ARC5, a cytosolic dynamin-like protein from plants, is part of the chloroplast division machinery

Abstract

Chloroplast division in plant cells is orchestrated by a complex macromolecular machine with components positioned on both the inner and outer envelope surfaces. The only plastid division proteins identified to date are of endosymbiotic origin and are localized inside the organelle. Employing positional cloning methods in Arabidopsis in conjunction with a novel strategy for pinpointing the mutant locus, we have identified a gene encoding a new chloroplast division protein, ARC5. Mutants of ARC5 exhibit defects in chloroplast constriction, have enlarged, dumbbellshaped chloroplasts, and are rescued by a wild-type copy of ARC5. The ARC5 gene product shares similarity with the dynamin family of GTPases, which mediate endocytosis, mitochondrial division, and other organellar fission and fusion events in eukaryotes. Phylogenetic analysis showed that ARC5 is related to a group of dynamin-like proteins unique to plants. A GFP-ARC5 fusion protein localizes to a ring at the chloroplast division site. Chloroplast import and protease protection assays indicate that the ARC5 ring is positioned on the outer surface of the chloroplast. Thus, ARC5 is the first cytosolic component of the chloroplast division complex to be identified. ARC5 has no obvious counterparts in prokaryotes, suggesting that it evolved from a dynamin-related protein present in the eukaryotic ancestor of plants. These results indicate that the chloroplast division apparatus is of mixed evolutionary origin and that it shares structural and mechanistic similarities with both the cell division machinery of bacteria and the dynamin-mediated organellar fission machineries of eukaryotes.T he chloroplasts of plants and algae are widely believed to have evolved only once from a free-living cyanobacterial endosymbiont (1). Over evolutionary time, many of the genes once present in the endosymbiont have been transferred to the nuclear genome where they have acquired sequences encoding transit peptides that direct their gene products back to the chloroplast (1, 2). This scenario describes the origin of the five previously identified plastid division proteins in plants, all of which evolved from related cell division proteins in cyanobacteria, are encoded in the nucleus, and are localized inside the chloroplast. These include FtsZ1 and FtsZ2, tubulin-like proteins that localize to a ring at the site of plastid constriction (3-10), MinD and MinE, which regulate placement of the plastid division site (11-13), and ARTEMIS, which appears to mediate constriction of the envelope membranes (14).Despite localization of the previously identified plastid division proteins inside the chloroplasts in plant cells, ultrastructural studies have shown that plastid division entails the coordinated activity of components localized outside as well as inside the organelle. In plants, the chloroplast division complex comprises electron-dense structures situated both on the stromal surface of the inner envelope membrane and on the cytosolic surface of the outer membrane (15). These structur...

FtsZ and the division of prokaryotic cells and organelles

“…The protein machine of chloroplasts, unlike that of bacteria, can be observed in electron micrographs of thin sections. One possible reason for its visibility is that this machine consists of at least three layers: an innermost ring comprising FtsZ, followed by the plastid dividing ring that is visible by negative staining but is of unknown composition 102 , and an outermost ring on the outside of the organelle that contains a dynamin-like protein (DRP) 103,104 . Dynamins are eukaryotic GTPases that assemble on membranes and mediate membrane fission 97 ; dynamin also has a role in cytokinesis of animal and plant cells 105,106 .…”

GFP‐targeting allows visualization of the apicoplast throughout the life cycle of live malaria parasites

“…In mammalian cells, fission of the mitochondrion (reviewed in Westermann, 2008) does not involve Ftsz. Dynamin-like proteins are known to be involved in mitochondrial fission in some organisms, including yeast and Caenorhabditis elegans (Bleazard et al, 1999;Labrousse et al, 1999), and also in the fission of plastids in plants (Gao et al, 2003;Miyagishima et al, 2003). It is known that Plasmodium and other apicomplexa encode dynamin proteins, one of which (dynamin-2) is similar to dynamin-like proteins that are known to be involved in mitochondrial fission in other organisms (Vaishnava and Striepen, 2006).…”

“…Here, Ftsz proteins form a ring structure within the stroma of the chloroplast, seemingly constricting the organelle in a drawstring action (reviewed in Margolin, 2005). Other proteins, including dynamin, are then recruited to the constricted regions of the chloroplast and this leads to fission of the organelle (Gao et al, 2003;Miyagishima et al, 2003). In the case of the mitochondrion, proteins, including dynamin, are similarly recruited to sites of constriction and appear to lead to mitochondrial fission (Bleazard et al, 1999;Labrousse et al, 1999).…”

A Conserved Transcription Factor Mediates Nuclear Control of Organelle Biogenesis in Anciently Diverged Land Plants

“…Although many nuclear genes that contribute functional components to the organelle have been described, including those of endosymbiotic origin that control division from within the organelle (Osteryoung et al, 1998), very few regulatory genes have been identified even in model organisms. One example, Accumulation and Replication of Chloroplasts5, encodes a cytoplasmically localized dynamin-like protein that regulates chloroplast division in Arabidopsis thaliana (Gao et al, 2003). A second example, the Golden2-like (GLK) genes, encode transcription factors that regulate chloroplast development in diverse species, namely in the monocot maize (Zea mays) and in the eudicot Arabidopsis (Langdale and Kidner, 1994;Hall et al, 1998;Rossini et al, 2001;Fitter et al, 2002).…”