POINT MUTATIONS 5 TO THE TRANSFER-RNA SELENOCYSTEINE TATA BOX ALTER RNA-POLYMERASE-III TRANSCRIPTION BY AFFECTING THE BINDING OF TBP

The selenocysteine tRNA(Sec) gene possesses two external promoter elem ents, one of which is constituted by a strong TATA box. Point mutant a nalysis performed in this study led to the conclusion that the functio nal TATA promoter actually encompasses the sequence -34 GGGTATAAAAGG-2 3. Individual changes at T-31 do not affect transcription much. Positi on T-29 is less permissive to mutation since transversion to a G, for example, is less well tolerated than at T-31. Interestingly, a double point mutation, converting GG(-33/-32) to TT, causes abrogation of tra nscription in vivo and severe reduction of transcription in vitro with human TBP. Therefore, data obtained underscore the fact that, in the Xenopus tRNA(Sec), these two Os are an integral part of the TATA promo ter. Gel retardation experiments indicate that the GG to TT substituti on, which led human TBP to lose its ability to support efficient trans cription in vitro, correlates with the appearance of an altered patter n of retarded complexes. Altogether, the data presented in this report support a model in which TBP interacts directly with the TATA element of the tRNA(Sec) gene, in contrast to the type of interaction propose d for classical TATA-less tRNA genes.