Chloroplast heteroplasmicity is stabilized by an amber-suppressor tryptophan tRNA(CUA).

Yu, Weizhu; Spreitzer, Robert J.

doi:10.1073/pnas.89.9.3904

Cited by 22 publications

(9 citation statements)

References 43 publications

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“…Chloroplast-based suppression of nonphotosynthetic phenotypes is well known in Chlamydomonas. Examples include point mutations that overcome translational incompetence in the psbC 59 UTR (Rochaix et al, 1989), a large intragenomic inversion that restored expression of a petD gene with a 59 UTR deletion (Higgs et al, 1998) and an amber suppressor tRNA that allowed expression of an rbcL mRNA possessing a stop codon (Yu and Spreitzer, 1992). In this context, the suppression we observed in all but spa19/23 was consistent with the known mutability of cpDNA, under conditions where expression of other essential genes remains sufficient.…”

Section: Dynamic Alteration Of Cpdna In Spa Strainsmentioning

confidence: 48%

Antisense Transcript and RNA Processing Alterations Suppress Instability of Polyadenylated mRNA in Chlamydomonas Chloroplasts

et al. 2004

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In chloroplasts, the control of mRNA stability is of critical importance for proper regulation of gene expression. The Chlamydomonas reinhardtii strain D26pAtE is engineered such that the atpB mRNA terminates with an mRNA destabilizing polyadenylate tract, resulting in this strain being unable to conduct photosynthesis. A collection of photosynthetic revertants was obtained from D26pAtE, and gel blot hybridizations revealed RNA processing alterations in the majority of these suppressor of polyadenylation (spa) strains, resulting in a failure to expose the atpB mRNA 39 poly(A) tail. Two exceptions were spa19 and spa23, which maintained unusual heteroplasmic chloroplast genomes. One genome type, termed PSþ, conferred photosynthetic competence by contributing to the stability of atpB mRNA; the other, termed PSÿ, was required for viability but could not produce stable atpB transcripts. Based on strand-specific RT-PCR, S1 nuclease protection, and RNA gel blots, evidence was obtained that the PSþ genome stabilizes atpB mRNA by generating an atpB antisense transcript, which attenuates the degradation of the polyadenylated form. The accumulation of double-stranded RNA was confirmed by insensitivity of atpB mRNA from PSþ genome-containing cells to S1 nuclease digestion. To obtain additional evidence for antisense RNA function in chloroplasts, we used strain D26, in which atpB mRNA is unstable because of the lack of a 39 stem-loop structure. In this context, when a 121-nucleotide segment of atpB antisense RNA was expressed from an ectopic site, an elevated accumulation of atpB mRNA resulted. Finally, when spa19 was placed in a genetic background in which expression of the chloroplast exoribonuclease polynucleotide phosphorylase was diminished, the PSþ genome and the antisense transcript were no longer required for photosynthesis. Taken together, our results suggest that antisense RNA in chloroplasts can protect otherwise unstable transcripts from 39!59 exonuclease activity, a phenomenon that may occur naturally in the symmetrically transcribed and densely packed chloroplast genome.

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Section: Dynamic Alteration Of Cpdna In Spa Strainsmentioning

confidence: 48%

Antisense Transcript and RNA Processing Alterations Suppress Instability of Polyadenylated mRNA in Chlamydomonas Chloroplasts

et al. 2004

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“…In Chlamydomonas , an amber suppressor tRNA was identified in the chloroplast (33), as it restored photosynthesis to a strain bearing a nonsense mutation in the rbcL gene, which encodes the Rubisco large subunit. Although the tRNA Trp which was mutated is a single-copy gene, the polyploid nature of the chloroplast allowed heteroplasmic maintenance of the suppressor.…”

Section: Discussionmentioning

confidence: 99%

A spontaneous tRNA suppressor of a mutation in the Chlamydomonas reinhardtii nuclear MCD1 gene required for stability of the chloroplast petD mRNA

