Diverse eukaryotes have retained mitochondrial homologues of the bacterial division protein FtsZ

Mitochondrial fission requires the division of both the inner and outer mitochondrial membranes. Dynamin-related proteins operate in division of the outer membrane of probably all mitochondria, and also that of chloroplasts--organelles that have a bacterial origin like mitochondria. How the inner mitochondrial membrane divides is less well established. Homologues of the major bacterial division protein, FtsZ, are known to reside inside mitochondria of the chromophyte alga Mallomonas, a red alga, and the slime mould Dictyostelium discoideum, where these proteins are likely to act in division of the organelle. Mitochondrial FtsZ is, however, absent from the genomes of higher eukaryotes (animals, fungi, and plants), even though FtsZs are known to be essential for the division of probably all chloroplasts. To begin to understand why higher eukaryotes have lost mitochondrial FtsZ, we have sampled various diverse protists to determine which groups have retained the gene. Database searches and degenerate PCR uncovered genes for likely mitochondrial FtsZs from the glaucocystophyte Cyanophora paradoxa, the oomycete Phytophthora infestans, two haptophyte algae, and two diatoms--one being Thalassiosira pseudonana, the draft genome of which is now available. From Thalassiosira we also identified two chloroplast FtsZs, one of which appears to be undergoing a C-terminal shortening that may be common to many organellar FtsZs. Our data indicate that many protists still employ the FtsZ-based ancestral mitochondrial division mechanism, and that mitochondrial FtsZ has been lost numerous times in the evolution of eukaryotes.     

Two Types of FtsZ Proteins in Mitochondria and Red-Lineage Chloroplasts: The Duplication of FtsZ Is Implicated in Endosymbiosis

The ancestors of plastids and mitochondria were once free-living bacteria that became organelles as a result of endosymbiosis. According to this theory, a key bacterial division protein, FtsZ, plays a role in plastid division in algae and plants as well as in mitochondrial division in lower eukaryotes. Recent studies have shown that organelle division is a process that combines features derived from the bacterial division system with features contributed by host eukaryotic cells. Two nonredundant versions of FtsZ, FtsZ1 and FtsZ2, have been identified in green-lineage plastids, whereas most bacteria have a single ftsZ gene. To examine whether there is also more than one type of FtsZ in red-lineage chloroplasts (red algal chloroplasts and chloroplasts that originated from the secondary endosymbiosis of red algae) and in mitochondria, we obtained FtsZ sequences from the complete sequence of the primitive red alga Cyanidioschyzon merolae and the draft sequence of the stramenopile (heterokont) Thalassiosira pseudonana. Phylogenetic analyses that included known FtsZ proteins identified two types of chloroplast FtsZ in red algae (FtsZA and FtsZB) and stramenopiles (FtsZA and FtsZC). These analyses also showed that FtsZB emerged after the red and green lineages diverged, while FtsZC arose by the duplication of an ftsZA gene that in turn descended from a red alga engulfed by the ancestor of stramenopiles. A comparison of the predicted proteins showed that like bacterial FtsZ and green-lineage FtsZ2, FtsZA has a short conserved C-termmal sequence (the C-terminal core domain), whereas FtsZB and FtsZC, like the green-lineage FtsZ1, lack this sequence. In addition, the Cyanidioschyzon and Dictyostelium genomes encode two types of mitochondrial FtsZ proteins, one of which lacks the C-terminal variable domain. These results suggest that the acquisition of an additional FtsZ protein with a modified C terminus was common to the primary and secondary endosymbioses that produced plastids and that this also occurred during the establishment of mitochondria, presumably to regulate the multiplication of these organelles.     

