The role of highly conserved single-stranded nucleotides of Xenopus 5S RNA in the binding of transcription factor IIIA

Abstract

The role of highly conserved single-stranded sequence elements of Xenopus 5S RNA in the binding of transcription factor IIIA (TFIIIA) was studied. A series of mutant 5S RNA genes were constructed with defined block sequence changes in regions corresponding to each of the single-stranded loops of the transcribed 5S RNA. The interaction of the resulting mutant 5S RNA molecules with TFIIIA was determined both by a direct binding assay and by a competition assay. With one exception, the substitution of highly conserved single-stranded loop sequences had only a modest effect on the binding of TFIIIA. The single exception was loop A, which ironically is not part of the protected site of TFIIIA on 5S RNA. The possible involvement of loop A in the coaxial stacking of the helical domains of 5S RNA, and how this might affect TFIIIA binding, are discussed.