Lineage-specific variations of congruent evolution among DNA sequences from three genomes, and relaxed selective constraints on rbcL in Cryptomonas (Cryptophyceae)

Abstract

Background: Plastid-bearing cryptophytes like Cryptomonas contain four genomes in a cell, the nucleus, the nucleomorph, the plastid genome and the mitochondrial genome. Comparative phylogenetic analyses encompassing DNA sequences from three different genomes were performed on nineteen photosynthetic and four colorless Cryptomonas strains. Twenty-three rbcL genes and fourteen nuclear SSU rDNA sequences were newly sequenced to examine the impact of photosynthesis loss on codon usage in the rbcL genes, and to compare the rbcL gene phylogeny in terms of tree topology and evolutionary rates with phylogenies inferred from nuclear ribosomal DNA (concatenated SSU rDNA, ITS2 and partial LSU rDNA), and nucleomorph SSU rDNA.

Results: Largely congruent branching patterns and accelerated evolutionary rates were found in nucleomorph SSU rDNA and rbcL genes in a clade that consisted of photosynthetic and colorless species suggesting a coevolution of the two genomes. The extremely accelerated rates in the rbcL phylogeny correlated with a shift from selection to mutation drift in codon usage of two-fold degenerate NNY codons comprising the amino acids asparagine, aspartate, histidine, phenylalanine, and tyrosine. Cysteine was the sole exception. The shift in codon usage seemed to follow a gradient from early diverging photosynthetic to late diverging photosynthetic or heterotrophic taxa along the branches. In the early branching taxa, codon preferences were changed in one to two amino acids, whereas in the late diverging taxa, including the colorless strains, between four and five amino acids showed changes in codon usage.

Conclusion: Nucleomorph and plastid gene phylogenies indicate that loss of photosynthesis in the colorless Cryptomonas strains examined in this study possibly was the result of accelerated evolutionary rates that started already in photosynthetic ancestors. Shifts in codon usage are usually considered to be caused by changes in functional constraints and in gene expression levels. Thus, the increasing influence of mutation drift on codon usage along the clade may indicate gradually relaxed constraints and reduced expression levels on the rbcL gene, finally correlating with a loss of photosynthesis in the colorless Cryptomonas paramaecium strains.

Figure 1

Unrooted maximum likelihood trees of DNA sequences representing three different genomes of the cryptophyte genus Cryptomonas. Figure 1A – Tree inferred from concatenated nuclear SSU rDNA, ITS2 and partial LSU rDNA sequences. Evolutionary model, GTR+I+Γ [51]; -ln L = 9254.5. Figure 1B – Nucleomorph SSU rDNA phylogeny. Evolutionary model, TVM+I+Γ [51]; -ln L = 4899.1. Figure 1C – Tree inferred from plastid-encoded rbcL genes (for a rooted tree including rbcL genes of other cryptophyte genera, see Additional file 3). Evolutionary model, GTR+I+Γ [51]; -ln L = 7857.4. Figure 1D (inlet) – Nuclear (top), nucleomorph (middle) and plastid (bottom) phylogeny scaled to the same substitution rate. Gray shaded areas in Figures 1A to C, presumed position of the root. In a rooted phylogeny inferred from a concatenated data set of nuclear (ITS2 excluded), nucleomorph and plastid DNA sequences with Guillardia theta as an outgroup, the root inserted between clade NoPyr and all other taxa (see Additional file 4). Evolutionary models were chosen according to the results of the Akaike information criterion in Modeltest (see Additional file 1 and Methods). Support values from left to right, maximum likelihood bootstrap/maximum parsimony bootstrap/distance (neighbor-joining) bootstrap/posterior probabilities (Figures 1A and B) or maximum likelihood bootstrap/maximum parsimony bootstrap/distance (neighbor-joining) bootstrap/logdet transformation bootstrap/posterior probabilities (Figure 1C). Cbo, Cryptomonas borealis; Ccu, C. curvata; Cgy, C. gyropyrenoidosa; Clu, C. lundii; Cma, C. marssonii; Cov, C. ovata; Cpa, C. paramaecium (colorless); Cpy, C. pyrenoidifera; Cte, C. tetrapyrenoidosa; blue, taxa of clade LB; red branches and strain designations, loss of photosynthesis; scale bars, substitutions per site.

Figure 2

Chart diagram displaying genetic divergences among the taxa and across the three data sets. A strain from a clade with inconspicuous branch lengths in all three phylogenies, Cryptomonas pyrenoidifera strain M1077, was chosen as a reference. The distance values represent the genetic divergences of strain M1077 to the other taxa. The distance values were extracted from the maximum likelihood distance matrices used otherwise by Paup to infer the neighbor-joining trees during phylogenetic analyses, and fed into a spread-sheet program. Strains CCMP 152, CCAC 0031 and M2180 were genetically identical to strains M1077, CCAP 979/46 and CCAC 0056, respectively, thus, were omitted from the chart diagram. Nucleus, concatenated nuclear SSU rDNA, ITS2 and partial LSU rDNA; nucleomorph, nucleomorph SSU rDNA; plastid, rbcL gene. Taxon designations (abscissa): py, C. pyrenoidifera CCAP 979/61; ma1, C. marssonii CCAC 0086; ma2, C. marssonii CCAC 0103; cu1, C. curvata CCAC 0006; cu2, C. curvata CCAC 0080; te1, C. tetrapyrenoidosa M1092; te2, C. tetrapyrenoidosa NIES 279; ov1, C. ovata CCAC 0064; ov2, C. ovata M1171; NP1, NoPyr strain CCAP 979/46; NP2, NoPyr strain CCAC 0109; NP3, NoPyr strain M0741; gy, C. gyropyrenoidosa CCAC 0108; lu, C. lundii CCAC 0107; bo1, C. borealis CCAC 0113; bo2, C. borealis SCCAP K-0063; pa1, C. paramaecium M2452; pa2, C. paramaecium CCAP 977/1; pa3, C. paramaecium CCAC 0056.