Chloroplast vesicle transport.

Photosynthesis is a well-known process that has been intensively investigated, but less is known about the biogenesis of the thylakoid membrane that harbors the photosynthetic machinery. Thylakoid membranes are constituted by several components, the major ones being proteins and lipids. However, neither of these two are produced in the thylakoid membranes themselves but are targeted there by different mechanisms. The interior of the chloroplast, the stroma, is an aqueous compartment that prevents spontaneous transport of single lipids and/or membrane proteins due to their hydrophobicities. Thylakoid targeted proteins are encoded either in the nucleus or plastid, and thus some cross the envelope membrane before entering one of the identified thylakoid targeting pathways. However, the pathway for all thylakoid proteins is not known. Lipids are produced at the envelope membrane and have been proposed to reach the thylakoid membrane by different means: invaginations of the envelope membrane, direct contact sites between these membranes, or through vesicles. Vesicles have been observed in chloroplasts but not much is yet known about the mechanism or regulation of their formation. The question of whether proteins can also make use of vesicles as one mechanism of transport remains to be answered. Here we discuss the presence of vesicles in chloroplasts and their potential role in transporting lipids and proteins. We additionally discuss what is known about the proteins involved in the vesicle transport and the gaps in knowledge that remain to be filled.

Understanding Plastid Vesicle Transport - Could it Provide Benefit for Human Medicine?

BACKGROUND: In plants, vesicle transport occurs in the secretory pathway in the cytosol, between the membranes of different compartments. Several protein components have been identified to be involved in the process and their functions were characterized. Both cargos and other molecules (such as hormones) have been shown to use vesicle transport, although the major constituents of vesicles are lipids which are transferred from donor to acceptor membranes. In humans, malfunction of the cytosolic vesicle transport system leads to different diseases. METHOD: To better understand and ultimately cure these human diseases, studying other model systems such as yeast can be beneficial. Plants with their cytosolic vesicle transport system could serve as another model system. However, this review focuses on plant vesicles not present in the cytosol but in the chloroplasts, where lipids produced in the surrounding envelope are transported through the aqueous stroma to the thylakoid membranes. Although chloroplast vesicles have found both biochemical and ultrastructural support, only two proteins have been characterized as components of the pathway. However, using bioinformatics a number of other proteins have been suggested as homologs to the cytosolic system. RESULTS & CONCLUSION: Based on these findings vesicles of chloroplasts are likely most similar to the vesicles trafficking from ER to Golgi, or may even be unique, but important experimental support is yet lacking. In this review, proposed vesicle transport components in chloroplasts are presented, and their possible future implementation for human medicine is discussed.

Vesicles Are Persistent Features of Different Plastids.

Peripheral vesicles in plastids have been observed repeatedly, primarily in proplastids and developing chloroplasts, in which they are suggested to function in thylakoid biogenesis. Previous observations of vesicles in mature chloroplasts have mainly concerned low temperature pretreated plants occasionally treated with inhibitors blocking vesicle fusion. Here, we show that such vesicle-like structures occur not only in chloroplasts and proplastids, but also in etioplasts, etio-chloroplasts, leucoplasts, chromoplasts and even transforming desiccoplasts without any specific pretreatment. Observations are made both in C3 and C4 species, in different cell types (meristematic, epidermis, mesophyll, bundle sheath and secretory cells) and different organs (roots, stems, leaves, floral parts and fruits). Until recently not much focus has been given to the idea that vesicle transport in chloroplasts could be mediated by proteins, but recent data suggest that the vesicle system of chloroplasts has similarities with the cytosolic coat protein complex II system. All current data taken together support the idea of an ongoing, active and protein-mediated vesicle transport not only in chloroplasts but also in other plastids, obviously occurring regardless of chemical modifications, temperature and plastid developmental stage.

Proteins affecting thylakoid morphology - the key to understanding vesicle transport in chloroplasts?

We recently showed that a Rab protein, CPRabA5e (CP = chloroplast localized), is located in chloroplasts of Arabidopsis thaliana where it is involved in various processes, such as thylakoid biogenesis and vesicle transport. Using a yeast two-hybrid method, CPRabA5e was shown to interact with a number of chloroplast proteins, including the CURVATURE THYLAKOID 1A (CURT1A) protein and the light-harvesting chlorophyll a/b binding protein (LHCB1.5). CURT1A has recently been shown to modify thylakoid architecture by inducing membrane curvature in grana, whereas LHCB1.5 is a protein of PSII (Photosystem II) facilitating light capture. LHCB1.5 is imported to chloroplasts and transported to thylakoid membranes using the post-translational Signal Recognition Particle (SRP) pathway. With this information as starting point, we here discuss their subsequent protein-protein interactions, given by the literature and Interactome 3D. CURT1A itself and several of the proteins interacting with CURT1A and LHCB1.5 have relations to vesicle transport and thylakoid morphology, which are also characteristics of cprabA5e mutants. This highlights the previous hypothesis of an alternative thylakoid targeting pathway for LHC proteins using vesicles, in addition to the SRP pathway.

