MIRO1 influences the morphology and intracellular distribution of mitochondria during embryonic cell division in Arabidopsis

Regulating the morphology and intracellular distribution of mitochondria is essential for embryo development in animals. However, the importance of such regulation is not clearly defined in plants. The evolutionarily conserved Miro proteins are known to be involved in the regulation of mitochondrial morphology and motility. We previously demonstrated that MIRO1, an Arabidopsis thaliana orthologue of the Miro protein, is required for embryogenesis. An insertional mutation in the MIRO1 gene causes arrest of embryonic cell division, leading to abortion of the embryo at an early stage. Here we investigated the role of MIRO1 in the regulation of mitochondrial behaviour in egg cells and early-stage embryos using GFP-labeled mitochondria. Two-photon laser scanning microscopy revealed that, in miro1 mutant egg cells, mitochondria are abnormally enlarged, although egg cell formation is nearly unaffected. After fertilization and subsequent zygotic cell division, the homozygous miro1 mutant two-celled embryo contained a significantly reduced number of mitochondria in its apical cell compared with the wild type, suggesting that the miro1 mutation inhibits proper intracellular distribution of mitochondria, leading to an arrest of embryonic cell division. Our findings suggest that proper mitochondrial morphology and intracellular distribution are maintained by MIRO1 and are vital for embryonic cell division.     

Promotion of respiration by auxin in the induction of cell division in suspension culture

Auxins (IAA, NAA, p-CPA) caused 20–30% promotion of respiration of auxin-dependent cell cultures ofNicotiana tabacum, Glycine max, Taraxacum mongolica, andAtriplex sp. and had small if any effect on respiration of auxin-independent cell cultures ofRubus sp. andScorzonera hispanica. Antiauxin (p-CPIBA) did not affect the respiration. The auxin effect on the respiration of tobacco cells was revealed 10 min after its addition to the suspension and reached a maximum value in 60 min. This stimulation preceded the induction of cell division by auxin. Mitochondria isolated from auxin-treated tobacco cells had greater oxidative and phosphorylative activity than mitochondria from untreated cells. However, isolated mitochondria did not respond to auxin. The inhibitors of respiration (cyanide, monoiodoacetate, malonate, and 2,4-dinitrophenol) eliminated auxin effect on the respiration and cell division. It is concluded that the promotion of respiration is a common event for the auxin effects both on cell extension and on cell division. This promotion is necessary for the induction of cell division and is exerted via direct activation of mitochondriain situ.     

Diatom motility: An explanation and a problem

Except for the lack of a centriole, interphase cell morphology and cell division in Stichococcus is similar to that in Klebsormidium. The cell in Stichococcus is largely filled by a chloroplast and pyrenoid, at the side of which are two mitochondria and one small peroxisome. The chloroplast/pyrenoid cleaves early in prophase, probably completely, and the nucleus is inserted between the two halves. A band of 3–5 microtubules always encircles the prophase nucleus; these disappear by metaphase. The spindle is open, the daughter nuclei remain far apart at telophase and during cytokinesis, and vacuoles collect between them; no phycoplast is associated with the cleavage furrow.

These results indicate a close phyletic relationship between Stichococcus and Klebsormidium, two organisms which are now considered to be more closely related to the progenitors of the higher land plants than most of the other members of the Ulotrichales.     

The primitive red algae Cyanidium caldarium and Cyanidioschyzon merolae as model system for investigating the dividing apparatus of mitochondria and plastids

The Cyanidiophyceae species Cyanidium caldarium and Cyanidioschyzon merolae have played important roles in showing the division mechanisms of mitochondria and plastids. The apparatus regulating mitochondrial and plastid divisions was formerly unknown. We first identified the division apparatus of plastids, called the plastid-dividing ring (PD ring), in C. caldarium and the division apparatus of mitochondria, called the mitochondrion-dividing ring (MD ring), in C. merolae. Eukaryotic cell division is therefore controlled by at least three dividing apparati (rings)—a contractile ring, an MD ring, and a PD ring—while bacterial division is controlled by a single bacterial contractile FtsZ ring. BioEssays 20 :344-354, 1998.© 1998 John Wiley & Sons, Inc.     

