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Nucleic Acids Research 2004 32(14):4297-4305; doi:10.1093/nar/gkh769
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Published online 10 August 2004

Nucleic Acids Research, Vol. 32 No. 14 © Oxford University Press 2004; all rights reserved

Position-dependent function of a B block promoter element implies a specialized chromatin structure on the S.cerevisiae U6 RNA gene, SNR6

Michael W. Kaiser, Jing Chi and David A. Brow*

Department of Biomolecular Chemistry, University of Wisconsin Medical School, Madison, WI 53706-1532, USA

* To whom correspondence should be addressed. Tel: +1 608 262 1475; Fax: +1 608 262 5253; Email: dabrow{at}wisc.edu
Present address: Michael W. Kaiser, Third Wave Technologies, Inc., Madison, WI 53719, USA

Received June 17, 2004; Revised and Accepted July 27, 2004

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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