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Volume 271, Number 45, Issue of November 8, 1996 pp. 28146-28153
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.

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Transcriptional Silencing of a tRNA₁^Gly Copy from within a Multigene Family Is Modulated by Distal cis Elements

(Received for publication, April 26, 1996, and in revised form, July 22, 1996)

From the Department of Microbiology and Cell Biology and Center for Genetic Engineering, Indian Institute of Science, Bangalore 560 012, India

Individual copies of tRNA₁^Gly from within the multigene family in Bombyx mori could be classified based on in vitro transcription in homologous nuclear extracts into three categories of highly, moderately, or weakly transcribed genes. Segregation of the poorly transcribed gene copies 6 and 7, which are clustered in tandem within 425 base pairs, resulted in enhancement of their individual transcription levels, but the linkage itself had little influence on the transcriptional status. For these gene copies, when fused together generating a single coding region, transcription was barely detectable, which suggested the presence of negatively regulating elements located in the far flanking sequences. They exerted the silencing effect on transcription overriding the activity of positive regulatory elements. Systematic analysis of deletion, chimeric, and mutant constructs revealed the presence of a sequence element TATATAA located beyond 800 nucleotides upstream to the coding region acting as negative modulator, which when mutated resulted in high level transcription. Conversely, a TATATAA motif reintroduced at either far upstream or far downstream flanking regions exerted a negative effect on transcription. The location of cis-regulatory sequences at such farther distances from the coding region and the behavior of TATATAA element as negative regulator reported here are novel. These element(s) could play significant roles in activation or silencing of genes from within a multigene family, by recruitment or sequestration of transcription factors.

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