An outer envelope membrane component of the plastid protein import apparatus plays an essential role in Arabidopsis

Translocon at the outer envelope membrane of chloroplasts, 34 kDa (Toc34) is a GTP-binding component of the protein import apparatus within the outer envelope membrane of plastids. The Arabidopsis genome encodes two homologues of Toc34, designated atToc33 and atToc34. In this report, we describe the identification and characterization of two atToc34 knockout mutants, plastid protein import 3-1 (ppi3-1) and ppi3-2. Aerial tissues of the ppi3 mutants appeared similar to the wild type throughout development, and contained structurally normal chloroplasts that were able to efficiently import the Rubisco small subunit precursor (prSS) in vitro. The absence of an obvious ppi3 phenotype in green tissues presumably reflects the ability of atToc33 to substitute for atToc34 in the mutant, and the relatively high level of expression of the atTOC33 gene in these tissues. In the roots, where atTOC33 is expressed at a much lower level, significant growth defects were observed in both mutants: ppi3 roots were approximately 20-30% shorter than wild-type roots. Attempts to identify a double homozygote lacking atToc34 and atToc33 (by crossing the ppi3 mutants with ppi1, an atToc33 knockout mutant) were unsuccessful, indicating that the function provided by atToc33/atToc34 is essential during early development. Plants that were homozygous for ppi1 and heterozygous for ppi3 displayed a chlorotic phenotype much more severe than that of the ppi1 single mutant. Furthermore, the siliques of these plants contained approximately 25% aborted seeds, indicating that the double homozygous mutation is embryo lethal. The data demonstrate that atToc33/atToc34 performs a central and essential role during plastid protein import, and indicate that the atToc34 isoform is relatively more important for plastid biogenesis in roots.