Types Of Microbodies Research Articles

A proteomic approach towards the identification of the matrix protein content of the two types of microbodies in Neurospora crassa

Microbodies (peroxisomes) comprise a class of organelles with a similar biogenesis but remarkable biochemical heterogeneity. Here, we purified the two distinct microbody family members of filamentous fungi, glyoxysomes and Woronin bodies, from Neurospora crassa and analyzed their protein content by HPLC/ESI-MS/MS. In the purified Woronin bodies, we unambiguously identified only hexagonal 1 (HEX1), suggesting that the matrix is probably exclusively filled with the HEX1 hexagonal crystal. The proteomic analysis of highly purified glyoxysomes allowed the identification of 191 proteins. Among them were 16 proteins with a peroxisomal targeting signal type 1 (PTS1) and three with a PTS2. The collection also contained the previously described N. crassa glyoxysomal matrix proteins FOX2 and ICL1 that lack a typical PTS. Three PTS1 proteins were identified that likely represent the long sought glyoxysomal acyl-CoA dehydrogenases of filamentous fungi. Two of them were demonstrated by subcellular localization studies to be indeed glyoxysomal. Furthermore, two PTS proteins were identified that are suggested to be involved in the detoxification of nitroalkanes. Since the glyoxysomal localization was experimentally demonstrated for one of these enzymes, a new biochemical reaction is expected to be associated with microbody function.

Dynamin‐like protein‐dependent formation of Woronin bodies in Saccharomyces cerevisiae upon heterologous expression of a single protein

Filamentous ascomycetes harbor Woronin bodies and glyoxysomes, two types of microbodies, within one cell at the same time. The dominant protein of the Neurospora crassa Woronin body, HEX1, forms a hexagonal core crystal via oligomerization and evidence has accumulated that Woronin bodies bud off from glyoxysomes. We analyzed whether HEX1 is sufficient to induce Woronin body formation upon heterologous expression in Saccharomyces cerevisiae, an organism devoid of this specialized organelle. In wild-type strain BY4742, initial import of HEX1 into existing peroxisomes enabled the formation of organelles with a hexagonal crystal. The observed structures mimicked the shape of genuine Woronin bodies, but exhibited a lower density and were significantly larger. Double-immunofluorescence analysis revealed that hexagonal HEX1 structures only occasionally co-localized with peroxisomal marker proteins, indicating that the Woronin-body-like structures are well separated from peroxisomes. In cells lacking Vps1p and Dnm1p, dynamin-like proteins required for the division of peroxisomes, the Woronin-body-like organelles remained attached to peroxisomes. The data indicate that Woronin bodies emerge after the formation of a HEX1 core crystal within peroxisomes followed by Vps1p- and Dnm1p-mediated fission.

Lysosomes and sulfide-oxidizing bodies in the bacteriocytes of Lucina pectinata , a cytochemical and microanalysis approach

Lucina pectinata is a large tropical clam living deeply burrowed in the black, reducing mud of mangrove swamps. It is known to possess hemoglobin in the cytoplasmic areas of its bacteriocytes, which harbor sulfide-oxidizing bacteria. The bacteriocytes also possess lysosome-like microbodies containing either membrane whorls or electron-dense granules in which free heme compounds have been identified. The cytochemical detection of acid phosphatase and arylsulphatase through EDX (energy-dispersive X-ray) microanalysis strongly suggests that the bacteriocytes of L. pectinata contain, in fact, two different types of microbodies. Some of these (devoid of dense granules) possess a variable amount of lysosomal enzymes and occasionally a limited quantity of iron, which may result from a recycling process of hemoglobin. Their main function seems to be the digestion of a limited proportion of symbiotic bacteria. They represent genuine secondary lysosomes with a functionally acidic pH. The second type of microbodies is characterized by dense granules containing sulfur and iron hemes but no lysosomal enzymes. Their sulfide-oxidizing activity was substantiated by benzyl viologen assay, with Na2S as a substrate. These microbodies appear to be similar to the sulfide-oxidizing bodies (SOBs) described in the bacteriocytes of other bivalve species with symbiotic thioautotrophic bacteria; however, their sulfide-oxidizing activity appears to be non-enzymatic. They are discrete organelles, characterized by a functionally basic pH and pseudoperoxidasic activity, and have been termed SOBs. Therefore, the bacteriocytes of L. pectinata possess at the same time functional lysosomes and functional SOBs.