Bioinformatic indications that COPI- and clathrin-based transport systems are not present in chloroplasts: an Arabidopsis model.

Coated vesicle transport occurs in the cytosol of yeast, mammals and plants. It consists of three different transport systems, the COPI, COPII and clathrin coated vesicles (CCV), all of which participate in the transfer of proteins and lipids between different cytosolic compartments. There are also indications that chloroplasts have a vesicle transport system. Several putative chloroplast-localized proteins, including CPSAR1 and CPRabA5e with similarities to cytosolic COPII transport-related proteins, were detected in previous experimental and bioinformatics studies. These indications raised the hypothesis that a COPI- and/or CCV-related system may be present in chloroplasts, in addition to a COPII-related system. To test this hypothesis we bioinformatically searched for chloroplast proteins that may have similar functions to known cytosolic COPI and CCV components in the model plants Arabidopsis thaliana and Oryza sativa (subsp. japonica) (rice). We found 29 such proteins, based on domain similarity, in Arabidopsis, and 14 in rice. However, many components could not be identified and among the identified most have assigned roles that are not related to either COPI or CCV transport. We conclude that COPII is probably the only active vesicle system in chloroplasts, at least in the model plants. The evolutionary implications of the findings are discussed.

A novel chloroplast localized Rab GTPase protein CPRabA5e is involved in stress, development, thylakoid biogenesis and vesicle transport in Arabidopsis.

A novel Rab GTPase protein in Arabidopsis thaliana, CPRabA5e (CP = chloroplast localized) is located in chloroplasts and has a role in transport. Transient expression of CPRabA5e:EGFP fusion protein in tobacco (Nicotiana tabacum) leaves, and immunoblotting using Arabidopsis showed localization of CPRabA5e in chloroplasts (stroma and thylakoids). Ypt31/32 in the yeast Saccharomyces cerevisiae are involved in regulating vesicle transport, and CPRabA5e a close homolog of Ypt31/32, restores the growth of the ypt31Δ ypt32(ts) mutant at 37 °C in yeast complementation. Knockout mutants of CPRabA5e displayed delayed seed germination and growth arrest during oxidative stress. Ultrastructural studies revealed that after preincubation at 4 °C mutant chloroplasts contained larger plastoglobules, lower grana, and more vesicles close to the envelopes compared to wild type, and vesicle formation being enhanced under oxidative stress. This indicated altered thylakoid development and organization of the mutants. A yeast-two-hybrid screen with CPRabA5e as bait revealed 13 interacting partner proteins, mainly located in thylakoids and plastoglobules. These proteins are known or predicted to be involved in development, stress responses, and photosynthesis related processes, consistent with the stress phenotypes observed. The results observed suggest a role of CPRabA5e in transport to and from thylakoids, similar to cytosolic Rab proteins involved in vesicle transport.

New putative chloroplast vesicle transport components and cargo proteins revealed using a bioinformatics approach: an Arabidopsis model.

Proteins and lipids are known to be transported to targeted cytosolic compartments in vesicles. A similar system in chloroplasts is suggested to transfer lipids from the inner envelope to the thylakoids. However, little is known about both possible cargo proteins and the proteins required to build a functional vesicle transport system in chloroplasts. A few components have been suggested, but only one (CPSAR1) has a verified location in chloroplast vesicles. This protein is localized in the donor membrane (envelope) and vesicles, but not in the target membrane (thylakoids) suggesting it plays a similar role to a cytosolic homologue, Sar1, in the secretory pathway. Thus, we hypothesized that there may be more similarities, in addition to lipid transport, between the vesicle transport systems in the cytosol and chloroplast, i.e. similar vesicle transport components, possible cargo proteins and receptors. Therefore, using a bioinformatics approach we searched for putative chloroplast components in the model plant Arabidopsis thaliana, corresponding mainly to components of the cytosolic vesicle transport system that may act in coordination with previously proposed COPII chloroplast homologues. We found several additional possible components, supporting the notion of a fully functional vesicle transport system in chloroplasts. Moreover, we found motifs in thylakoid-located proteins similar to those of COPII vesicle cargo proteins, supporting the hypothesis that chloroplast vesicles may transport thylakoid proteins from the envelope to the thylakoid membrane. Several putative cargo proteins are involved in photosynthesis, thus we propose the existence of a novel thylakoid protein pathway that is important for construction and maintenance of the photosynthetic machinery.