Anaerobic peroxisomes in Mastigamoeba balamuthi

The adaptation of eukaryotic cells to anaerobic conditions is reflected by substantial changes to mitochondrial metabolism and functional reduction. Hydrogenosomes belong among the most modified mitochondrial derivative and generate molecular hydrogen concomitant with ATP synthesis. The reduction of mitochondria is frequently associated with loss of peroxisomes, which compartmentalize pathways that generate reactive oxygen species (ROS) and thus protect against cellular damage. The biogenesis and function of peroxisomes are tightly coupled with mitochondria. These organelles share fission machinery components, oxidative metabolism pathways, ROS scavenging activities, and some metabolites. The loss of peroxisomes in eukaryotes with reduced mitochondria is thus not unexpected. Surprisingly, we identified peroxisomes in the anaerobic, hydrogenosome-bearing protist Mastigamoeba balamuthi. We found a conserved set of peroxin (Pex) proteins that are required for protein import, peroxisomal growth, and division. Key membrane-associated Pexs (MbPex3, MbPex11, and MbPex14) were visualized in numerous vesicles distinct from hydrogenosomes, the endoplasmic reticulum (ER), and Golgi complex. Proteomic analysis of cellular fractions and prediction of peroxisomal targeting signals (PTS1/PTS2) identified 51 putative peroxisomal matrix proteins. Expression of selected proteins in Saccharomyces cerevisiae revealed specific targeting to peroxisomes. The matrix proteins identified included components of acyl-CoA and carbohydrate metabolism and pyrimidine and CoA biosynthesis, whereas no components related to either β-oxidation or catalase were present. In conclusion, we identified a subclass of peroxisomes, named “anaerobic” peroxisomes that shift the current paradigm and turn attention to the reductive evolution of peroxisomes in anaerobic organisms.

Sperm ultrastructure in Culmenella rezvoji (Lindholm, 1929) (Gastropoda: Hygrophila)

The taxonomic position of the genus within Hygrophila remains uncertain. The spermatozoa of , a species from the Far East of Russia, were examined using a combination of light, scanning and transmission electron microscopy with the objective to assess the utility of sperm characters for clarifying the phylogenetic relationships of the genus. The spermatozoa of C. rezvoji are divided into four regions: head, midpiece, glycogen piece and endpiece. The head contains a slender, cone-shaped acrosome and a conical nucleus with a sinistrally coiled keel. The acrosome consists of an apical vesicle and a thick-walled pedestal with an electron lucent canal partially filled with a patchy electron-dense material. The midpiece contains the mitochondrial derivative that encloses apically three parallel glycogen-filled tracts (helices) positioned in such a way that in the sperm cross section two helices lie opposite each other and equidistant from the third helix. The surface of the sperm above one of the helices forms a high, narrow ridge; the ridges above the remaining two helices have a much lower profile. The boundary between the midpiece and glycogen piece is demarcated by a constriction (annulus) consisting of an anterior electron-dense ring and a posterior conical cylinder connected to the ring with thin filaments. The structure of spermatozoa in Culmenella is consistent with the general pattern of sperm morphology common to all studied species of Hygrophila, but the spermatozoa of Culmenella also have distinctive characters (three glycogen helices and high-profile surface ridge in the apical portion of the midpiece) that should be potentially useful in resolving the taxonomic position of this genus.

The cytochemical localization of succinic dehydrogenase in the sperm mitochondria of the gastropod Biomphalaria glabrata. A comparison of two methods.

The mitochondrial derivative of the sperm of the gastropod pulmonate Biomphalaria glabrata was studies to ascertain succinic dehydrogenase localization cytochemically. Two techniques were compared. One technique depends on a tetrazolium salt that yields an osmiophilic formazan upon reduction. The other technique is dependent on the reduction of copper ferricyanide. The effects of several electron transport inhibitors were studied. The reaction product observed in the matrix of the mitochondrial derivative using the former technique is sensitive to rotenone and is believed to be nicotinamide adenine dinucleotide-dependent. The reaction product observed in the intracristal spaces using the copper ferricyanide method is insensitive to rotenone and is believed to cytochemically demonstrate succinic dehydrogenase in this material.