Import of a Cytosolic Protein into Lysosomes by Chaperone-mediated Autophagy Depends on Its Folding State

Import of a Cytosolic Protein into Lysosomes by Chaperone-mediated Autophagy Depends on Its Folding State

Characterization of Aspergillus nidulans peroxisomes by immunoelectron microscopy.

In previous work, we have demonstrated that oleate induces a massive proliferation of microbodies (peroxisomes) in Aspergillus nidulans. Although at a lower level, proliferation of peroxisomes also occurs in cells growing under conditions that induce penicillin biosynthesis. Here, microbodies in oleate-grown A. nidulans cells were characterized by using several antibodies that recognize peroxisomal enzymes and peroxins in a broad spectrum of eukaryotic organisms such as yeast, and plant, and mammalian cells. Peroxisomes were immunolabeled by anti-SKL and anti-thiolase antibodies, which suggests that A. nidulans conserves both PTS1 and PTS2 import mechanisms. Isocitrate lyase and malate synthase, the two key enzymes of the glyoxylate cycle, were also localized in these organelles. In contrast to reports of Neurospora crassa, our results demonstrate that A. nidulans contains only one type of microbody (peroxisomes) that carry out the glyoxylate cycle and contain 3-ketoacyl-CoA thiolase and proteins with the C-terminal SKL tripeptide.

The role of peroxisomes in ether lipid synthesis. Back to the roots of PAF.

Peroxisomes belong to a family of subcellular organelles called microbodies that play indispensable roles in cellular metabolism in protozoa, fungi, plant and mammalian cells’. These organelles show a greater variety in enzyme content than other subcellular organelles. Well over 50 enzymes have been reported to be located in microbodies of some kind, but no microbody contains all of them. This is illustrated in table I for a few major microbody processes. Whereas fatty acid [3-oxidation is present in all microbody types, various oxidases producing H202 and catalase are not present in glycosomes, the microbody originally discovered in Trypanosomatidae 2.The glyoxylate cycle is only found in the microbodies of plant seeds and those of yeast and Tetrahymena. Photorespiration and glycolysis are restricted to only one type of microbody, i.e. plant leaf peroxisomes and glycosomes, respectively. Ether lipid synthesis is confined to mammalian peroxisomes3 and, as we recently demonstrated, to trypanosomal glycosomes4, but is not found in plant and yeast microbodies.

Microbodies in fungi: a review

Microbodies are ubiquitous organelles in fungal cells, occurring in both vegetative hyphae and spores. They are bounded by a single membrane and may contain a crystalloid inclusion with subunits spaced at regular intervals. Typically, they contain catalase which reacts with the cytochemical stain 3,3′-diaminobenzidine to yield an electron-opaque product, urate oxidase,l-α-hydroxy acid oxidase andd-amino acid oxidase. Their fragility and the necessity to disrupt the tough fungal cell wall before isolating them make them difficult to isolate. Analysis of enzymes in purified or partially purified microbodies from fungi indicates that they participate in fatty acid degradation, the glyoxylate cycle, purine metabolism, methanol oxidation, assimilation of nitrogenous compounds, amine metabolism and oxalate synthesis. In organisms where microbodies are known to contain enzymes of the glyoxylate cycle, they are known as glyoxysomes; where they are known to contain peroxidatic activity, they are known as peroxisomes. In some cases microbodies contain enzymes for only a portion of a pathway or cycle. Thus, they must be involved in metabolic cooperation with other organelles, particularly mitochondria. The number, size and shape of microbodies in cells, their buoyant density and their enzyme contents may vary with the composition of the medium; their proliferation in cells is regulated by the growth environment. The isolation from the same organism of microbodies with different buoyant densities and different enzymes suggests strongly that more than one type of microbody can be formed by fungi.