Two Dictyostelium orthologs of the prokaryotic cell division protein FtsZ localize to mitochondria and are required for the maintenance of normal mitochondrial morphology

In bacteria, the protein FtsZ is the principal component of a ring that constricts the cell at division. Though all mitochondria probably arose through a single, ancient bacterial endosymbiosis, the mitochondria of only certain protists appear to have retained FtsZ, and the protein is absent from the mitochondria of fungi, animals, and higher plants. We have investigated the role that FtsZ plays in mitochondrial division in the genetically tractable protist Dictyostelium discoideum, which has two nuclearly encoded FtsZs, FszA and FszB, that are targeted to the inside of mitochondria. In most wild-type amoebae, the mitochondria are spherical or rod-shaped, but in fsz-null mutants they become elongated into tubules, indicating that a decrease in mitochondrial division has occurred. In support of this role in organelle division, antibodies to FszA and FszA-green fluorescent protein (GFP) show belts and puncta at multiple places along the mitochondria, which may define future or recent sites of division. FszB-GFP, in contrast, locates to an electron-dense, submitochondrial body usually located at one end of the organelle, but how it functions during division is unclear. This is the first demonstration of two differentially localized FtsZs within the one organelle, and it points to a divergence in the roles of these two proteins.     

Triple immunofluorescent labeling of FtsZ, dynamin, and EF-Tu reveals a loose association between the inner and outer membrane mitochondrial division machinery in the red alga Cyanidioschyzon merolae.

In the mitochondria of primitive eukaryotes, FtsZ and dynamin are part of the machinery involved in division of the inner and outer membranes, respectively. These genes also commonly function in the same manner during chloroplast division. In this study, a relationship between the localization of the inner and outer division machinery was directly shown for the first time. Triple immunofluorescent labeling was performed in the red alga Cyanidioschyzon merolae by a device using narrow bandpass filter sets and bright photostable dyes. FtsZ (CmFtsZ1) and dynamin (CmDnm1) localizations were examined simultaneously throughout the mitochondrial division cycle with an alternative mitochondrial marker protein, the mitochondrial translation elongation factor EF-Tu, whose localization was also shown to be identical to the mitochondrial matrix. FtsZ and dynamin did not necessarily co-localize when both were recruited to the mitochondrial constriction site, indicating that inner and outer dividing machineries are not in tight association during the late stage of division.     

Structure, function and evolution of the mitochondrial division apparatus

Mitochondria are derived from free-living alpha-proteobacteria that were engulfed by eukaryotic host cells through the process of endosymbiosis, and therefore have their own DNA which is organized using basic proteins to form organelle nuclei (nucleoids). Mitochondria divide and are split amongst the daughter cells during cell proliferation. Their division can be separated into two main events: division of the mitochondrial nuclei and division of the matrix (the so-called mitochondrial division, or mitochondriokinesis). In this review, we first focus on the cytogenetical relationships between mitochondrial nuclear division and mitochondriokinesis. Mitochondriokinesis occurs after mitochondrial nuclear division, similar to bacterial cytokinesis. We then describe the fine structure and dynamics of the mitochondrial division ring (MD ring) as a basic morphological background for mitochondriokinesis. Electron microscopy studies first identified a small electron-dense MD ring in the cytoplasm at the constriction sites of dividing mitochondria in the slime mold Physarum polycephalum, and then two large MD rings (with outer cytoplasmic and inner matrix sides) in the red alga Cyanidioschyzon merolae. Now MD rings have been found in all eukaryotes. In the third section, we describe the relationships between the MD ring and the FtsZ ring descended from ancestral bacteria. Other than the GTPase, FtsZ, mitochondria have lost most of the proteins required for bacterial cytokinesis as a consequence of endosymbiosis. The FtsZ protein forms an electron transparent ring (FtsZ or Z ring) in the matrix inside the inner MD ring. For the fourth section, we describe the dynamic association between the outer MD ring with a ring composed of the eukaryote-specific GTPase dynamin. Recent studies have revealed that eukaryote-specific GTPase dynamins form an electron transparent ring between the outer membrane and the MD ring. Thus, mitochondriokinesis is thought to be controlled by a mitochondrial division (MD) apparatus including a dynamic trio, namely the FtsZ, MD and dynamin rings, which consist of a chimera of rings from bacteria and eukaryotes in primitive organisms. Since the genes for the MD ring and dynamin rings are not found in the prokaryotic genome, the host genomes may make these rings to actively control mitochondrial division. In the fifth part, we focus on the dynamic changes in the formation and disassembly of the FtsZ, MD and dynamin rings. FtsZ rings are digested during a later period of mitochondrial division and then finally the MD and dynamin ring apparatuses pinched off the daughter mitochondria, supporting the idea that the host genomes are responsible for the ultimate control of mitochondrial division. We discuss the evolution, from the original vesicle division (VD) apparatuses to VD apparatuses including classical dynamin rings and MD apparatuses. It is likely that the MD apparatuses involving the dynamic trio evolved into the plastid division (PD) apparatus in Bikonta, while in Opisthokonta, the MD apparatus was simplified during evolution and may have branched into the mitochondrial fusion apparatus. Finally, we describe the possibility of intact isolation of large MD/PD apparatuses, the identification of all their proteins and their related genes using C. merolae genome information and TOF-MS analyses. These results will assist in elucidating the universal mechanism and evolution of MD, PD and VD apparatuses.     

