Abstract
Most nucleus-encoded chloroplast proteins rely on an N-terminal transit peptide (TP) as a post-translational sorting signal for directing them to the organelle. Although Toc159 is known to be a receptor for specific preprotein TPs at the chloroplast surface, the mechanism for its own targeting and integration into the chloroplast outer membrane is not completely understood. In a previous study, we identified a novel TP-like sorting signal at the C-terminus (CT) of a Toc159 homolog from the single-cell C4 species, Bienertia sinuspersici. In the current study, we have extended our understanding of the sorting signal using transient expression of fluorescently-tagged fusion proteins of variable-length, and with truncated and swapped versions of the CT. As was shown in the earlier study, the 56 residues of the CT contain crucial sorting information for reversible interaction of the receptor with the chloroplast envelope. Extension of this region to 100 residues in the current study stabilized the interaction via membrane integration, as demonstrated by more prominent plastid-associated signals and resistance of the fusion protein to alkaline extraction. Despite a high degree of sequence similarity, the plastid localization signals of the equivalent CT regions of Arabidopsis thaliana Toc159 homologs were not as strong as that of the B. sinuspersici counterparts. Together with computational and circular dichroism analyses of the CT domain structures, our data provide insights into the critical elements of the CT for the efficient targeting and anchorage of Toc159 receptors to the dimorphic chloroplasts in the single-cell C4 species.
Highlights
In plant cells, chloroplasts are one of the many types of plastids, which play crucial roles in photosynthesis and other metabolic pathways including amino acid and lipid synthesis, and nitrogen and sulfur assimilation (Keeling, 2004)
Our previous enhanced green fluorescent protein (EGFP)-fusion experiments were based on the ChloroP-predicted transit peptide (TP)-like region at the CT of BsToc159 plus five additional residues, which successfully directed the reversible association of the passenger proteins with the outer envelope of chloroplasts (Lung and Chuong, 2012)
We sought to further elucidate the functional region of the novel TP-like sorting signal used by BsToc159 and identify the essential region which mediates the successful integration of the receptor into the outer envelope membrane of chloroplasts
Summary
Chloroplasts are one of the many types of plastids, which play crucial roles in photosynthesis and other metabolic pathways including amino acid and lipid synthesis, and nitrogen and sulfur assimilation (Keeling, 2004). Assembly of the correct plastid proteome is crucial for proper functioning of plants and their responses to developmental and external cues. The targeting and translocation processes are facilitated by information embedded within the N-terminal sequences of preproteins, known as transit peptides (TPs). Preprotein translocation across the envelope is mediated by the coordinate action of two multiprotein complexes, commonly known as the Translocon at the outer envelope membrane of chloroplasts (Toc) and the Translocon at the inner envelope membrane of chloroplasts (Tic)
Sign-in/Register to view highlights & summary 
Published Version (
Free)
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call