Protein import mechanisms of chloroplasts and mitochondria

Chloroplasts and mitochondria are endosymbiotic organelles that function to control energy metabolism in plants. Many lines of evidence support the theory that ancestral eukaryotes acquired these two organelles via endosymbiosis of cyanobacteria and proteobacteria, respectively. Most of the genes in the endosymbionts’ genomes were then transferred to the nucleus of the host over a billion years of evolution. Each of the genomes of Arabidopsis chloroplast and mitochondrion contains over 100 protein-coding genes. Many proteins encoded by nuclear genes have to be transported into these two organelles to maintain their functions. The first entry points on chloroplasts and mitochondria are the TOC (translocase of the outer chloroplast envelope) and TOM (translocase of the outer mitochondrial membrane) complexes on the outer membranes (OM), respectively. Chloroplast targeting signals (transit peptides) and mitochondrial targeting signals (presequences) at N-termini of precursor proteins translated in the cytosol are recognized by receptors on the TOC and TOM, respectively.

A model was proposed for pSSU recognition and TOC translocation. First, the transit peptide of pSSU is phosphorylated by cytosolic STY kinases at Ser34, allowing it to bind to 14-3-3 and HSP70 proteins. Formation of the pSSU/14-3-3/HSP70 complex increased the speed of import of pSSU into the chloroplast compared to free pSSU, possibly by enhancing its binding to the Toc33 and Toc159 receptors of the TOC complexes. Hydrolysis of GTP at Toc33 dissociates pSSU from this complex, and an as yet unknown phosphatase then dephosphorylates the pSer34 allowing it to be imported into chloroplasts via the combined action of Toc159 and Toc75 (Oreb et al. 2011).

Unlike the precursors for chloroplast import, the presequences of the proteins imported to the mitochondria were not known to be phosphorylated. It was believed that phosphorylation and dephosphorylation of presequences of precursor proteins was not involved in the mitochondrial import.

Our studies (Law et al., 2015) showed that:

• The presequences of multiple organellar RNA editing factors pMORF2/3/5/6 are phosphorylated by STY kinases
• Phosphorylation of the presequence of pMORF3 by STY8 kinase enables it to bind to both HSP70 and 14-3-3 proteins in vitro
• The phosphorylation of pMORF3 by STY kinases does not enhance, but rather impedes import into mitochondria.
• AtPAP2 interacts with the presequences of a number of pMORF proteins and import rate of the pMORF3 into pap2 T-DNA mitochondria is slow.Based on our findings, we propose a model to explain how phosphorylation of transit peptide/presequences affect the import of certain precursor proteins into chloroplasts and mitochondria

We also found that STY8, STY17, and STY46 kinases have different substrate specificity. The growth phenotypes of their T-DNA lines are also different (May and Soll 2000). We postulate that differential phosphorylation of the presequences of different preproteins by various kinases may modulate their import rate into mitochondria and thus affect the import priority (Law et al., 2018).

FdC1 and Leaf-Type Ferredoxins Channel Electrons From Photosystem I to Different Downstream Electron Acceptors

Plant-type ferredoxins in Arabidopsis transfer electrons from the photosystem I to multiple redox-driven enzymes involved in the assimilation of carbon, nitrogen, and sulfur. Leaf-type ferredoxins also modulate the switch between the linear and cyclic electron routes of the photosystems. Recently, two novel ferredoxin homologs with extra C-termini were identified in the Arabidopsis genome (AtFdC1, AT4G14890; AtFdC2, AT1G32550). FdC1 was considered as an alternative electron acceptor of PSI under extreme ferredoxin-deficient conditions. Here, we showed that FdC1 could interact with some, but not all, electron acceptors of leaf-type Fds, including the ferredoxin-thioredoxin reductase (FTR), sulfite reductase (SiR), and nitrite reductase (NiR). Photoreduction assay on cytochrome c and enzyme assays confirmed its capability to receive electrons from PSI and donate electrons to the Fd-dependent SiR and NiR but not to the ferredoxin-NADP+ oxidoreductase (FNR). Hence, FdC1 and leaf-type Fds may play differential roles by channeling electrons from photosystem I to different downstream electron acceptors in photosynthetic tissues. In addition, the median redox potential of FdC1 may allow it to receive electrons from FNR in non-photosynthetic plastids.
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