A plant-specific dynamin-related protein forms a ring at the chloroplast division site

Chloroplasts have retained the bacterial FtsZ for division, whereas mitochondria lack FtsZ except in some lower eukaryotes. Instead, mitochondrial division involves a dynamin-related protein, suggesting that chloroplasts retained the bacterial division system, whereas a dynamin-based system replaced the bacterial system in mitochondria during evolution. In this study, we identified a novel plant-specific group of dynamins from the primitive red alga Cyanidioschyzon merolae. Synchronization of chloroplast division and immunoblot analyses showed that the protein (CmDnm2) associates with the chloroplast only during division. Immunocytochemical analyses showed that CmDnm2 appears in cytoplasmic patches just before chloroplast division and is recruited to the cytosolic side of the chloroplast division site to form a ring in the late stage of division. The ring constricts until division is complete, after which it disappears. These results show that a dynamin-related protein also participates in chloroplast division and that its behavior differs from that of FtsZ and plastid-dividing rings that form before constriction at the site of division. Combined with the results of a recent study of mitochondrial division in Cyanidioschyzon, our findings led us to hypothesize that when first established in lower eukaryotes, mitochondria and chloroplasts divided using a very similar system that included the FtsZ ring, the plastid-dividing/mitochondrion-dividing ring, and the dynamin ring.     

An evolutionary puzzle: chloroplast and mitochondrial division rings

Consistent with their bacterial origin, chloroplasts and primitive mitochondria retain a FtsZ ring for division. However, chloroplasts and mitochondria have lost most of the proteins required for bacterial division other than FtsZ and certain homologues of the Min proteins, but they do contain plastid and mitochondrion dividing rings, which were recently shown to be distinct from the FtsZ ring. Moreover, recent studies have revealed that rings of the eukaryote-specific dynamin-related family of GTPases regulate the division of chloroplasts and mitochondria, and these proteins emerged early in eukaryotic evolution. These findings suggest that the division of chloroplasts and primitive mitochondria involve very similar systems, consisting of an amalgamation of rings from bacteria and eukaryotes.     

The cellular machineries responsible for the division of endosymbiotic organelles

Chloroplasts (plastids) and mitochondria evolved from endosymbiotic bacteria. These organelles perform vital functions in photosynthetic eukaryotes, such as harvesting and converting energy for use in biological processes. Consistent with their evolutionary origins, plastids and mitochondria proliferate by the binary fission of pre-existing organelles. Here, I review the structures and functions of the supramolecular machineries driving plastid and mitochondrial division, which were discovered and first studied in the primitive red alga Cyanidioschyzon merolae. In the past decade, intact division machineries have been isolated from plastids and mitochondria and examined to investigate their underlying structure and molecular mechanisms. A series of studies has elucidated how these division machineries assemble and transform during the fission of these organelles, and which of the component proteins generate the motive force for their contraction. Plastid- and mitochondrial-division machineries have important similarities in their structures and mechanisms despite sharing no component proteins, implying that these division machineries evolved in parallel. The establishment of these division machineries might have enabled the host eukaryotic ancestor to permanently retain these endosymbiotic organelles by regulating their binary fission and the equal distribution of resources to daughter cells. These findings provide key insights into the establishment of endosymbiotic organelles and have opened new avenues of research into their evolution and mechanisms of proliferation.     