The cell wall amidase AmiB is essential for Pseudomonas aeruginosa cell division,drug resistance and viability

The physiological function of cell wall amidases has been investigated in several proteobacterial species. In all cases, they have been implicated in the cleavage of cell wall material synthesized by the cytokinetic ring. Although typically non‐essential, this activity is critical for daughter cell separation and outer membrane invagination during division. In Escherichia coli, proteins with LytM domains also participate in cell separation by stimulating amidase activity. Here, we investigated the function of amidases and LytM proteins in the opportunistic pathogen Pseudomonas aeruginosa. In agreement with studies in other organisms, ^PaAmiB and three LytM proteins were found to play crucial roles in P. aeruginosa cell separation, envelope integrity and antibiotic resistance. Importantly, the phenotype of amidase‐defective P. aeruginosa cells also differed in informative ways from the E. coli paradigm; ^PaAmiB was found to be essential for viability and the successful completion of cell constriction. Our results thus reveal a key role for amidase activity in cytokinetic ring contraction. Furthermore, we show that the essential function of ^PaAmiB can be bypassed in mutants activated for a Cpx‐like envelope stress response, suggesting that this signaling system may elicit the repair of division machinery defects in addition to general envelope damage.     

The machinery of mitochondrial fusion, division, and distribution, and emerging connections to apoptosis

Mitochondrial undergo regulated fusion and division in many organisms and cell types, and each event is mediated by a different complex of proteins each containing at least one large GTPase. The mitochondrial fusion and division molecular machinery is in large part conserved; recent studies show a functional connection between some of these proteins and the apoptotic cascade. Mitochondria also undergo directed movement in cells, and the gene products that attach and propel mitochondria along cytoskeletal elements (actin filaments in some organisms, microtubules in others) are becoming gradually elucidated.     

Suspended animation,diapause and quiescence

Developing organisms require nutrients to support cell division vital for growth and development. An adaptation to stress, used by many organisms, is to reversibly enter an arrested state by reducing energy-requiring processes, such as development and cell division. This “wait it out” approach to survive stress until the environment is conductive for growth and development is used by many metazoans. Much is known about the molecular regulation of cell division, metazoan development and responses to environmental stress. However, how these biological processes intersect is less understood. Here, we review studies conducted in Caenorhabditis elegans that investigate how stresses such as oxygen deprivation (hypoxia and anoxia), exogenous chemicals or starvation affect cellular processes in the embryo, larvae or adult germline. Using C. elegans to identify how stress signals biological arrest can help in our understanding of evolutionary pressures as well as human health-related issues.     

Programmed cell death in parasitic protozoans that lack mitochondria

In multicellular organisms and in all protozoans harbouring mitochondria, the pathways leading to programmed cell death (PCD) are localized in the mitochondria. Intriguingly, unicellular parasites devoid of mitochondria such as Trichomonas vaginalis and Giardia intestinalis undergo a form of cell death resembling apoptosis, the most frequent form of PCD. This reinforces the idea that PCD must have evolved before the evolution of multicellularity. Moreover, this leads to the hypothesis of an early emergence of death pathways in eukaryotes preceding mitochondrial endosymbiosis and brings into question the central role of mitochondria in PCD.     

Behavior of mitochondria and their nuclei during amoebae cell proliferation inPhysarum polycephalum

Summary We investigated the manner of mitochondrial DNA (mtDNA) replication and distribution during the culture ofPhysarum polycephalum amoebae cells by microphotometry, anti-BrdU immunofluorescence microscopy, and quantitative hybridization analysis. In amoebae cells ofP. polycephalum, the number of mitochondria per cell and the shape of both mitochondria and mitochondrial nuclei (mt-nuclei) noticeably changed over the culture period. At the time of transfer, about 27 short ellipsoidal shaped mitochondria, which each contained a small amount of DNA, were observed in each cell. The number of mitochondria per cell decreased gradually, while the amount of mtDNA in an mt-nucleus and the length of mt-nuclei increased gradually. Midway through the middle logarithmic growth phase, the number of mitochondria per cell reached a minimum (about 10 mitochondria per cell), but most mtnuclei assumed an elongated shape and contained a large amount of mtDNA. During the late log- and stationary-growth phase, the number of mitochondria per cell increased gradually, while the amount of DNA in an mt-nucleus and mt-nuclei length decreased gradually. Upon completion of the stationary phase, the number and condition of mitochondria within cells returned to that first observed at the time of transfer. The total amount of mtDNA in a cell increased about 1.6-fold the first day, decreased immediately, then maintained a constant level ranging from 130 to 160 T. Except for the fact that mtDNA synthesis began earlier than synthesis of cell nuclei, the rate of increase in mtDNA paralleled that of cell-nuclear DNA throughout the culture. These results indicate that mtDNA is continuously replicated in pace with cell proliferation and the rate of mitochondrial division varies during culture; this mitochondrial division does not synchronize with either mtDNA replication or cell division. Furthermore, we observed the spatial distribution of DNA replication sites along mt-nuclei. Replication began at several sites scattered along an mt-nucleus, and the number of replication sites increased as the length of mt-nuclei increased. These results indicate that mtDNA replication progresses in adjacent replicons, which are collectively termed a mitochondrial replicon cluster.Abbreviations DAPI 4,6-diamidino-2-phenylindole - VIMPCS video-intensified microscope photon counting system - BrdU 5-bromodeoxyuridine - FITC fluorescein isothiocyanate     

