The number of completely sequenced plastid genomes available is growing rapidly. This array of sequences presents new opportunities to perform comparative analyses. In comparative studies, it is often useful to compare across wide phylogenetic spans and, within angiosperms, to include representatives from basally diverging lineages such as the genomes reported here: Nuphar advena (from a basal-most lineage) and Ranunculus macranthus (a basal eudicot). We report these two new plastid genome sequences and make comparisons (within angiosperms, seed plants, or all photosynthetic lineages) to evaluate features such as the status of ycf15 and ycf68 as protein coding genes, the distribution of simple sequence repeats (SSRs) and longer dispersed repeats (SDR), and patterns of nucleotide composition. The Nuphar [GenBank:NC_008788] and Ranunculus [GenBank:NC_008796] plastid genomes share characteristics of gene content and organization with many other chloroplast genomes. Like other plastid genomes, these genomes are A+T-rich, except for rRNA and tRNA genes. Detailed comparisons of Nuphar with Nymphaea, another Nymphaeaceae, show that more than two-thirds of these genomes exhibit at least 95% sequence identity and that most SSRs are shared. In broader comparisons, SSRs vary among genomes in terms of abundance and length and most contain repeat motifs based on A and T nucleotides. SSR and SDR abundance varies by genome and, for SSRs, is proportional to genome size. Long SDRs are rare in the genomes assessed. SSRs occur less frequently than predicted and, although the majority of the repeat motifs do include A and T nucleotides, the A+T bias in SSRs is less than that predicted from the underlying genomic nucleotide composition. In codon usage third positions show an A+T bias, however variation in codon usage does not correlate with differences in A+T-richness. Thus, although plastome nucleotide composition shows "A+T richness", an A+T bias is not apparent upon more in-depth analysis, at least in these aspects. The pattern of evolution in the sequences identified as ycf15 and ycf68 is not consistent with them being protein-coding genes. In fact, these regions show no evidence of sequence conservation beyond what is normal for non-coding regions of the IR.

/pdf/comparative-chloroplast-genomics-analyses-including-new-t3ad60y9an.pdf

Comparative chloroplast genomics: Analyses including new sequences from the angiosperms Nuphar advena and Ranunculus macranthus

The field of comparative plastid genomics has burgeoned during the past decade, largely due to the availability of rapid, less expensive genome sequencing technologies. Currently there are 200 plastid genomes (plastomes) publicly available with 65% of these from seed plants. Comparative analyses have demonstrated that there is an overall stability in plastome architecture, gene and intron content, and gene order across seed plants. However, a number of unrelated lineages of both gymnosperms and angiosperms do not follow this pattern and have experienced considerable genomic upheaval. Within angiosperms these lineages with highly rearranged plastomes exhibit three other phenomena: highly accelerated rates of nucleotide substitutions, an increase in the number of dispersed repeats, many of which are associated with rearrangement endpoints, and biparental plastid inheritance. The correlation between genomic upheaval and these other phenomena suggest that aberrant DNA repair mechanisms may be involved in destabilizing these plastid genomes. Experimental studies support this idea because knocking out DNA repair genes destabilizes plastomes. Further studies of nuclear-plastid interactions, especially in seed plant lineages with highly rearranged plastomes, are needed to clarify the causes of the plastome instability. The large number of plastome sequences has also provided valuable data for resolving phylogenetic relationships among seed plants. This is especially true for angiosperms where these data have been instrumental in clarifying relationships among the early diverging clades, an endeavor that had stymied plant biologists for over a decade. The most recent plastome phylogenies clearly identify Amborella as the earliest diverging lineage of flowering plants and provide strong support for the position of magnoliids as sister to a large clade that includes eudicots and monocots. This robust phylogenetic estimate provides an evolutionary framework for examining patterns and rates of change in plastid genomes across angiosperms.

Plastid Genomes of Seed Plants

Production of heterologous proteins in plants: strategies for optimal expression.

Towards high-yield production of pharmaceutical proteins with plant cell suspension cultures

The molecular era has fundamentally reshaped our knowledge of the evolution and diversification of angiosperms. One outstanding question is the phylogenetic placement of Amborella trichopoda Baill., commonly thought to represent the first lineage of extant angiosperms. Here, we leverage publicly available data and provide a broad coalescent-based species tree estimation of 45 seed plants. By incorporating 310 nuclear genes, our coalescent analyses strongly support a clade containing Amborella plus water lilies (i.e., Nymphaeales) that is sister to all other angiosperms across different nucleotide rate partitions. Our results also show that commonly applied concatenation methods produce strongly supported, but incongruent placements of Amborella: slow-evolving nucleotide sites corroborate results from coalescent analyses, whereas fast-evolving sites place Amborella alone as the first lineage of extant angiosperms. We further explored the performance of coalescent versus concatenation methods using nucleotide sequences simulated on (i) the two alternate placements of Amborella with branch lengths and substitution model parameters estimated from each of the 310 nuclear genes and (ii) three hypothetical species trees that are topologically identical except with respect to the degree of deep coalescence and branch lengths. Our results collectively suggest that the Amborella alone placement inferred using concatenation methods is likely misled by fast-evolving sites. This appears to be exacerbated by the combination of long branches in stem group angiosperms, Amborella, and Nymphaeales with the short internal branch separating Amborella and Nymphaeales. In contrast, coalescent methods appear to be more robust to elevated substitution rates.

