The Saccharomyces cerevisiae U6 RNA gene, SNR6, is transcribed by RNA polymerase III (Pol III), but lacks the intragenic B block promoter element found in most other Pol III transcription units. Rather, the SNR6 B block element is located 120 bp downstream of the terminator. In contrast, the Schizosaccharomyces pombe U6 RNA gene has an intragenic B block sequence in a short intron. We show that the S.pombe U6 intron, when inserted into SNR6, can functionally replace the downstream B block in vitro but not in vivo. The in vivo expression defect is caused by at least three different effects of the insertion: (i) the S.pombe intron is inefficiently spliced in S.cerevisiae due to the short distance between the 5' splice site and branchpoint; (ii) the S.pombe B block sequence is suboptimal for S.cerevisiae; and (iii) a B block does not function well within the context of the SNR6 intron, especially when the gene is present at its normal chromosomal locus rather than on a plasmid. This last observation suggests that the chromatin structure of the SNR6 locus favors utilization of a downstream B block element. We also provide evidence that splicing of U6 RNA reduces its activity, presumably due to alterations in U6 RNA structure, localization and/or assembly into the spliceosome.

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Journal Article | Research Support, U.S. Gov't, P.H.S.

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Kaiser MW, Chi J, Brow DA

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