| 1. |
- Byström, Anders S, et al.
(författare)
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Differentially expressed trmD ribosomal protein operon of Escherichia coli is transcribed as a single polycistronic mRNA species
- 1989
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Ingår i: Journal of Molecular Biology. - : Academic Press. - 0022-2836 .- 1089-8638. ; 208:4, s. 575-586
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Tidskriftsartikel (refereegranskat)abstract
- The trmD operon is a four-cistron operon in which the first and fourth genes encode ribosomal proteins S16 (rpsP) and L19 (rplS), respectively. The second gene encodes a 21,000 Mr polypeptide of unknown function and the third gene (trmD) encodes the enzyme tRNA(m1G37)methyltransferase, which catalyzes the formation of 1-methylguanosine (m1G) next to the 3' end of the anticodon (position 37) of some tRNAs in Escherichia coli. Here we show under all regulatory conditions studied, transcription initiates at one unique site, and the entire operon is transcribed into one polycistronic mRNA. Between the promoter and the first gene, rpsP, an attenuator-like structure is found (delta G = -18 kcal; 1 cal = 4.184 J), followed by four uridine residues. This structure is functional in vitro, and terminates more than two-thirds of the transcripts. The different parts of the trmD operon mRNA decay at a uniform rate. The stability of the trmD mRNA is not reduced with decreasing growth rate, which is in contrast to what has been found for other ribosomal protein mRNAs. Furthermore, earlier experiments have shown the existence of differential expression as well as non-co-ordinate regulation within the operon. Our results are consistent with the regulation of the trmD operon being due to some mechanism(s) operating at the post-transcriptional level, and do not involve differential degradation of different mRNA segments, internal promoters or internal terminators.
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| 2. |
- Kirsebom, Leif A, et al.
(författare)
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Differential effects of mutations in the protein and RNA moieties of RNase P on the efficiency of suppression by various tRNA suppressors
- 1988
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Ingår i: Journal of Molecular Biology. - : Elsevier BV. - 0022-2836 .- 1089-8638. ; 204:4, s. 879-888
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Tidskriftsartikel (refereegranskat)abstract
- We have studied the efficiency of suppression by tRNA suppressors in vivo in strains of Escherichia coli that harbor a mutation in the rnpA gene, the gene for the protein component (C5) of RNase P, and in strains that carry several different alleles of the rnpB gene, the gene for the RNA component (M1) of RNase P. Depending on the genetic background, different efficiencies of suppression by the various tRNA suppressors were observed. Thus, mutations in rnpA have separable and distinct effects from mutations in rnpB on the processing of tRNA precursors by RNase P. In addition, the efficiency of suppression by several derivatives of E. coli tRNA(Tyr) Su3 changed as the genetic background was altered.
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| 3. |
- Kirsebom, Leif A, et al.
(författare)
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Reaction in vitro of some mutants of RNase P with wild-type and temperature-sensitive substrates
- 1989
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Ingår i: Journal of Molecular Biology. - : Elsevier BV. - 0022-2836 .- 1089-8638. ; 207:4, s. 837-840
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- The reaction of wild-type and two mutant derivatives of RNase P have been examined with wild-type and mutant substrates. We show that a mutant derivative of tRNA(Tyr)Su3, tRNA(Tyr)Su3A15, in which the G15.C48(57) base-pair essential for folding of the tRNA moiety is altered, is a temperature-sensitive suppressor in vivo. The precursor to tRNA(Tyr)Su3A15 is cleaved in a temperature-sensitive manner in vitro by RNase P and with a higher Km compared to the precursor to tRNA(Tyr)Su3. The precursor to tRNA(Tyr)Su3A2, another temperature-sensitive suppressor in vivo in which the G2.C71(80) base-pair in the acceptor stem is changed to A2.C71(80), behaves like the precursor to tRNA(Tyr)Su3 in vitro; that is, it is not cleaved in a temperature-sensitive manner. Therefore, there are at least two ways in which a suppressor tRNA can acquire a temperature-sensitive phenotype in vivo. One of the mutant derivatives of RNase P we have tested, rnpA49, which affects the protein cofactor of the enzyme, has a decreased kcat compared to wild-type, which can explain its phenotype in vivo.
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| 4. |
- Wikström, P M, et al.
(författare)
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Non-autogenous control of ribosomal protein synthesis from the trmD operon in Escherichia coli
- 1988
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Ingår i: Journal of Molecular Biology. - : Elsevier BV. - 0022-2836 .- 1089-8638. ; 203:1, s. 141-152
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Tidskriftsartikel (refereegranskat)abstract
- The trmD operon of Escherichia coli encodes the ribosomal proteins S16 and L19, the tRNA(m1G37)methyltransferase and a 21,000 Mr protein of unknown function. Here we demonstrate that, in contrast to the expression of other ribosomal protein operons, the amount of trmD operon mRNA and the rate of synthesis of the proteins encoded by the operon respond to increased gene dosage. The steady-state level of the mRNA was about 18 times higher, and the relative rate of synthesis of the ribosomal proteins S16 and L19, the tRNA(m1G37)methyltransferase and the 21,000 Mr protein was 15, 9, 25 and 23 times higher, respectively, in plasmid-containing cells than in plasmid-free cells. Overproduced tRNA(m1G37)methyltransferase and 21,000 Mr protein were as stable as E. coli total protein, whereas the two ribosomal proteins were degraded to a large extent. The steady-state amount of S16 and L19 in the plasmid-containing cells exceeded that in plasmid-free cells by threefold and twofold, respectively. No significant effect on the synthesis of the trmD operon proteins from the chromosomally located genes was observed when parts of the operon were expressed on different plasmids. Taken together, these results suggest that the expression of the trmD operon is not subject to transcriptional or translational feedback regulation, and demonstrate that not all ribosomal protein operons are regulated in the same manner. We propose that ribosomal protein operons that do not encode proteins that bind directly to rRNA are not under autogenous control. Metabolic regulation at the transcriptional level and protein degradation are plausible mechanisms for the control of expression of such operons.
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| 5. |
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