The plastid is an organelle vital to all photosynthetic and some non-photosynthetic eukaryotes. In the model plant Arabidopsis thaliana, a number of nuclear genes encoding plastid proteins have been found to be necessary for embryo development. However, the exact roles of plastids in this process remain largely unknown. Here we use publicly available datasets to obtain insights into the relevance of plastid activities to A. thaliana embryogenesis. By searching the SeedGenes database (http://www.seedgenes.org) and recent literature, we found that, of the 339 non-redundant genes required for proper embryo formation, 108 genes likely encode plastid-targeted proteins. Nineteen of these genes are necessary for development of preglobular embryos and/or their conversion to globular embryos, of which 13 genes encode proteins involved in non-photosynthetic metabolism. By contrast, among 38 genes which are dispensable for globular embryo formation but necessary for further development, only one codes for a protein involved in metabolism. Products of 21 of the 38 genes play roles in plastid gene expression and maintenance. Examination of RNA profiles of embryos at distinct growth stages obtained in laser-capture microdissection coupled with DNA microarray experiments revealed that most of the identified genes are expressed throughout embryo morphogenesis and maturation. These findings suggest that metabolic activities are required at preglobular and throughout all stages of embryo development, whereas plastid gene expression becomes necessary during and/or after the globular stage to sustain various activities of the organelle including photosynthetic electron transport.

Thylakoidal processing peptidase (TPP) is responsible for removing amino-terminal thylakoid-transfer signals from several proteins in the thylakoid lumen. Three TPP isoforms are encoded by the nuclear genome of Arabidopsis thaliana. Previous studies showed that one of them termed plastidic type I signal peptidase 1 (Plsp1) was necessary for processing three thylakoidal proteins and one protein in the chloroplast envelope in vivo. The lack of Plsp1 resulted in seedling lethality, apparently due to disruption of proper thylakoid development. The physiological roles of the other two TPP homologs remain unknown. Here we show that the three A. thaliana TPP isoforms evolved to acquire diverse functions. Phylogenetic analysis revealed that TPP may have originated before the endosymbiotic event, and that there are two groups of TPP in seed plants: one includes Plsp1 and another comprises the other two A. thaliana TPP homologs, which are named as Plsp2A and Plsp2B in this study. The duplication leading to the two groups predates the gymnosperm-angiosperm divergence, and the separation of Plsp2A and Plsp2B occurred after the Malvaceae-Brassicaceae diversification. Quantitative reverse transcription-PCR assay revealed that the two PLSP2 genes were co-expressed in both photosynthetic tissues and roots, whereas the PLSP1 transcript accumulated predominantly in photosynthetic tissues. Both PLSP2 genes were expressed in the aerial parts of the plsp1-null mutant at levels comparable to those in wild-type plants. The seedling-lethal phenotype of the plsp1-null mutant could be rescued by a constitutive expression of Plsp1 cDNA but not by that of Plsp2A or Plsp2B. These results indicate that Plsp1 and Plsp2 evolved to function differently, and that neither of the Plsp2 isoforms is necessary for proper thylakoid development in photosynthetic tissues.