Occurrence, characterization and development of two different types of microbodies in the nematophagous fungusArthrobotrys oligospora

The occurrence of microbodies in different cells of the nematophagous fungus Arthrobotrys oligospora has been investigated. In the predacious phase this organism forms complex 3-dimensional network traps. Mature trap cells generally were crowded with “special” microbodies which possessed an electron dense matrix and were surrounded by a membrane of approx. 9 nm. These organelles developed during the early stages of trap formation and were derived from specialized regions of the endoplasmic reticulum. Cytochemical staining experiments revealed that the electron-dense microbodies contained catalase and d-amino acid oxidase and thus must be considered peroxisomal in nature. Electron-dense bodies were absent in normal vegetative cells of the fungus. These cells contained “normal” microbodies which developed from each other by the separation of small organelles from mature ones. As in yeasts, the metabolic function of these latter organelles was dependent upon environmental conditions.

Occurrence, characterization and development of two different types of microbodies in the nematophagous fungus Arthrobotrys oligospora

Occurrence, characterization and development of two different types of microbodies in the nematophagous fungus Arthrobotrys oligospora

Developmental studies of NAD-malate dehydrogenase isozymes in the cotyledon of cucumber seedlings grown in darkness and in light

The total activity of NAD-malate dehydrogenase (E.C. 1.1.1.37; MDH) in the cotyledons of cucumber (Cucumis sativus L.) seeds and seedlings increased 10fold during the first 3 days of germination in darkness and then declined gradually to one third of the peak activity after 10 days. Exposure of the seedlings to light at day 3 sharply reduced the activity to one third of the peak activity within 1 day but the residual activity remained fairly constant from day 4 to day 10. In extracts of cotyledons of 2-day-old, dark-grown seedlings and of cotyledons of 6-day-old, light-grown seedlings MDH activity was resolved into 5 isozymes by starch gel electrophoresis. Three of these isozymes were identified as cytosol isozymes, one each as a mitochondrial isozyme and a microbody (glyoxysome or peroxisome) isozyme. During germination in darkness, the activities of the five isozymes increased at different rates. However, they all peaked together at day 3 and then declined gradually at a similar rate. Light applied at day 3 selectively eliminated two cytosol isozymes but did not affect the subsequent developmental pattern of the third cytosol isozyme, the mitochondrial isozyme and the microbody isozyme. Such a selective elimination of two cytosol isozymes, together with their absence in roots and green leaves, indicates that these two isozymes participate in the mobilization of food reserves during germination. The glyoxysomal and peroxisomal isozymes were not resolved in starch gel electrophoresis. Their similarities are discussed in connection with the relationship of the two enzymes and of the two types of microbodies during the greening process.

Microbodies of the rat renal proximal tubule: ultrastructural and cytochemical investigations.

The present study was undertaken to provide a detailed morphological and cytochemical characterization of the microbodies of the rat renal proximal tubule following perfusion fixation. The following observations were made: 1) Two basic types of microbodies (Mb-I and Mb-II) can be identified. Mb-I have both circular and tubular profiles which are located peripherally within the granular matrix of these microbodies. Mb-II have marginal plates and crystalloid inclusion in addition to circular and tubular profiles. 2) Circular and tubular profiles, 100 nm in diameter, described by previous investigators as being infrequent in occurrence, are the most consistent morphological characteristic of rat renal microbodies after perfusion fixation. These profiles have a homogeneous center surrounded by a double or single ring of granules. The uniform size and spacing of these granules within profiles establish a basic 100 A periodicity found in both types of microbodies. 3) Evidence is presented which suggests that both "nucleoids" and "tubular protrusion rods" as described by other investigators of the rat renal microbodies may result from poor fixation and/or osmotic stress. 4) The density of the matrix of Mb-I is, in contrast to previous reports, greater than the density of adjacent mitochondria. 5) Marginal plates or crystalloid inclusions were demonstrated in some microbodies (Mb-II) of all the rats studied; periodicities of 100, 200, and 300 A were identified within these structures. 6) Both types of microbodies were positive for catalase activity, but were negative for acid phosphatase activity. On the basis of both morphological and cytochemical criteria, it seems plausible that these two populations of renal microbodies (Mb-I and Mb-II) represent a morphological and functional continuum.

