- Relaxed Cleavage Specificity of Hyperactive Variants of Escherichia coli RNase E on RNA I
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Dayeong Bae , Hana Hyeon , Eunkyoung Shin , Ji , Kangseok Lee
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J. Microbiol. 2023;61(2):211-220. Published online February 22, 2023
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DOI: https://doi.org/10.1007/s12275-023-00013-z
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Abstract
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RNase E is an essential enzyme in Escherichia coli. The cleavage site of this single-stranded specific endoribonuclease is
well-characterized in many RNA substrates. Here, we report that the upregulation of RNase E cleavage activity by a mutation
that affects either RNA binding (Q36R) or enzyme multimerization (E429G) was accompanied by relaxed cleavage specificity.
Both mutations led to enhanced RNase E cleavage in RNA I, an antisense RNA of ColE1-type plasmid replication,
at a major site and other cryptic sites. Expression of a truncated RNA I with a major RNase E cleavage site deletion at the
5′-end (RNA I-
5) resulted in an approximately twofold increase in the steady-state levels of RNA I-
5 and the copy number
of ColE1-type plasmid in E. coli cells expressing wild-type or variant RNase E compared to those expressing RNA I. These
results indicate that RNA I-
5 does not efficiently function as an antisense RNA despite having a triphosphate group at the
5′-end, which protects the RNA from ribonuclease attack. Our study suggests that increased cleavage rates of RNase E lead
to relaxed cleavage specificity on RNA I and the inability of the cleavage product of RNA I as an antisense regulator in vivo
does not stem from its instability by having 5′-monophosphorylated end.
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Citations
Citations to this article as recorded by 
- Engineering an Escherichia coli based in vivo mRNA manufacturing platform
Edward Curry, George Muir, Jixin Qu, Zoltán Kis, Martyn Hulley, Adam Brown Biotechnology and Bioengineering.2024; 121(6): 1912. CrossRef
- Transcript-specific selective translation by specialized ribosomes bearing genome-encoded heterogeneous rRNAs in V. vulnificus CMCP6
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Younkyung Choi , Minju Joo , Wooseok Song , Minho Lee , Hana Hyeon , Hyun-Lee Kim , Ji-Hyun Yeom , Kangseok Lee , Eunkyoung Shin
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J. Microbiol. 2022;60(12):1162-1167. Published online November 24, 2022
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DOI: https://doi.org/10.1007/s12275-022-2437-9
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Abstract
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Ribosomes composed of genome-encoded heterogeneous
rRNAs are implicated in the rapid adaptation of bacterial
cells to environmental changes. A previous study showed that
ribosomes bearing the most heterogeneous rRNAs expressed
from the rrnI operon (I-ribosomes) are implicated in the preferential
translation of a subset of mRNAs, including hspA
and tpiA, in Vibrio vulnificus CMCP6. In this study, we show
that HspA nascent peptides were predominantly bound to
I-ribosomes. Specifically, I-ribosomes were enriched more
than two-fold in ribosomes that were pulled down by immunoprecipitation
of HspA peptides compared with the proportion
of I-ribosomes in crude ribosomes and ribosomes pulled
down by immunoprecipitation of RNA polymerase subunit
ß peptides in the wild-type (WT) and rrnI-completed strains.
Other methods that utilized the incorporation of an affinity
tag in 23S rRNA or chimeric rRNA tethering 16S and 23S
rRNAs, which generated specialized functional ribosomes
in Escherichia coli, did not result in functional I-ribosomes
in V. vulnificus CMCP6. This study provides direct evidence
of the preferential translation of hspA mRNA by I-ribosomes.
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Citations
Citations to this article as recorded by 
- Functional conservation of specialized ribosomes bearing genome-encoded variant rRNAs in Vibrio species
Younkyung Choi, Eunkyoung Shin, Minho Lee, Ji-Hyun Yeom, Kangseok Lee, Bashir Sajo Mienda PLOS ONE.2023; 18(12): e0289072. CrossRef - Relaxed Cleavage Specificity of Hyperactive Variants of Escherichia coli RNase E on RNA I
Dayeong Bae, Hana Hyeon, Eunkyoung Shin, Ji-Hyun Yeom, Kangseok Lee Journal of Microbiology.2023; 61(2): 211. CrossRef
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