Protein transport into mitochondria

Mitochondria are made up of two membrane systems that subdivide this organelle into two aqueous subcompartments: the matrix, which is enclosed by the inner membrane, and the intermembrane space, which is located between the inner and the outer membrane. Protein import into mitochondria is a complex reaction, as every protein has to be routed to its specific destination within the organelle. In the past few years, studies with mitochondria of Neurospora crassa and Saccharomyces cerevisiae have led to the identification of four distinct translocation machineries that are conserved among eukaryotes. These translocases, in a concerted fashion, mediate import and sorting of proteins into the mitochondrial subcompartments.     

PARC6, a novel chloroplast division factor, influences FtsZ assembly and is required for recruitment of PDV1 during chloroplast division in Arabidopsis

Chloroplast division in plant cells is accomplished through the coordinated action of the tubulin-like FtsZ ring inside the organelle and the dynamin-like ARC5 ring outside the organelle. This coordination is facilitated by ARC6, an inner envelope protein required for both assembly of FtsZ and recruitment of ARC5. Recently, we showed that ARC6 specifies the mid-plastid positioning of the outer envelope proteins PDV1 and PDV2, which have parallel functions in dynamin recruitment. PDV2 positioning involves direct ARC6–PDV2 interaction, but PDV1 and ARC6 do not interact indicating that an additional factor functions downstream of ARC6 to position PDV1. Here, we show that PARC6 (paralog of ARC6), an ARC6-like protein unique to vascular plants, fulfills this role. Like ARC6, PARC6 is an inner envelope protein with its N-terminus exposed to the stroma and Arabidopsis parc6 mutants exhibit defects of chloroplast and FtsZ filament morphology. However, whereas ARC6 promotes FtsZ assembly, PARC6 appears to inhibit FtsZ assembly, suggesting that ARC6 and PARC6 function as antagonistic regulators of FtsZ dynamics. The FtsZ inhibitory activity of PARC6 may involve its interaction with the FtsZ-positioning factor ARC3. A PARC6–GFP fusion protein localizes both to the mid-plastid and to a single spot at one pole, reminiscent of the localization of ARC3, PDV1 and ARC5. Although PARC6 localizes PDV1, it is not required for PDV2 localization or ARC5 recruitment. Our findings indicate that PARC6, like ARC6, plays a role in coordinating the internal and external components of the chloroplast division complex, but that PARC6 has evolved distinct functions in the division process.     

Mitochondrial division prevents neurodegeneration

Mitochondrial division is mediated by the conserved dynamin-related GTPase DNM1L/DRP1. DNM1L assembles onto the surface of mitochondria and constricts this tubular organelle. Alterations in mitochondrial division are linked to many neurodegenerative diseases. However, the in vivo function of mitochondrial division is poorly understood. In our recent paper, we studied the physiological role of mitochondrial division in postmitotic neurons using the cre-loxP system. We found that the loss of DNM1L resulted in increased oxidative damage in mitochondria, impaired respiration and neurodegeneration in postmitotic neurons. Suggesting a decrease in mitochondrial turnover, mitophagy-related proteins such as LC3, SQSTM1/p62 and ubiqutin accumulated in division-defective mitochondria. These findings suggest that mitochondrial division functions as an important quality control mechanism that suppresses oxidative damage and neurodegeneration in vivo

     

Origin and evolution of the chloroplast division machinery

Chloroplasts were originally established in eukaryotes by the endosymbiosis of a cyanobacterium; they then spread through diversification of the eukaryotic hosts and subsequent engulfment of eukaryotic algae by previously nonphotosynthetic eukaryotes. The continuity of chloroplasts is maintained by division of preexisting chloroplasts. Like their ancestors, chloroplasts use a bacterial division system based on the FtsZ ring and some associated factors, all of which are now encoded in the host nuclear genome. The majority of bacterial division factors are absent from chloroplasts and several new factors have been added by the eukaryotic host. For example, the ftsZ gene has been duplicated and modified, plastid-dividing (PD) rings were most likely added by the eukaryotic host, and a member of the dynamin family of proteins evolved to regulate chloroplast division. The identification of several additional proteins involved in the division process, along with data from diverse lineages of organisms, our current knowledge of mitochondrial division, and the mining of genomic sequence data have enabled us to begin to understand the universality and evolution of the division system. The principal features of the chloroplast division system thus far identified are conserved across several lineages, including those with secondary chloroplasts, and may reflect primeval features of mitochondrial division. Shin-ya Miyagishima is the recipient of the Botanical Society Award for Young Scientists, 2004.     

