Molecular Mechanism of the Specificity of Protein Import into Chloroplasts and Mitochondria in Plant Cells

Abstract

Plants possess both types of endosymbiotic organelles, chloroplasts and mitochondria. Transit peptides and presequences function as signal sequences for specific import into chloroplasts and mitochondria, respectively. However, how these highly similar signal sequences confer the protein import specificity remains elusive. Here, we show that mitochondrial-or chloroplast-specific import involves two distinct steps, specificity determination and translocation across envelopes, which are mediated by the N-terminal regions and functionally interchangeable C-terminal regions, respectively, of transit peptides and presequences. A domain harboring multiple-arginine and hydrophobic sequence motifs in the N-terminal regions of presequences was identified as the mitochondrial specificity factor. The presence of this domain and the absence of arginine residues in the N-terminal regions of otherwise common targeting signals confers specificity of protein import into mitochondria and chloroplasts, respectively. AtToc159, a chloroplast import receptor, also contributes to determining chloroplast import specificity. We propose that common ancestral sequences were functionalized into mitochondrial-and chloroplast-specific signal sequences by the presence and absence, respectively, of multiple-arginine and hydrophobic sequence motifs in the N-terminal region.

Cross-Species Functional Conservation and Possible Origin of the N-Terminal Specificity Domain of Mitochondrial Presequences

“…Thus, plant cells should have distinct mechanisms for protein targeting to the chloroplast and mitochondria (Schleiff and Becker, 2011;Garg and Gould, 2016). Indeed, we recently found that chloroplast transit peptides and presequences consist of two functionally different domains in plant cells (Lee et al, 2019). The N-terminal specificity domain confers import specificity, whereas the C-terminal domain is interchangeable between the transit peptide and presequence, and functions in the translocation of organellar proteins across the envelope membranes (Lee et al, 2019).…”

“…Indeed, we recently found that chloroplast transit peptides and presequences consist of two functionally different domains in plant cells (Lee et al, 2019). The N-terminal specificity domain confers import specificity, whereas the C-terminal domain is interchangeable between the transit peptide and presequence, and functions in the translocation of organellar proteins across the envelope membranes (Lee et al, 2019). The N-terminal specificity domain of the mitochondrial presequence contains two sequence elements: multiple arginine residues and a hydrophobic sequence motif (Abe et al, 2000;Lee et al, 2019).…”

“…The N-terminal specificity domain confers import specificity, whereas the C-terminal domain is interchangeable between the transit peptide and presequence, and functions in the translocation of organellar proteins across the envelope membranes (Lee et al, 2019). The N-terminal specificity domain of the mitochondrial presequence contains two sequence elements: multiple arginine residues and a hydrophobic sequence motif (Abe et al, 2000;Lee et al, 2019). Arginine residues play a crucial role in another signal sequence known as the twin-arginine translocation (TAT) signal sequence (Alami et al, 2002;Cline, 2015).…”

“…Arginine residues play a crucial role in another signal sequence known as the twin-arginine translocation (TAT) signal sequence (Alami et al, 2002;Cline, 2015). Similar to the arginines in the TAT signal sequence, arginine residues in presequences cannot be functionally replaced with another positively charged residue, lysine (Lee et al, 2019). In the TAT signal sequence, another important sequence motif is the hydrophobic (h-) region downstream of the conserved twinarginine (RR) motif (Ulfig et al, 2017).…”

Major changes in plastid protein import and the origin of the Chloroplastida

“…The most N-terminal domain carries the charged residues critical for distinguishing between mitochondrial- and plastid targeting (Fig. 1), while the C-terminus is exchangeable 84 . Because the plastid is younger and because the photosynthetic organelle evolved in a eukaryotic cell instead of contributing to its actual origin, we understand more about the origin of the plastid NTS.…”