Functional pleiotropy of an intramolecular triplex-forming fragment from the 3 '-UTR of the rat Pigr gene

A microsatellite-containing 359-bp restriction fragment, isolated from the rat Pigr gene (murine polymeric immunoglobulin receptor gene) 3'-untranslat ed region (3'-UTR) and inserted into 3'-UTR or 3' flanking positions in tra nscription units of supercoiled plasmids, attenuates luciferase reporter ge ne expression in orientation- and position-dependent ways following transie nt transfection of human 293 cells. The same fragment stimulates orientatio n- dependent gene expression in a 5' flanking position. Plasmid linearizati on abrogates both orientation- and position-dependent responses. Cell-free translation reveals that 5' and 3' flanking expression responses are propor tional to increased and decreased luciferase mRNA levels, whereas 3'-UTR ex pression is associated with control mRNA levels. Hypersensitivity to nuclea ses S1 and P1, gel mobility differences between supercoiled plasmids carryi ng opposing microsatellite orientations, and anomalous melting profiles of this fragment are also observed. These results suggest that functional plei otropy of this fragment depends on the DNA context of its purine-rich micro satellite strand and on DNA supercoiling. Intramolecular triplexes stabiliz ed by supercoiling and secondary structures of purine repeat-rich mRNAs may also confer regulatory properties to similar genomic elements.