Bacterial cell division: regulating Z-ring formation

The earliest stage of cell division in bacteria is the formation of a Z ring, composed of a polymer of the FtsZ protein, at the division site. Z rings appear to be synthesized in a bi‐directional manner from a nucleation site (NS) located on the inside of the cytoplasmic membrane. It is the utilization of a NS specifically at the site of septum formation that determines where and when division will occur. However, a Z ring can be made to form at positions other than at the division site. How does a cell regulate utilization of a NS at the correct location and at the right time? In rod‐shaped bacteria such as Escherichia coli and Bacillus subtilis, two factors involved in this regulation are the Min system and nucleoid occlusion. It is suggested that in B. subtilis, the main role of the Min proteins is to inhibit division at the nucleoid‐free cell poles. In E. coli it is currently not clear whether the Min system can direct a Z ring to the division site at mid‐cell or whether its main role is to ensure that division inhibition occurs away from mid‐cell, a role analogous to that in B. subtilis. While the nucleoid negatively influences Z‐ring formation in its vicinity in these rod‐shaped organisms, the exact relationship between nucleoid occlusion and the ability to form a mid‐cell Z ring is unresolved. Recent evidence suggests that in B. subtilis and Caulobacter crescentus, utilization of the NS at the division site is intimately linked to the progress of a round of chromosome replication and this may form the basis of achieving co‐ordination between chromosome replication and cell division.     

Application of 13C-[2] - and 13C-[1,2] acetate in metabolic labelling studies of yeast and insect cells

The advantage of using ¹³C-labelled glucose in metabolic studies is that it is an important carbon and energy source for almost all biotechnologically and medically important organisms. On the other hand, the disadvantage is its relatively high cost in the labelling experiments. Looking for cheaper alternatives we found that ¹³C-[2] acetate or ¹³C-[1,2] acetate is a prospective compound for such experiments. Acetate is well incorporated by many organisms, including mammalian and insect cell cultures as preferred source of acetyl-CoA. Our experimental results using ¹³C NMR demonstrated that acetate was efficiently incorporated into glutamate and alanine secreted by the insect cell culture. Using D-stat culture of Saccharomyces uvarum on glucose/¹³C-acetate mineral media we demonstrated that the labelling patterns of proteinogenic amino acids can be well predicted on the basis of specific substrate consumption rates using the modified scheme of yeast metabolism and stoichiometric modelling. According to this scheme aspartate and alanine in S. uvarum under the experimental conditions used is synthesised in the mitochondria. Synthesis of alanine in the mitochondria was also demonstrated for Spodoptera frugiperda. For both organisms malic enzyme was also operative. For S. uvarum it was shown that the activity of malic enzyme is sufficient for supporting the mitochondrial biosynthetic reactions with NADPH.     

Untersuchungen über Beziehungen zwischen Zellteilung und Morphogenese bei Gewebekulturen vou Daucus carota III. Der Eiufluss des Kinetins auf die Ultrastruktur der Zellen

Relationship between cell division and morphogenesis of tissue culture of Daucus carota. III. The influence of kinetin on the ultrastructure of cells.— A previous paper reported a close correlation between cell division as influenced by kinetin and respiration, i.e. a high kinetin induced cell division rate corresponds with low oxygen consumption of the explants. This earlier observation was now confirmed. No significant differences between the ultrastructure of mitochondria of explants cultured with kinetin and those of explants cultured without kinetin could be observed, although the number of mitochondria per cell seems to be increased in explants cultured without kinetin. However, also the number of ribosomes in cells of explants cultured without kinetin seems to be increased as compared with those treated with kinetin. Kinetin apparently has no significant influence on the ultrastructure of cell organells under the conditions employed.     