Plant mitochondrial dynamics

Higher plant mitochondria are dynamic, pleomorphic organelles. The higher plant chondriome (all mitochondria in a cell collectively) is typically composed of numerous, physically discrete, mitochondria. However, frequent inter-mitochondrial fusion, enabling the mixing and recombination of mtDNA, ensures that the higher plant chondriome functions, at least genetically, as a discontinuous whole. Nothing is known about the genes controlling mitochondrial fusion in plants; there are no plant homologues of most of the genes known to be involved in fusion in other organisms. In contrast, the mitochondrial fission apparatus is generally conserved. Higher plant mitochondria use dynamin-like and Fis-type proteins for division; like yeast and animals, higher plants have lost the mitochondrial-specific form of the prokaryote-derived protein, FtsZ. In addition to being providers of energy for life, mitochondria provide a trigger for death. The role of mitochondrial dynamics in the initiation and promulgation of cell death is conserved in higher plants although there are specific differences in the genes and mechanisms involved relative to other higher eukaryotes.     

Plant mitochondrial dynamics

Higher plant mitochondria are dynamic, pleomorphic organelles. The higher plant chondriome (all mitochondria in a cell collectively) is typically composed of numerous, physically discrete, mitochondria. However, frequent inter-mitochondrial fusion, enabling the mixing and recombination of mtDNA, ensures that the higher plant chondriome functions, at least genetically, as a discontinuous whole. Nothing is known about the genes controlling mitochondrial fusion in plants; there are no plant homologues of most of the genes known to be involved in fusion in other organisms. In contrast, the mitochondrial fission apparatus is generally conserved. Higher plant mitochondria use dynamin-like and Fis-type proteins for division; like yeast and animals, higher plants have lost the mitochondrial-specific form of the prokaryote-derived protein, FtsZ. In addition to being providers of energy for life, mitochondria provide a trigger for death. The role of mitochondrial dynamics in the initiation and promulgation of cell death is conserved in higher plants although there are specific differences in the genes and mechanisms involved relative to other higher eukaryotes.     

Filamentous temperature-sensitive Z (FtsZ) isoforms specifically interact in the chloroplasts and in the cytosol of Physcomitrella patens

Plant filamentous temperature-sensitive Z (FtsZ) proteins have been reported to be involved in biological processes related to plastids. However, the precise functions of distinct isoforms are still elusive. Here, the intracellular localization of the FtsZ1-1 isoform in a moss, Physcomitrella patens, was examined. Furthermore, the in vivo interaction behaviour of four distinct FtsZ isoforms was investigated. Localization studies of green fluorescent protein (GFP)-tagged FtsZ1-1 and fluorescence resonance energy transfer (FRET) analyses employing all dual combinations of four FtsZ isoforms were performed in transient protoplast transformation assays. FtsZ1-1 is localized to network structures inside the chloroplasts and exerts influence on plastid division. Interactions between FtsZ isoforms occur in distinct ordered structures in the chloroplasts as well as in the cytosol. The results expand the view of the involvement of Physcomitrella FtsZ proteins in chloroplast and cell division. It is concluded that duplication and diversification of ftsZ genes during plant evolution were the main prerequisites for the successful remodelling and integration of the prokaryotic FtsZ-dependent division mechanism into the cellular machineries of distinct complex processes in plants.     

Organelle division: Self-assembling GTPase caught in the middle

Recent findings indicate that the dynamin GTPase helps to divide animal and fungal mitochondria, and that the tubulin-like FtsZ GTPase is involved in division of, not only most bacteria, but also chloroplasts and probably mitochondria of unicellular eukaryotes.     

