Abstract

BackgroundSynonymous sites are freer to vary because of redundancy in genetic code. Messenger RNA secondary structure restricts this freedom, as revealed by previous findings in mitochondrial genes that mutations at third codon position nucleotides in helices are more selected against than those in loops. This motivated us to explore the constraints imposed by mRNA secondary structure on evolutionary variability at all codon positions in general, in chloroplast systems.ResultsWe found that the evolutionary variability and intrinsic secondary structure stability of these sequences share an inverse relationship. Simulations of most likely single nucleotide evolution in Psilotum nudum and Nephroselmis olivacea mRNAs, indicate that helix-forming propensities of mutated mRNAs are greater than those of the natural mRNAs for short sequences and vice-versa for long sequences. Moreover, helix-forming propensity estimated by the percentage of total mRNA in helices increases gradually with mRNA length, saturating beyond 1000 nucleotides. Protection levels of functionally important sites vary across plants and proteins: r-strategists minimize mutation costs in large genes; K-strategists do the opposite.ConclusionMrna length presumably predisposes shorter mRNAs to evolve under different constraints than longer mRNAs. The positive correlation between secondary structure protection and functional importance of sites suggests that some sites might be conserved due to packing-protection constraints at the nucleic acid level in addition to protein level constraints. Consequently, nucleic acid secondary structure a priori biases mutations. The converse (exposure of conserved sites) apparently occurs in a smaller number of cases, indicating a different evolutionary adaptive strategy in these plants. The differences between the protection levels of functionally important sites for r- and K-strategists reflect their respective molecular adaptive strategies. These converge with increasing domestication levels of K-strategists, perhaps because domestication increases reproductive output.

Highlights

  • Synonymous sites are freer to vary because of redundancy in genetic code

  • We found that for shorter sequences, a greater percentage of the secondary structure of mutated mRNAs is composed of helices, than that for the natural mRNAs

  • Secondary structural folding stability of nucleic acid sequences modulates their evolutionary variability We tested for correlations between the length-adjusted residual stability measure for each gene and that gene's relative fit to four pre-defined substitution rate categories (M1, M2, M3 and M4)

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Summary

Methods

Sequence Dataset RNA-synthesis (sense) strand sequences were extracted from NCBI [59] using in-house PERL scripts for thirty-five genes (atpB, petA, petL, rps, rpl, rpl, rps, rps, atpI, atpH, rpoB, rps, rps, psbK, psbL, psbE, psbF, rps, rpl, psbH, psbN, psbB, psaA, psaB, psaC, atpE, psbA, psbD, psaJ, clp, atpF, psbC, rpl, psbI, and rps8) of 17 plant species (Oryza sativa (japonica cultivargroup) (NC_001320; [60]); Triticum aestivum (NC_002762; [61]); Zea mays (NC_001666; [62]); Calycanthus floridus var. glaucus (NC_004993; [63]); Arabidopsis thaliana (NC_000932; [64]); Pinus koraiensis (NC_004677; [65]); Pinus thunbergii (NC_001631; [66]); Nephroselmis olivacea (NC_000927; [67]); Marchantia polymorpha (NC_001319; [68]); Mesostigma viride (NC_002186; [69]); Chlorella vulgaris (NC_001865; [70]); Chlamydomonas reinhardtii (NC_005353; [71]); Psilotum nudum (NC_003386; [72]); Chaetosphaeridium globosum (NC_004115; [73]); Physcomitrella patens subsp. patens (NC_005087; [74]); Spinacea oleracea (NC_002202; [75]) and Adiantum capillus-veneris (NC_004766; [76]) belonging to the family Viridiplantae.We considered as the anti-sense (coding) strand, the strand that coded for a majority of 23 out of 35 sampled genes. For the 12 sense-stranded coded genes (clpP, psbE, psbF, petL, petA, psaJ, psbC, psbB, psbD, psbK, psbH, rps18), we considered the sequence as it is and for the remaining genes (atpI, atpF, atpH, atpB, atpE, psaA, psaB, psaC, psbA, psbL, psbN, psbI, rpoB, rpl, rpl, rpl, rpl, rps, rps, rps, rps, rps, rps8), we analyzed the reverse complementary sequence. This combination of 17 species and 35 genes was chosen because it consists of an ample number of species and at the same time, a sufficiently large number of homologous genes (concatenated alignment of 27,465 nucleotides) for building the http://www.biomedcentral.com/1471-2164/9/48 chloroplast phylogeny.

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