The mitochondria ofPolytoma papillatum at two different stages of the vegetative cell cycle

Summary The examination of adjacent sections ofPolytoma papillatum cells by the electron microscope enabled us to construct three dimensional models of the chondriomes. These reconstructions show that in two predivision cells the chondriome primarily consists of one large and highly convoluted mitochondrion. One and three small ovoid mitochondria also exist. Examination of two new daughter cells reveales 30 and 47 mitochondria of various shapes and sizes. This indicates that during the growth phase ofPolytoma (vegetative cell cycle) preponderantly one mitochondrion is formed by fusion of those numerous mitochondria found after cell cleavage. Our observations will be compared with results on this subject in other organisms.We are greatly indebted to Prof. Dr. C. G.Arnold (Erlangen) for his support in this work.     

Mitochondria are involved in apoptosis induced by ultraviolet radiation in lepidopteran Spodoptera litura cell line

Abstract Mitochondria are involved in apoptosis of mammalian cells and even single‐cell organisms, but mitochondria are not required in apoptosis in cultured Drosophila cells such as S2 and BG2 cell lines. It is not very clear whether mitochondria are involved in apoptosis in other insect cells such as lepidopteran cell lines. Thus, we determined to elucidate the role of mitochondria in apoptosis induced by ultraviolet radiation in Spodoptera litura (Lepidoptera: Noctuidae) cell line (SL‐ZSU‐1). The Western blot results suggested that cytochrome c in the ultraviolet‐treated SL‐1 cells was released from the mitochondria to cytosol as early as 4 h after the induction of ultraviolet radiation and increased in the cytosolic fractions in a time‐dependent manner. Flow cytometric analysis of mitochondrial membrane potential (ΔΨm) of SL‐ZSU‐1 cell treated with ultraviolet‐C (UV‐C) light indicated the decrease in mitochondrial membrane potential was dependent on the times of ultraviolet treatment. Both of them are different from apoptosis in cultured Drosophila melanogaster cell lines (S2 and BG2) and it appears evident mitochondria are involved in apoptosis of the studied lepidopteran cells.     

Ricerche Sulle Infrastrutture Cellulari Dello Spadice Di Arum

Abstract

Electronmicroscope studies on fine structure of the ARUM spadix. — Three main stages of spadix development have been recognized. In the first stage — during wigh the growth would be embrional and by cell division, while respiration would sharply increase — new mitochondria are continously generated, mainly by the activity of long organules (chondriochontha). The morphology of these mitochondria is quite peculiar, showing bended cristae, often ring — or arc-shaped, and closed. In the second stage — during wich the growth would be by cell enlargment and respiration would further increase — the production of new mitochondria is decreasing, but thed are still numerous in each cell, and show a larger number of cristae (or tubules) than mitochondria observed in the first stage. A generalized vesicle production by the endoplasmic reticulum and the cristae of mitochondria is apparent at the beginning of the third stage — during wich the spadix gradually involves and respiration would fall down. Later, presumably in concurrence with the loss of cell vitality, the membranes of mitochopdria are disrupted, and all the cell structuree less evident. A great interest is presented by the vesicle production; indeed visicle production by the endoplasmic reticulum, and often also by mitochondria, can be observed in the senescent cells, already present and scattered in the tissues of the very young spadices. This ultimate process is howerer often preceded by the re-absorption of the outer membranes of these organules, in connection of the cristae, wich therefore become opened and in communication with the cell cavity.     

Trichomonas vaginalis and early evolving DNA and protein sequences of the CDC2/28 protein kinase family

The human sexually transmittted parasite Trichomonas vaginalis is a representative of one of the three earliest evolving eukaryotic lineages. We investigated whether T. vaginalis has DNA sequences and peptides related to cell division control molecules universal among yeasts and higher eukaryotes. A T. vaginalis ceil division control (CDC2/28) homologue was amplified by the polymerase chain reaction and sequenced. The absolute similarity with other CDC2/28 genes was 47%, with conservative replacement similarity of 67%. Western blots demonstrated a single T. vaginalis peptide reactive with antiserum to the PSTAIRE peptide, an expressed component of CDC2/28 genes in higher eukaryotes. Although eukaryotic, T. vaginalis has properties similar to those of bacteria and is the earlist evolving eukaryote reported to possess CDC2/28 DNA and peptide homologues. These observations suggest that the molecular origins of cell division control in eukaroytes preceded mitochondria, 28S ribosomes and regulated glycolysis.     