The Spermatozoon of Arthropoda

The spermatozoa of some species of Diptera belonging to the Psychodidae are described. In the 4 species examined, they appear needle-shaped, non-motile, anteriorly flattened, posteriorly bifid. Flagella, and any microtubule system, are lacking, and the only organelles encountered are an extremely elongated nucleus, a prominent acrosome, a well developed subacrosomal body, a slender, somewhat elongated mitochondrial derivative and a conspicuous multilayered membrane wall, produced by the superimposition of the plasma membrane and the acrosomal membrane. The mitochondrion is devoid of a crystalline axis, while the acrosome contains a longitudinal, cross-striated para-crystalline body. Centrioles are absent throughout the spermatid stage. Sperms and spermatids are organized in syncytia of 210 elements.

THE CYTOCHEMICAL LOCALIZATION OF GLYCOLYTIC AND OXIDATIVE ENZYMES WITHIN MITOCHONDRIA OF SPERMATOZOA OF SOME PULMONATE GASTROPODS

In this electron microscopic cytochemical study, the periodic acid-thiosemicarbazide-silver proteinate procedure was used to demonstrate glycogen stores within the mitochondrial derivative of sperm of pulmonate gastropods. In the presence of phenazine methosulfate and tetrazolium salt, enzymatic activity for glyceraldehyde 3-phosphate dehydrogenase and lactate dehydrogenase is shown in the matrix and in the compartment containing glycogen, but in the absence of phenazine methosulfate, tetrazolium reductase activity in the matrix is emphasized. Activity for NADH2-tetrazolium reductase and succinate dehydrogenase is also demonstrated in the matrix. Using 3,3'-diaminobenzidine tetra-HCl, cytochrome c oxidase activity is shown in the paracrystalline mitochondrial structure. The interrelation between glycolytic and oxidative pathways in this highly compartmentalized mitochondrion is considered.

LOCALISATION MITOCHONDRIALE D'ENZYMES LIEES AU METABOLISME DU GLYCOGENE DANS LE SPERMATOZOIDE DE L'ESCARGOT

The mitochondrial derivative of the sperm cell of the Gastropod Helix is endowed with a compartment loaded with glycogen (Personne et André, 1964). The aim of this work is to establish whether this mitochondrial glycogen is synthesized in the mitochondrial derivative itself or elsewhere in the cell. For this purpose, living sperm were first incubated in a medium containing glucose 1-phosphate, then fixed, sectioned, and stained specifically for polysaccharides by the phosphotungstic acid technique or the periodic acid-thiosemicarbazide-silver proteinate technique. Comparison with controls shows that a synthesis of glycogen occurred during the time of incubation. It can be inferred from this result that an amylophosphorylase system controlling the metabolism of polysaccharides is present in the mitochondrial derivative itself. Results obtained with the iodine technique indicate that the original glycogen molecules are elongated during the experiment. It seems probable that the amylophosphorylase system demonstrated here accounts for at least a part of the proteinaceous coat morphologically detectable around each individual glycogen particle.

OBSERVATIONS ON SPERMIOGENESIS IN THE FUNGUS GNAT SCIARA COPROPHILA

Although 9-membered centrioles are found in somatic tissues of Sciara, the centriole which lies at the spindle pole of the second meiotic division in male Sciara is composed of a row of approximately 70 short tubules in an oval array. Shortly after telophase of this unequal division, in the daughter cell destined to undergo spermiogenesis, microtubules become confluent with the tubules of the centriole. These tubules have the same density as other cytoplasmic microtubules after glutaraldehyde-OsO4 fixation and, like them, are not preserved with OsO4 fixation. As the centriole, now with approximately 70 attached, posteriorly directed, doublet tubules, migrates from the polar to the apolar end of the nucleus to take a final position in an oval groove which forms in the nuclear envelope, the tubules lengthen and become demonstrable after OsO4 fixation and more electron opaque than other cytoplasmic microtubules following glutaraldehyde-OsO4 fixation. Later, a singlet tubule appears peripherally to each doublet of the oval and 4 "arms" develop at specific sites on the tubules. Posteriorly, where the oval of tubules becomes discontinuous and forms a spiral, the arrangement of arms is different and the singlet tubules are lacking. Dense solid bodies develop inside this odd flagellum and become enclosed by a smooth double membrane. A single mitochondrial derivative has three components: a central area of homogeneous, moderately electron-opaque, proteinaceous material; a peripheral ring of cristae; and a crystalloid which is specifically oriented with respect to the flagellar tubules.