enhancer or downstream of the gene s timulated gene expression 2-3-fold. These effects could be reversed by cotransfection of a competitor plasmid carrying (G)(25).(C)(25) seque nces. The results suggest that a G.C-binding transcription factor modu lates gene expression in this system and that promoter strength can be regulated by providing protein binding sites in trans. Although const ructs containing longer tracts of alternating (C-G), (T-G), or (A-T) s equences inhibited CAT expression when inserted in the 5'-untranslated region of the CAT gene, the amount of CAT mRNA was unaffected. Hence, these inhibitions must be due to posttranscriptional events, presumab ly at the level of translation. These effects of microsatellite sequen ces on gene expression are discussed with respect to recent data on re lated simple repeat sequences which cause several human genetic diseas es.