Fine structural and cytochemical analysis of phragmosomes (microbodies) during cytokinesis inAllium root tip cells

The ontogeny and distribution of phragmosomes (microbodies) during cytokinesis inAllium sativum root tip cells have been studied and complemented with a cytochemical analysis of reactivity with diaminobenzidine (DAB). Incubation in different DAB media revealed the presence of catalase but not peroxidase in these organelles, identifying them as a type of microbody associated with the forming cell plate. Only vacuoles, segments of endoplasmic reticulum and portions of the mature walls stained positively with DAB for peroxidase activity. Microbodies begin to appear in the region of the future cell plate as cells enter late anaphase. They exhibit a moderately electron-opaque anucleoid matrix and are continuous with segments of endoplasmic reticulum (ER). Certain observations have led us to consider that certain aspects of plate formation inAllium require the participation of microbodies: (a) their pronounced numerical increase at the onset of plate formation, (b) their intimate association with regions of the plate where vesicle fusion is in progress, and (c) their rapid numerical decline following vesicular fusion and concomitant cell plate formation. The characteristic spatial association observed between microbodies and the plate-forming vesicles may well reflect their mutual involvement in the metabolism of carbohydrates comprising the middle lamella, being coordinated by metabolic activities in the cytosol, mitochondria and dictyosomes.

Ultrastructure of the zoospores of Entophlyctis confervae-glomeratae, Rhizophydium patellarium and Catenaria anguillulae

The fine structure of zoospores of Entophlyctis confervae-glomeratae, Rhizophydium patellarium, and Catenaria anguillulae has been described and compared. The zoospores of E. confervae-glomeratae contain many posteriorly located mitochondria in a petal-like arrangement. On the inner (proximal) side of a kinetosome are two electron-dense, slightly curved bands with the convex side directed inward towards the eccentrically located nucleus. Microtubules radiate from the convex side of the bands. Two or three lipid bodies are seen usually in the anterior region of the zoospore with microbodies often associated with them. Ribosomes are scattered throughout the cytoplasm. The zoospores of R. patellarium contain a single, bowl-shaped mitochondrion in the posterior region and a centrally located nucleus. Ribosomes are mainly clustered between the mitochondrion and the nucleus and are partially enclosed by a double-membrane envelope. No microtubules are seen associating with the kinetosome and the non-functional centriole lies parallel to the kinetosome. In the anterior region there is a single lipid body as well as two types of microbodies: very electron-dense microbodies un-associated with any other organelle, and a finely granular microbody associated closely with the lipid body. The zoospores of C. anguillulae contain a single basal mitochondrion with the associated side-body complex. From the proximal region of the kinetosome, a set of 27 microtubules radiate anteriorly and surround the base of the cone-shaped nucleus. Ribosomes are enclosed within a distinct nuclear cap. Gamma bodies are present in the cytoplasm. Taxonomic implications resulting from this study are discussed.

Localization of peroxidase activity in microbodies of fetal mouse liver.

The peroxidase activity of microbodies in fetal mouse liver was studied by light and electron microscopy. Two types of microbodies were present; a small population of bodies that lacked a nucleoid, predominant on the 16th day of gestation, and a larger population of nucleoid-bearing microbodies, predominant on the 19th day, in association with the rough endoplasmic reticulum from which they probably originate. Both types of bodies were visualized when incubated for peroxidase activity but were negative (19th day) for acid phosphatase activity. The findings suggest that the anucleoid- and nucleoid-bearing organelles together constitute the microbody population of the fetal liver.