Chloroplast DNA sequence of the green alga Oedogonium cardiacum (Chlorophyceae): unique genome architecture, derived characters shared with the Chaetophorales and novel genes acquired through horizontal transfer

Abstract

Background: To gain insight into the branching order of the five main lineages currently recognized in the green algal class Chlorophyceae and to expand our understanding of chloroplast genome evolution, we have undertaken the sequencing of chloroplast DNA (cpDNA) from representative taxa. The complete cpDNA sequences previously reported for Chlamydomonas (Chlamydomonadales), Scenedesmus (Sphaeropleales), and Stigeoclonium (Chaetophorales) revealed tremendous variability in their architecture, the retention of only few ancestral gene clusters, and derived clusters shared by Chlamydomonas and Scenedesmus. Unexpectedly, our recent phylogenies inferred from these cpDNAs and the partial sequences of three other chlorophycean cpDNAs disclosed two major clades, one uniting the Chlamydomonadales and Sphaeropleales (CS clade) and the other uniting the Oedogoniales, Chaetophorales and Chaetopeltidales (OCC clade). Although molecular signatures provided strong support for this dichotomy and for the branching of the Oedogoniales as the earliest-diverging lineage of the OCC clade, more data are required to validate these phylogenies. We describe here the complete cpDNA sequence of Oedogonium cardiacum (Oedogoniales).

Results: Like its three chlorophycean homologues, the 196,547-bp Oedogonium chloroplast genome displays a distinctive architecture. This genome is one of the most compact among photosynthetic chlorophytes. It has an atypical quadripartite structure, is intron-rich (17 group I and 4 group II introns), and displays 99 different conserved genes and four long open reading frames (ORFs), three of which are clustered in the spacious inverted repeat of 35,493 bp. Intriguingly, two of these ORFs (int and dpoB) revealed high similarities to genes not usually found in cpDNA. At the gene content and gene order levels, the Oedogonium genome most closely resembles its Stigeoclonium counterpart. Characters shared by these chlorophyceans but missing in members of the CS clade include the retention of psaM, rpl32 and trnL(caa), the loss of petA, the disruption of three ancestral clusters and the presence of five derived gene clusters.

Conclusion: The Oedogonium chloroplast genome disclosed additional characters that bolster the evidence for a close alliance between the Oedogoniales and Chaetophorales. Our unprecedented finding of int and dpoB in this cpDNA provides a clear example that novel genes were acquired by the chloroplast genome through horizontal transfers, possibly from a mitochondrial genome donor.

Figure 1

Gene map of Oedogonium cpDNA. Thick lines on the inner circle represent the two copies of the IR (IR_A and IR_B), with the arrows indicating the transcription direction of the IR-encoded rRNA operon. Genes (filled boxes) on the outside of the outer circle are transcribed in a clockwise direction. Introns are represented by open boxes and intron ORFs are denoted by narrow, filled boxes. The rpoB gene consists of two separate ORFs (rpoBa and rpoBb) that are not associated with sequences typical of group I or group II introns. tRNA genes are indicated by the one-letter amino acid code followed by the anticodon in parentheses (Me, elongator methionine; Mf, initiator methionine). Only one of the two isomeric forms of the genome is shown here; these isomers differ with respect to the relative orientation of the SC regions.

Figure 2

Conservation of gene pairs in Oedogonium and other UTC algal cpDNAs. For each gene pair, adjoining termini of the genes are indicated. Filled boxes indicate the presence of gene pairs with the same relative polarities in two or more genomes. Grey or open boxes indicate the absence of gene pairs. A grey box indicates that the two genes associated with a gene pair are found in the genome but are unlinked. An open box indicates that one or both genes associated with a gene pair are absent from the genome. Two horizontal lines at the bottom of the figure denote the gene pairs that are shared specifically by Chlamydomonas and Scenedesmus (CS clade) and by Oedogonium and Stigeoclonium (OCC clade). The gene pairs shared by the prasinophycean Nephroselmis and the streptophyte Mesostigma viride were presumably present in the last common ancestor of all green algae. Chlorella vulgaris belongs to the Trebouxiophyceae, whereas Oltmannsiellopsis and Pseudendoclonium are representatives of the Ulvophyceae.

Figure 3

Distribution of introns in Oedogonium and other UTC algal cpDNAs. Circles denote the presence of group I introns and squares denote the presence of group II introns. Divided squares represent trans-spliced group II introns. Open symbols denote the absence of intron ORFs, whereas filled symbols denote their presence. Intron insertion sites are designated as indicated in Table 4. The column at the extreme right indicates the introns of Chlamydomonas species other than C. reinhardtii that are known to have homologues in completely sequenced UTC algal genomes. References for the latter introns are as follows: psaB [70]; psbA [71]; psbC [70]; rrs [72]; and rrl [73-76]. An asterisk denotes the absence of the ORF in some Chlamydomonas species.

Figure 4

Consensus secondary structure model of the Oedogonium and Stigeoclonium psaC introns. The model is displayed according to Michel et al. [39]. Exon sequences are shown in lowercase letters. Positions exhibiting different nucleotides are denoted by dots. Positions showing deletions/additions are denoted by arrows labelled with two numbers; the left and right numbers refer to the nucleotides found in the Oedogonium and Stigeoclonium introns, respectively. Conserved base pairings are represented by dashes. Roman numerals specify the six major structural domains, whereas uppercase letters in bold denote the helices in domain I. Upper and lower numbers inside the variable loops indicate the sizes of these loops in the Oedogonium and Stigeoclonium introns, respectively. Tertiary interactions are represented by dashed lines, curved arrows, or Greek lettering. Nucleotides that potentially participate in the δ-δ' interaction are boxed. EBS and IBS are exon-binding and intron-binding sites, respectively. The putative site of lariat formation is denoted by an asterisk.