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Distribution of 5'-triphosphate termini on the mRNA of Escherichia coli
1Department of Microbiology, University of California, Los Angeles 90024.
Abstract:
We have determined the distribution of 5'-nucleoside triphosphates on the RNA in Escherichia coli. These groups represent the initial nucleoside triphosphate incorporated when RNA polymerase initiates transcription. It was estimated that at least 15% of polysome-associated messengers had triphosphates. This was interpreted to mean that removal of the triphosphate or messenger leader is not necessary for the functioning of most mRNAs but that a substantial amount of messenger processing occurs in the polysome pool. We found that the ratio of GTP- to ATP-initiated messengers was about 2 to 1. Since prior work has indicated that G- and A-initiated RNAs decay at the same rate and since a compilation of messenger start sites shows an A preference, this value implies that there is a significant physiological selection of G-initiated transcripts. We also characterized the 5'-terminal groups on RNAs in other fractions. A small amount was found associated with 30S ribosomes, presumably in initiation complexes; such complexes have not previously been detected in situ. In addition, it was concluded that the 5' terminus of rRNA precursors is processed more rapidly than is implied by the current literature.
Insights
Researchers studied 5'-nucleoside triphosphates in E. coli RNA, finding most mRNAs function without triphosphate removal. Significant processing occurs in polysomes, with a 2:1 GTP- to ATP-initiated messenger ratio suggesting physiological selection.
Area of Science:
- Molecular Biology
- Microbiology
- Genetics
Background:
- RNA polymerase initiates transcription by incorporating nucleoside triphosphates.
- The 5' end of mRNA contains a triphosphate group that can be modified or removed.
- Understanding RNA processing and initiation is crucial for gene expression regulation.
Purpose of the Study:
- To determine the distribution of 5 eal'-nucleoside triphosphates on RNA in Escherichia coli.
- To investigate the functional significance of the triphosphate group in mRNA.
- To analyze the ratio of GTP- to ATP-initiated transcripts and its implications for gene regulation.
Main Methods:
- Analysis of 5 eal'-nucleoside triphosphate distribution on RNA fractions from Escherichia coli.
- Quantification of triphosphate-containing mRNAs in polysome-associated fractions.
- Characterization of 5 eal'-terminal groups on various RNA species, including those associated with 30S ribosomes.
Main Results:
- At least 15% of polysome-associated messenger RNAs (mRNAs) retained their 5 eal'-triphosphate groups.
- The ratio of guanosine triphosphate (GTP)- to adenosine triphosphate (ATP)-initiated messengers was approximately 2:1.
- 5 eal'-terminal groups were detected on RNAs in 30S ribosome initiation complexes, previously undetected in situ.
- The processing of the 5 eal' terminus of ribosomal RNA (rRNA) precursors was found to be faster than previously suggested.
Conclusions:
- Removal of the 5 eal'-triphosphate group is not essential for the function of most mRNAs, indicating substantial processing occurs within the polysome pool.
- The observed GTP- to ATP-initiated messenger ratio suggests significant physiological selection favoring G-initiated transcripts, despite prior evidence of an A preference in messenger start sites.
- The detection of initiation complexes on 30S ribosomes provides in situ evidence for early-stage transcription initiation.
- The rapid processing of rRNA precursors necessitates a re-evaluation of current models.