Erik Richly

Evolutionary analysis of Arabidopsis, cyanobacterial, and chloroplast genomes reveals plastid phylogeny and thousands of cyanobacterial genes in the nucleus

Chloroplasts were once free-living cyanobacteria that became endosymbionts, but the genomes of contemporary plastids encode only ≈5–10% as many genes as those of their free-living cousins, indicating that many genes were either lost from plastids or transferred to the nucleus during the course of plant evolution. Previous estimates have suggested that between 800 and perhaps as many as 2,000 genes in the Arabidopsis genome might come from cyanobacteria, but genome-wide phylogenetic surveys that could provide direct estimates of this number are lacking. We compared 24,990 proteins encoded in the Arabidopsis genome to the proteins from three cyanobacterial genomes, 16 other prokaryotic reference genomes, and yeast. Of 9,368 Arabidopsis proteins sufficiently conserved for primary sequence comparison, 866 detected homologues only among cyanobacteria and 834 other branched with cyanobacterial homologues in phylogenetic trees. Extrapolating from these conserved proteins to the whole genome, the data suggest that ≈4,500 of Arabidopsis protein-coding genes (≈18% of the total) were acquired from the cyanobacterial ancestor of plastids. These proteins encompass all functional classes, and the majority of them are targeted to cell compartments other than the chloroplast. Analysis of 15 sequenced chloroplast genomes revealed 117 nuclear-encoded proteins that are also still present in at least one chloroplast genome. A phylogeny of chloroplast genomes inferred from 41 proteins and 8,303 amino acids sites indicates that at least two independent secondary endosymbiotic events have occurred involving red algae and that amino acid composition bias in chloroplast proteins strongly affects plastid genome phylogeny.

Covariations in the nuclear chloroplast transcriptome reveal a regulatory master-switch

The evolution of the endosymbiotic progenitor into the chloroplast organelle was associated with the transfer of numerous chloroplast genes into the nucleus. Hence, inter‐organellar signalling, and the co‐ordinated expression of sets of nuclear genes, was set up to control the metabolic and developmental status of the chloroplast. Here, we show by the differential‐expression analysis of 3,292 genes, that most of the 35 environmental and genetic conditions tested, including plastid signalling mutations, elicit only three main classes of response from the nuclear chloroplast transcriptome. Two classes, probably involving GUN (genomes uncoupled)‐type plastid signalling, are characterized by alterations, in opposite directions, in the expression of largely overlapping sets of genes.

Evolutionary diversification of mitochondrial proteomes: implications for human disease

By combining comparative genomics and computational identification of protein targeting, we predicted the size and composition of the mitochondrial proteome for ten species. Functional mitochondria could harbour from a few hundred to more than 3000 gene products, and protein relocation from and to mitochondria occurred during evolution. Although each genome studied contains lineage-specific mitochondrial proteins, conserved mitochondrial proteins also exist. Their functions mostly relate to transport and metabolism, and in humans their mutated states are frequently associated with disease.

Gene-sequence-tag expression analyses of 1,800 genes related to chloroplast functions

Abstract. Quantification of the expression levels of nuclear genes encoding plastid proteins under different genetic or environmental conditions can contribute to the genetic dissection of plastid functions. To facilitate such measurements, a set of 1,827 Arabidopsis thaliana genes coding for plastid proteins was PCR-amplified from genomic DNA and spotted on nylon membranes to generate an array of chloroplast-specific gene-sequence-tags (GSTs). The sensitivity and reliability of the experimental system was evaluated and a procedure was developed for detecting differential gene expression. The GST array was found to serve as a reliable monitor of changes in gene expression induced by environmental and genetic alteration of chloroplast functions. Based on comparisons of dark- versus light-grown seedlings, and wild-type versus prpl11-1 plants, lists of differentially expressed genes are provided which include 193/7 and 25/42 up/down-regulated genes, respectively. The cut-off values for differential expression were 2.5-times (up) and 0.40 (down). Additional up-regulated genes with relatively low expression ratios (from 1.5- to 2.5-times) or down-regulated with relatively high ratios (0.4–0.67) can be accessed at the website: http://www.mpiz-koeln.mpg.de/~richly/GST-array.html. A sample of genes analysed by quantitative reverse transcription PCR confirmed the expression profiles monitored by the GST array. Differential hybridisation experiments with the prpl11-1 mutant revealed the existence of regulatory networks sensing the protein state of the chloroplast and transmitting the signal to the nucleus.

Functional genomics of Arabidopsis photosynthesis

The study of photosynthesis in higher plants will benefit significantly from the sequencing of the entire genome of Arabidopsis thaliana and from the recent development of new genomic technologies. For large sets of genes, it is now possible to identify their functions and levels of expression, and the post-translational modifications and interactions of their products. This functional genomic approach to photosynthesis is summarised here by considering the application of knock-out mutant screens (forward genetics), the identification of mutant alleles of genes whose sequence is known (reverse genetics), the use of microarrays to detect gene expression at genome level, the separation of proteins by two-dimensional electrophoresis, and their identification by mass spectrometry and bioinformatics. This emerging, integrated approach to the elucidation of gene function is presented and its potential is discussed.

Protein-protein and protein-function relationships in Arabidopsis photosystem I: cluster analysis of PSI polypeptide levels and photosynthetic parameters in PSI mutants.

In flowering plants, photosystem I (PSI) mediates electron transport across the thylakoid membrane and contains at least 14 proteins. The availability of co-suppression and/or mutant lines deficient for individual PSI polypeptides in Arabidopsis thaliana allows one to assign functions to PSI subunits. We have performed cluster analysis on an extensive set of data on PSI polypeptide levels in ten different PSI mutants. This type of analysis serves to group proteins that exhibit similar changes in amount in different genotypes, and also identifies genotypes which show similar PSI compositions. The interdependence of levels of PSI-C, -D and -E, of -H and -L, and of Lhca2 and 3, which was previously proposed based on the study of single genotypes or on cross-linking experiments, was confirmed by our analyses. In addition, the levels of the lumenal subunits F and N are found to be interdependent. The incorporation of photosynthetic parameters into the cluster analysis revealed that the level of photosynthetic state transitions correlates with the abundance of PSI-H in all 8 genotypes tested, supporting the hypothesis that PSI-H serves as a docking site for LHCII during state transitions.