Chloroplasts are subcellular organelles that provide plants with abundant metabolic capabilities, most notably the ability to capture carbon from atmospheric carbon dioxide through the process of photosynthesis. It has been proposed1 that the plant kingdom began to evolve when a bacterium, similar to a present-day photosynthetic cyanobacterium, was engulfed by a host cell, and from this bacterial ancestor, chloroplasts eventually arose in the cellular descendants of the host cell. During chloroplast evolution, thousands of genes transferred from the intracellular bacterial genome to the host genome. However, these relocated genes encode proteins that need to be targeted to their site of function within the chloroplast. Writing in Nature, Chen et al.2 report the identification of a component of the system that imports proteins into chloroplasts. Their finding illuminates how this evolved, and also provides mechanistic insight into how import is co-ordinated across the two membrane layers that form the chloroplast’s outer envelope.
Most chloroplast proteins are made in the cytoplasm. They contain specific amino-acid sequences, termed transit peptides, that are used to direct these proteins from the cytoplasm, across the two membrane layers of the chloroplast and into the interior of the organelle3. Chen and colleagues’ work addresses some key questions regarding this protein-import system. The first is, how do the multi-protein complexes, found at the outer and inner membranes of the chloroplast envelope (termed TOC and TIC, respectively) mediate transport in a coordinated way that prevents the mistargeting or misfolding of proteins as they transit through the intermembrane space? Under normal conditions, protein import across the outer and inner membranes seems to occur essentially simultaneously through the TOC and TIC complexes4. However, whether there is a physical connection between TOC and TIC, and if so, what its nature is, has been a mystery.
Chen et al. report the identification of a previously unknown component of the TIC complex, a protein that they name TIC236, which acts as a link between TIC and TOC. The authors discovered TIC236 using a biochemical approach to identify proteins that are associated with TOC components. TIC236 is anchored in the inner membrane, where it interacts with components of the TIC complex. Part of TIC236 extends into the inter-membrane space, where it interacts with TOC75, a membrane protein that forms part of the channel in the TOC complex (Fig. 1a).
The authors report that, in the plant Arabidopsis thaliana, mutations that block the expression of TIC236 are lethal, and mutations that impair TIC236 function reduce the rates of protein import into chloroplasts compared to the import rate in wild-type A. thaliana. These results, in addition to the authors’ studies of protein–protein interactions, provide compelling evidence that TIC236 provides a key physical link between TIC and TOC. Chen and colleagues conducted a phylo-genetic analysis that provides evidence for the co-evolution of the interacting domains of TOC75 and TIC236, supporting their hypothesis that these proteins evolved as components of interacting complexes. The ability to couple transit through TOC and TIC offers a way of ensuring efficient protein import. This is probably crucial during seedling development, when the rate of protein import into chloroplasts is high, and more than half of the total protein in a cell can be within the chloroplasts5.
The second key question this work addresses is, how did the TOC and TIC import systems evolve? Evidence for analogous systems that allow protein import into bacteria is lacking6, and, consequently, the evolutionary origin of the chloroplast protein-import system has been an open question. TOC75 is related to the OMP85 family of membrane proteins found on the outer membranes of chloroplasts, energy-generating organelles called mitochondria, and in Gram-negative bacteria, a group that includes the photo-synthetic cyano-bacteria7. Gram-negative bacteria have two membrane layers, and membrane proteins on their surface can be assembled and transported to the cell exterior with the help of protein complexes called TAM or BAM on the outer membrane of the cell. Membrane proteins in these complexes are members of the OMP85 family7. It has been proposed8 that TOC75 is derived from an ancestral protein related to components of the BAM or TAM systems in the ancestral bacterium that gave rise to the chloroplast.
Chen and colleagues demonstrate that TIC236 is related to TamB, which aids protein transport9 between bacterial membrane proteins that form a secretion system (called Sec), located on the inner membrane, and BAM or TAM components in the outer membrane (Fig. 1b). It therefore seems that a mechanism for coupling inner- and outer-membrane transport in chloroplasts has been evolutionarily conserved from an ancestral bacterial system.
However, despite this conservation, the chloroplast protein-import system has evolved to function in the reverse direction relative to the direction of transport in the bacterial export system5. The BAM and TAM complexes facilitate export of bacterial proteins from the cytoplasm to the outer membrane, whereas the TOC and TIC complexes import proteins from outside the chloroplast to inside it. This remarkable reversal of the direction of protein transport probably resulted from the gain of other TOC or TIC proteins that evolved from host-encoded genes to adapt the complexes for the purposes of protein import. These include TOC and TIC receptors and molecular motor proteins known to facilitate transport into the chloroplast3.
Chen and colleagues’ results provide convincing evidence for the origin of key elements of the chloroplast protein-import system from an ancestral bacterial protein-export system. Their insights also reveal the adaptation and consequent reversal of an existing protein-targeting pathway that was essential for the ancestral bacteria to successfully take up residence in a host cell, thereby enabling the host to take advantage of its guest’s photosynthetic and metabolic capabilities.
Nature 564, 45-46 (2018)