Observations of mitochondria and mitochondrial nuclei by double staining with rhodamine-123 and DAPI in synchronous cultures ofCatharanthus roseus

Mitochondria in cells ofCatharanthus roseus (L.) G. Don in synchronous cell division cultures were observed by double staining using fluorescence microscopy. The cells were stained with 4′-6-diamidino-2-phenylindole (DAPI) first and subsequently stained with rhodamine 123 (r-123). Immediately after staining with r-123, yellowishgreen, elongated and moving mitochondria were observed upon excitation at 485 nm. When the excitation filters were replaced by a UV filter (360 nm), 1 to 7 mitochondrial nucleoids were visible in each mitochondrion in the same field. Changes in the lengths of mitochondria during the cell cycle obtained from the observations under fluorescence microscopy by this staining method suggest the occurrence of multiplication of mitochondria concurrent with the cell cycle ofC. roseus.     

Hydrogen peroxide induces caspase activation and programmed cell death in the amitochondrial Tritrichomonas foetus

Tritrichomonas foetus is an amitochondrial parasite protist which lacks typical eukaryote organelles such as mitochondria and peroxisomes, but possesses the hydrogenosome, a double-membrane-bound organelle that produces ATP. The cell death of amitochondrial organisms is poorly studied. In the present work, the cytotoxic effects of hydrogen peroxide on T. foetus and its participation on cell death were analyzed. We took advantage of several microscopy techniques, including videomicroscopy, light microscopy immunocytochemistry for detection of caspase activation, and scanning and transmission electron microscopy. We report here that in T. foetus: (1) H₂O₂ leads to loss of motility and induces cell death, (2) the dying cells exhibit some characteristics similar to those found during the death of other organisms, and (3) a caspase-like protein seems to be activated during the death process. Thus, we propose that, although T. foetus does not present mitochondria nor any known pathways of cell death, it is likely that it bears mechanisms of cell demise. T. foetus exhibits morphological and physiological alterations in response to H₂O₂ treatment. The hydrogenosome, a unique organelle which is supposed to share a common ancestral origin with mitochondria and has an important role in oxidative responses in trichomonads, is a candidate for participating in this event.Abbreviations TUNEL Terminal deoxyribonucleotide transferase (TdT)-mediated dUTP nick-end labeling - PARP Poly (ADP-ribose) polymerase - DAPI 4,6-Diamidino-2-phenylindole dihydrochloride     

The effects of mesenchymal stem cell mitochondrial transplantation on doxorubicin‐mediated nephrotoxicity in rats

The effect of dysfunctional mitochondria in several cell pathologies has been reported in renal diseases, including diabetic nephropathy and acute kidney injury. Previous studies have reported that mitochondrial transplantation provided surprising results in myocardial and liver ischemia, as well as in Parkinson's disease. We aimed to investigate the beneficial effects of isolated mitochondria transplantation from mesenchymal stem cells (MSCs) in vivo, to mitigate renal damage that arises from doxorubicin‐mediated nephrotoxicity and its action mechanism. In this study, a kidney model of doxorubicin‐mediated nephrotoxicity was used and isolated mitochondria from MSCs were transferred to the renal cortex of rats. The findings showed that the rate of isolated mitochondria from MSCs maintains sufficient membrane integrity, and was associated with a beneficial renal therapeutic effect. Following doxorubicin‐mediated renal injury, isolated mitochondria or vehicle infused into the renal cortex and rats were monitored for five days. This study found that mitochondrial transplantation decreased cellular oxidative stress and promoted regeneration of tubular cells after renal injury (P < .001, P = .009). Moreover, mitochondrial transplantation reduced protein accumulation of tubular cells and reversed renal deficits (P = .01, P < .001). Mitochondrial transplantation increased Bcl‐2 levels, and caspase‐3 levels decreased in injured renal cells (P < .015, P < .001). Our results provide a direct link between mitochondria dysfunction and doxorubicin‐mediated nephrotoxicity and suggest a therapeutic effect of transferring isolated mitochondria obtained from MSCs against renal injury. To our knowledge, this study is the first study in the literature that showed good therapeutic effects of mitochondrial transplantation in a nephrotoxicity model, which is under‐researched.     

On the Organization of the Naked Filose Amoeba,Nuclearia moebiusi Frenzel, 1897 (Sarcodina,Filosea) and Its Implications1

The ultrastructural organization of Nuclearia moebiusi is described. The organism has filose pseudopodia without supportive microtubules; it has mitochondria with flattened cristae, lacks extrusomes, microtubule-organizing centers, and cytoplasmic microtubules. During cell division, microtubules appear only within the nucleus, and reorganization of the nuclear envelope occurs late in the nuclear division cycle. Ultrastructural studies reveal that Nuclearia spp. have a pattern of cellular organization distinct from that of other amoebae. Comparison of ultrastructural features suggests that this organism is only distantly related to other rhizopod amoebae (including other filose amoebae) and to the heliozoa.