Analysis of the upstream region of the Escherichia coli rnd gene encoding RNase D. Evidence for translocational regulation of a putative tRNA processing enzyme

J. Zhang, Murray P Deutscher

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

The upstream region of the Escherichia coli rnd gene has been analyzed. A single promoter and transcription start site were identified in the 170-nucleotide upstream region of the clone based on deletion analysis, nuclease S1 mapping, primer extension, and quantitative dot-blots. The start site of transcription is 70 nucleotides upstream of the UUG initiation codon of the RNase D coding region. Between the RNA initiation site and the coding region is a GC-rich hairpin structure followed by 8 T residues that looks like a ρ-independent transcription terminator. Surprisingly, deletion of the hairpin structure elevates rnd mRNA levels only slightly (<2-fold), but it dramatically decreases RNase D expression (>95%), measured both by activity and immunoblotting. No evidence for an effect on mRNA stability or processing was observed. The uncommon UUG translation initiation codon was converted to AUG by site-directed mutagenesis. This alteration led to an 11-fold elevation of RNase D expression in single-copy plasmids. These data indicate that RNase D epxression is negatively regulated at the translational level by the initiation codon, and that an upstream structure (or sequence) also influences translation of this gene.

Original languageEnglish
Pages (from-to)18228-18233
Number of pages6
JournalJournal of Biological Chemistry
Volume264
Issue number30
StatePublished - Jan 1 1989
Externally publishedYes

Fingerprint

Ribonuclease III
Gene encoding
Initiator Codon
Transfer RNA
Escherichia coli
Transcription Initiation Site
Enzymes
Nucleotides
Processing
Genes
Mutagenesis
Messenger RNA
RNA Stability
Transcription
Site-Directed Mutagenesis
Immunoblotting
Plasmids
Clone Cells
RNA

ASJC Scopus subject areas

  • Biochemistry

Cite this

Analysis of the upstream region of the Escherichia coli rnd gene encoding RNase D. Evidence for translocational regulation of a putative tRNA processing enzyme. / Zhang, J.; Deutscher, Murray P.

In: Journal of Biological Chemistry, Vol. 264, No. 30, 01.01.1989, p. 18228-18233.

Research output: Contribution to journalArticle

@article{8b20291fe041494f8f58043158415235,
title = "Analysis of the upstream region of the Escherichia coli rnd gene encoding RNase D. Evidence for translocational regulation of a putative tRNA processing enzyme",
abstract = "The upstream region of the Escherichia coli rnd gene has been analyzed. A single promoter and transcription start site were identified in the 170-nucleotide upstream region of the clone based on deletion analysis, nuclease S1 mapping, primer extension, and quantitative dot-blots. The start site of transcription is 70 nucleotides upstream of the UUG initiation codon of the RNase D coding region. Between the RNA initiation site and the coding region is a GC-rich hairpin structure followed by 8 T residues that looks like a ρ-independent transcription terminator. Surprisingly, deletion of the hairpin structure elevates rnd mRNA levels only slightly (<2-fold), but it dramatically decreases RNase D expression (>95{\%}), measured both by activity and immunoblotting. No evidence for an effect on mRNA stability or processing was observed. The uncommon UUG translation initiation codon was converted to AUG by site-directed mutagenesis. This alteration led to an 11-fold elevation of RNase D expression in single-copy plasmids. These data indicate that RNase D epxression is negatively regulated at the translational level by the initiation codon, and that an upstream structure (or sequence) also influences translation of this gene.",
author = "J. Zhang and Deutscher, {Murray P}",
year = "1989",
month = "1",
day = "1",
language = "English",
volume = "264",
pages = "18228--18233",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "30",

}

TY - JOUR

T1 - Analysis of the upstream region of the Escherichia coli rnd gene encoding RNase D. Evidence for translocational regulation of a putative tRNA processing enzyme

AU - Zhang, J.

AU - Deutscher, Murray P

PY - 1989/1/1

Y1 - 1989/1/1

N2 - The upstream region of the Escherichia coli rnd gene has been analyzed. A single promoter and transcription start site were identified in the 170-nucleotide upstream region of the clone based on deletion analysis, nuclease S1 mapping, primer extension, and quantitative dot-blots. The start site of transcription is 70 nucleotides upstream of the UUG initiation codon of the RNase D coding region. Between the RNA initiation site and the coding region is a GC-rich hairpin structure followed by 8 T residues that looks like a ρ-independent transcription terminator. Surprisingly, deletion of the hairpin structure elevates rnd mRNA levels only slightly (<2-fold), but it dramatically decreases RNase D expression (>95%), measured both by activity and immunoblotting. No evidence for an effect on mRNA stability or processing was observed. The uncommon UUG translation initiation codon was converted to AUG by site-directed mutagenesis. This alteration led to an 11-fold elevation of RNase D expression in single-copy plasmids. These data indicate that RNase D epxression is negatively regulated at the translational level by the initiation codon, and that an upstream structure (or sequence) also influences translation of this gene.

AB - The upstream region of the Escherichia coli rnd gene has been analyzed. A single promoter and transcription start site were identified in the 170-nucleotide upstream region of the clone based on deletion analysis, nuclease S1 mapping, primer extension, and quantitative dot-blots. The start site of transcription is 70 nucleotides upstream of the UUG initiation codon of the RNase D coding region. Between the RNA initiation site and the coding region is a GC-rich hairpin structure followed by 8 T residues that looks like a ρ-independent transcription terminator. Surprisingly, deletion of the hairpin structure elevates rnd mRNA levels only slightly (<2-fold), but it dramatically decreases RNase D expression (>95%), measured both by activity and immunoblotting. No evidence for an effect on mRNA stability or processing was observed. The uncommon UUG translation initiation codon was converted to AUG by site-directed mutagenesis. This alteration led to an 11-fold elevation of RNase D expression in single-copy plasmids. These data indicate that RNase D epxression is negatively regulated at the translational level by the initiation codon, and that an upstream structure (or sequence) also influences translation of this gene.

UR - http://www.scopus.com/inward/record.url?scp=0024452693&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0024452693&partnerID=8YFLogxK

M3 - Article

VL - 264

SP - 18228

EP - 18233

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 30

ER -