Murakami

Kuehnle

Stern

2005

Nucleic Acids Research

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Numerous nuclear gene products are required for the correct expression of organellar genes. One such gene in the green alga Chlamydomonas reinhardtii is MCD1, whose product is required for stability of the chloroplast-encoded petD mRNA. In mcd1 mutants, which are non-photosynthetic, petD mRNA is degraded by a 5′–3′ exonuclease activity, resulting in a failure to synthesize its product, subunit IV of the cytochrome b 6/f complex. Here, we report the sequence of the wild-type MCD1 gene, which encodes a large and novel putative protein. Analysis of three mutant alleles showed that two harbored large deletions, but that one allele, mcd1-2, had a single base change resulting in a nonsense codon near the N-terminus. This same mutant allele can be suppressed by a second-site mutation in the nuclear MCD2 gene, whereas mcd2-1 cannot suppress the deletion in mcd1-1 (Esposito,D. Higgs,D.C. Drager,R.G. Stern, D.B. and Girard-Bascou,J. (2001) Curr. Genet., 39, 40–48). We report the cloning of mcd2-1, and show that the mutation lies in a tRNASer(CGA), which has been modified to translate the nonsense codon in mcd1-2. We discuss how the existence of a large tRNASer gene family may permit this suppression without pleiotropic consequences.

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“…Compensation by mutations in other genes (i.e., extragenic or intergenic compensation) is often mediated by the relationship between gene products. For example, a compensatory mutation in a gene involved in translation can ameliorate the phenotypic effects of a mutation by occasionally mistranslating the mutant mRNA into a normal protein product (Hartman and Roth 1973;Waterston and Brenner 1978;Murgola 1985;Yu and Spreitzer 1992;El Mezaine et al 1998); often these compensatory mutations turn out to be point-mutations in certain tRNA genes. Alternatively, compensation can be mediated through biochemical pathways (Hartman and Roth 1973).…”

Section: Evidence For Compensatory Mutationsmentioning

confidence: 99%

Compensating for Our Load of Mutations: Freezing the Meltdown of Small Populations

2000

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We have investigated the reduction of fitness caused by the fixation of new deleterious mutations in small populations within the framework of Fisher's geometrical model of adaptation. In Fisher's model, a population evolves in an n-dimensional character space with an adaptive optimum at the origin. The model allows us to investigate compensatory mutations, which restore fitness losses incurred by other mutations, in a context-dependent manner. We have conducted a moment analysis of the model, supplemented by the numerical results of computer simulations. The mean reduction of fitness (i.e., expected load) scaled to one is approximately n/(n+2Ne), where Ne is the effective population size. The reciprocal relationship between the load and Ne implies that the fixation of deleterious mutations is unlikely to cause extinction when there is a broad scope for compensatory mutations, except in very small populations. Furthermore, the dependence of load on n implies that pleiotropy plays a large role in determining the extinction risk of small populations. Differences and similarities between our results and those of a previous study on the effects of Ne and n are explored. That the predictions of this model are qualitatively different from studies ignoring compensatory mutations implies that we must be cautious in predicting the evolutionary fate of small populations and that additional data on the nature of mutations is of critical importance.

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Chloroplast heteroplasmicity is stabilized by an amber-suppressor tryptophan tRNA(CUA).

Abstract: Photosynthesis-deficient mutants of the green alga Chlamydomonas reinhardti were previously shown to arise from nonsense mutations within the chloroplast rbcL gene, which encodes the large subunit of ribulose-1,5-bisphosphate

Cited by 22 publications

References 43 publications

Antisense Transcript and RNA Processing Alterations Suppress Instability of Polyadenylated mRNA in Chlamydomonas Chloroplasts

Antisense Transcript and RNA Processing Alterations Suppress Instability of Polyadenylated mRNA in Chlamydomonas Chloroplasts

A spontaneous tRNA suppressor of a mutation in the Chlamydomonas reinhardtii nuclear MCD1 gene required for stability of the chloroplast petD mRNA

Compensating for Our Load of Mutations: Freezing the Meltdown of Small Populations

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