Transit Peptides From Photosynthesis-Related Proteins Mediate Import of a Marker Protein Into Different Plastid Types and Within Different Species.

Álvaro Eseverri,Luis M Rubio,Elena Caro,Vicente Medina,Can Baysal,Paul Christou,Teresa Capell

doi:10.3389/fpls.2020.560701

Abstract

Nucleus-encoded plastid proteins are synthesized as precursors with N-terminal targeting signals called transit peptides (TPs), which mediate interactions with the translocon complexes at the outer (TOC) and inner (TIC) plastid membranes. These complexes exist in multiple isoforms in higher plants and show differential specificity and tissue abundance. While some show specificity for photosynthesis-related precursor proteins, others distinctly recognize nonphotosynthetic and housekeeping precursor proteins. Here we used TPs from four Arabidopsis thaliana proteins, three related to photosynthesis (chlorophyll a/b binding protein, Rubisco activase) and photo-protection (tocopherol cyclase) and one involved in the assimilation of ammonium into amino-acids, and whose expression is most abundant in the root (ferredoxin dependent glutamate synthase 2), to determine whether they were able to mediate import of a nuclear-encoded marker protein into plastids of different tissues of a dicot and a monocot species. In A. thaliana, import and processing efficiency was high in all cases, while TP from the rice Rubisco small chain 1, drove very low import in Arabidopsis tissues. Noteworthy, our results show that Arabidopsis photosynthesis TPs also mediate plastid import in rice callus, and in leaf and root tissues with almost a 100% efficiency, providing new biotechnological tools for crop improvement strategies based on recombinant protein accumulation in plastids by the expression of nuclear-encoded transgenes.

Highlights

Plastids are double-membrane organelles found within plants and algae cells
Here we selected three of these transit peptides (TPs) from Arabidopsis proteins directly involved in photosynthesis (CAB6, chlorophyll a/b binding protein; RCA, Rubisco activase) or photo-protection (TOCC, tocopherol cyclase) and one involved in the assimilation of ammonium into amino-acids (GLTB2, ferredoxin dependent glutamate synthase 2) (Figure 1A; Supplementary Figure 2)
CAB6, RCA and TOCC are highly expressed in green tissue (Figure 1B, left panel), and accumulate preferentially in leaves (Figure 1B, right panel), while GLTB2 expression and protein accumulation are most abundant in roots, as evidenced by data from the Plant eFP browser (Waese et al, 2017) and AtProteome (Baerenfaller et al, 2008) databases

Summary

Introduction

Plastids are double-membrane organelles found within plants and algae cells. Phylogenetic analyses show that plastids originated from endosymbiosis of a cyanobacterial ancestor (Yoon et al, 2004). Plastid genomes of land plants have suffered huge reduction by transferring genes to the nuclear genome (Kleine et al, 2009). Such transfer underscores the importance of differentially expressing nuclear genes encoding plastid proteins according to tissue and developmental stage (Kleffmann et al, 2004) and the regulation of the import of proteins from the cytosol (Jarvis and López-Juez, 2013; Chu and Li, 2018; Chu et al, 2020), as these processes determine plastid biogenesis and plant development. It seems that TPs adopt alpha-helical structures in membrane-mimetic environments and that this structure might play a role in TP recognition (Bruce, 2001)

Methods

Results

Conclusion