Isolation of mitochondrial and plastid ftsZ genes and analysis of the organelle targeting sequence in the diatom Chaetoceros neogracile (Diatoms,Bacillariophyceae)

Mitochondria and plastids multiply by division in eukaryotic cells. Recently, the eukaryotic homolog of the bacterial cell division protein FtsZ was identified and shown to play an important role in the organelle division process inside the inner membrane. To explore the evolution of FtsZ proteins, and to accumulate data on the protein import system in mitochondria and plastids of the red algal lineage, one mitochondrial and three plastid ftsZ genes were isolated from the diatom Chaetoceros neogracile, whose plastids were acquired by secondary endosymbiotic uptake of a red alga. Protein import into organelles depends on the N‐terminal organelle targeting sequences. N‐terminal bipartite presequences consisting of an endoplasmic reticulum signal peptide and a plastid transit peptide are required for protein import into diatom plastids. To characterize the organelle targeting peptides of C. neogracile, we observed the localization of each green fluorescent protein‐tagged predicted organelle targeting peptide in cultured tobacco cells and diatom cells. Our data suggested that each targeting sequences functioned both in tobacco cultured cells and diatom cells.     

Mitochondrial protein import: from transport pathways to an integrated network

Mitochondria, the powerhouses of the cell, import most of their proteins from the cytosol. It was originally assumed that mitochondria imported precursor proteins via a general pathway but recent studies have revealed a remarkable variety of import pathways and mechanisms. Currently, five different protein import pathways can be distinguished. However, the import machineries cooperate with each other and are connected to other systems that function in the respiratory chain, mitochondrial membrane organization, protein quality control and endoplasmic reticulum-mitochondria junctions. In this Opinion, we propose that mitochondrial protein import should not be seen as an independent task of the organelle and that a network of cooperating machineries is responsible for major mitochondrial functions.     

Mechanisms of iron-sulfur protein maturation in mitochondria, cytosol and nucleus of eukaryotes

Iron-sulfur (Fe/S) clusters are important cofactors of numerous proteins involved in electron transfer, metabolic and regulatory processes. In eukaryotic cells, known Fe/S proteins are located within mitochondria, the nucleus and the cytosol. Over the past years the molecular basis of Fe/S cluster synthesis and incorporation into apoproteins in a living cell has started to become elucidated. Biogenesis of these simple inorganic cofactors is surprisingly complex and, in eukaryotes such as Saccharomyces cerevisiae, is accomplished by three distinct proteinaceous machineries. The "iron-sulfur cluster (ISC) assembly machinery" of mitochondria was inherited from the bacterial ancestor of mitochondria. ISC components are conserved in eukaryotes from yeast to man. The key principle of biosynthesis is the assembly of the Fe/S cluster on a scaffold protein before it is transferred to target apoproteins. Cytosolic and nuclear Fe/S protein maturation also requires the function of the mitochondrial ISC assembly system. It is believed that mitochondria contribute a still unknown compound to biogenesis outside the organelle. This compound is exported by the mitochondrial "ISC export machinery" and utilised by the "cytosolic iron-sulfur protein assembly (CIA) machinery". Components of these two latter systems are also highly conserved in eukaryotes. Defects in the mitochondrial ISC assembly and export systems, but not in the CIA machinery have a strong impact on cellular iron uptake and intracellular iron distribution showing that mitochondria are crucial for both cellular Fe/S protein assembly and iron homeostasis.     

植物叶绿体分裂的分子机制

叶绿体是植物细胞内一种重要的细胞器．它不仅是光合作用的场所,还是其它多种中间代谢的场所．叶绿体起源于蓝细菌,与其原核祖先类似,通过二分裂方式进行增殖．最近的研究表明,叶绿体的分裂装置包含原核起源和真核起源的蛋白质,它们在叶绿体的内膜内侧和外膜外侧协同作用以完成叶绿体的分裂．在过去十几年里,包括丝状温度敏感蛋白Z（FtsZ）、Min系统蛋白、质体分裂蛋白（PDV）和ARC蛋白等在内的多个叶绿体分裂相关组分被分离鉴定．本文简要介绍了叶绿体分裂装置各成员的发现、叶绿体被膜的收缩和叶绿体分裂位点的选择机制．另外,植物发育过程中叶绿体分裂可能受到细胞的控制,但目前对细胞如何调控叶绿体分裂知之甚少．本文对该领域的最新研究进展也进行了综述．