/pdf/coalescent-versus-concatenation-methods-and-the-placement-of-3jc2zhvni6.pdf

Coalescent versus Concatenation Methods and the Placement of Amborella as Sister to Water Lilies

A detailed comparison was made of codon usage of chloroplast genes with their host (nuclear) genes in the four angiosperm species Oryza sativa, Zea mays, Triticum aestivum and Arabidopsis thaliana. The average GC content of the entire genes, and at the three codon positions individually, was higher in nuclear than in chloroplast genes, suggesting different genomic organization and mutation pressures in nuclear and chloroplast genes. The results of Nc-plots and neutrality plots suggested that nucleotide compositional constraint had a large contribution to codon usage bias of nuclear genes in O. sativa, Z. mays, and T. aestivum, whereas natural selection was likely to be playing a large role in codon usage bias in chloroplast genomes. Correspondence analysis and chi-test showed that regardless of the genomic environment (species) of the host, the codon usage pattern of chloroplast genes differed from nuclear genes of their host species by their AU-richness. All the chloroplast genomes have predominantly A- and/or U-ending codons, whereas nuclear genomes have G-, C- or U-ending codons as their optimal codons. These findings suggest that the chloroplast genome might display particular characteristics of codon usage that are different from its host nuclear genome. However, one feature common to both chloroplast and nuclear genomes in this study was that pyrimidines were found more frequently than purines at the synonymous codon position of optimal codons.

Comparative studies on codon usage pattern of chloroplasts and their host nuclear genes in four plant species.

Plants are often threatened by pathogen attack or other external stresses during growth and development. To decrease loss in productivity, plants respond to biotic and abiotic stresses by triggering a series of biochemical reactions, termed hypersensitive reaction (HR), which is associated with synthesis of secondary metabolites, cell wall rigidification, lignin and suberin deposition, and induction of a variety of novel proteins. In the HR response, pathogenesis-related proteins (PRs) that are suggested to be effective in inhibiting growth, multiplication and/or spread of pathogens, and are responsible for the state of systemic acquired resistance (SAR; Van Loon 1997), are significantly induced and accumulated abundantly in host plants (Van Loon and Van Strien 1999). PR proteins can be divided into 17 groups on the basis of their sequence characteristics, immunological relationships and biological activities (Liu et al. 2005). However, only 14 distinct families of PR proteins have been found in plants (Sarowar et al. 2005). Not all families are found in any one plant species (Van Loon and Van Strien 1999). Among the diverse PRs, PR-1 is the dominant group, whose function is not well known. The limited antifungal activity suggests function of PR-1 in plant defence (Kitajima and Sato 1999). However, the mode of action and its relationship to other types of proteins still remain unknown. The first PR1 protein was discovered in 1970. Since then, a number of PR-1 proteins have been identified in Arabidopsis (Uknes et al. 1992), maize (Morris et al. 1998), wheat (Molina et al. 1999), tomato (Tornero et al. 1994), tobacco (Pfitzner and Goodman 1987), barley (Muradov et al. 1993), rice (Agrawal et al. 2000a,b; Kim et al. 2001) and pepper (Kim and Hwang 2000). Rice is a vitally important crop for human consumption. The completion of the Oryza sativa genome

Computational identification of novel PR-1-type genes in Oryza sativa.

The initiation of protein biosynthesis is a major determi-nant of the efficiency of gene expression at the transla-tional level. As indicated in many studies, the nucleotide sequences around the AUG translation initiation codon (AUG co ntext) act as an important signal to trigger the initiation of translation event (Kozak 1987; Miyasaka et al. 2002). It is also known that the occurrence of a purine at the –3 and a G at the +4 position is the most impo rtant feature of the AUG conte xt in eukaryotes, and the strong preference for G at the +4 position promotes14933 for the process of recognition of the actual initiation codon by the ribosome subunits (Kozak 1997, 1999). However, there is presently little knowledge about the sequence cha ra-cteri stics around – especially before – the initiator codon, and their effects on translation initiation efficiency. Recen -tly, the whole genome sequences of Arab idopsis thaliana , Homo sapiens, Drosophila melanogaster (Kaul et al. 2000; Adams et al. 2000; Venter et al. 2001), and the draft genome sequence of Oryza sativa have been deter-mined (Yu et al. 2002; Goff et al. 2002). Therefore, it is of interest to determine whether hitherto unknown se-quence elements might affect translation initiation in the genes of these eukaryotes. We have done this through a systematic analysis of codon and nucleotides biases around the AUG codon in these four species (a monocot, dicot, invertebrate and vertebrate). Moreover, we also examined the relationship between the conservation of nucleotides around the initiator codon and gene expre ssion level.

Qingpo Liu

Papers

Comparative studies on codon usage pattern of chloroplasts and their host nuclear genes in four plant species.

Computational identification of novel PR-1-type genes in Oryza sativa.

Comparative studies on sequence characteristics around translation initiation codon in four eukaryotes.