Mitochondrial Derivatives Research Articles

The length of a short sperm: Elongation and shortening during spermiogenesis in Cotesia congregata (Hymenoptera, Braconidae)

The spermatozoon of the parasitoid wasp Cotesia congregata is an extremely short gamete measuring less than 7 μm; it is as yet the shortest flagellated sperm to be identified. The mature sperm consists of an acrosome, surrounded by an extra cellular coat, a condensed nucleus, two uncoiled mitochondrial derivatives and a short axoneme. Testes of young adults contain a continuum of differentiation stages. Initially, the flagellum is approximately 5 μm long. It conserves its length in round, elongated and mature spermatids, but is reduced to less than 3 μm in mature spermatozoa. The nucleus is 2 μm in diameter when round, 10 μm long when it becomes a long boat-hull shaped filament, and then reduces to 3.6 μm. Thus, during development the gamete reaches a total length of 15 μm before finally reducing to less than half that length. Some traits of mature sperm anatomy are similar to related species of the Braconidae family, but others seem to be specific and could be due to the shortness of the cell. This uncommon elongation and subsequent shortening of such a tiny flagellated cell constitutes a model for both nucleus and cilium development.

Structural and ultrastructural characterization of the spermatozoa in Pachycondyla striata and P. marginata

This study characterizes the male gametic cells in two ant species (Pachycondyla striata and Pachnoda marginata) by light and transmission electron microscopies. The sperm of both species is composed of head, flagellum, and transition regions. The head consists of the acrosome, tapered and bilayer, and a cylindrical nucleus. In P. marginata, the intranuclear material is homogeneously compacted, and in P. striata, there are translucent inclusions and density variations. In the transition region of both species, centriolar adjunct, nuclear base, and the modified basal body (structure of the spermatozoa equivalent to the spermatids’ centriole) coexist and are progressively replaced by two oval and symmetrical mitochondrial derivatives and an axoneme 9 + 9 + 2, respectively, in addition to two symmetrical triangular accessory bodies. Moreover, cytochemical analysis with Ethanolic Phosphotungstic Acid Method (E-PTA) revealed species-specific features not previously reported in the Formicidae, such as the presence of E-PTA negative acrosomal vesicle, E-PTA positive perforatorium, and E-PTA positive nucleus, all in P. marginata. Mitochondrial derivatives exhibited distinct behavior in its internal composition, with E-PTA negative mitochondrial cristae and E-PTA positive paracrystalline in both species. Sperm morphology in Pachycondyla has marked morphological similarities with other ant groups, and its specific features can be useful tools to understand phylogenetic relationships within the Formicidae and Hymenoptera.

Structure and ultrastructure of spermatozoon in six species of Drosophilidae (Diptera).

The Drosophilidae family is formed by Brachycera Diptera distributed widely across different regions of the planet. It is composed of about 4000 species, 304 of which are found in Brazil. The objective of this work was to characterize morphologically the structure of the male internal reproductive apparatus and the ultrastructure of the spermatozoon in four Neotropical (Drosophila cardini, D. mercatorum, D. nebulosa and D. sturtevanti) and two invasive (D. simulans and Zaprionus indianus) species of drosophilids. The structural aspect of the internal reproductive apparatus corresponds with that described for other drosophilids; however, there are differences in the size and coloration of the structures, such as the testes, in each species analyzed. The spermatozoon of these species was seen to be long and fine, presenting morphological variation. The ultrastructure of the spermatozoon revealed that the morphological pattern is similar to that found in the majority of insects. The head region presents a nucleus with condensed chromatin and the acrosome positioned laterally to the nucleus. In the tail region, the axoneme presents the 9+9+2 pattern commonly described for other species of Diptera. The species presented differences regarding the shape and size of the mitochondrial derivatives. Cytochemical analysis using EPTA also revealed differences in terms of the location of the basic proteins in the mitochondrial derivates. The results obtained contribute to expanding the database for the Drosophilidae family, providing information that may contribute to intra- and inter-specific identification and supplying phylogenetic analyses.

Knockdown of Dynamitin in testes significantly decreased male fertility in Drosophila melanogaster

Dynamitin (Dmn) is a major component of dynactin, a multiprotein complex playing important roles in a variety of intracellular motile events. We previously found that Wolbachia bacterial infection resulted in a reduction of Dmn protein. As Wolbachia may modify sperm in male hosts, we speculate that Dmn may have a function in male fertility. Here we used nosGal4 to drive Dmn knock down in testes of Drosophila melanogaster to investigate the functions of Dmn in spermatogenesis. We found that knockdown of Dmn in testes dramatically decreased male fertility, overexpression of Dmn in Wolbachia-infected males significantly rescued male fertility, indicating an important role of Dmn in inducing male fertility defects following Wolbachia infection. Some scattered immature sperm with late canoe-shaped head distributed in the end of Dmn knockdown testis and only about half mature sperm were observed in the Dmn knockdown testis relative to those in the control. Transmission electron microscopy (TEM) exhibited fused spermatids in cysts and abnormal mitochondrial derivatives. Immunofluorescence staining showed significantly less abundance of tubulin around the nucleus of spermatid and scattered F-actin cones to different extents in the individualization complex (IC) during spermiogenesis in Dmn knockdown testes, which may disrupt the nuclear condensation and sperm individualization. Since dynein-dynactin complex has been shown to mediate transport of many cellular components, including mRNAs and organelles, these results suggest that Dmn may play an important role in Drosophila spermiogenesis by affecting transport of many important cytoplasmic materials.

Testis-Specific Bb8 Is Essential in the Development of Spermatid Mitochondria.

Mitochondria are essential organelles of developing spermatids in Drosophila, which undergo dramatic changes in size and shape after meiotic division, where mitochondria localized in the cytoplasm, migrate near the nucleus, aggregate, fuse and create the Nebenkern. During spermatid elongation the two similar mitochondrial derivatives of the Nebenkern start to elongate parallel to the axoneme. One of the elongated mitochondrial derivatives starts to lose volume and becomes the minor mitochondrial derivative, while the other one accumulates paracrystalline and becomes the major mitochondrial derivative. Proteins and intracellular environment that are responsible for cyst elongation and paracrystalline formation in the major mitochondrial derivative need to be identified. In this work we investigate the function of the testis specific big bubble 8 (bb8) gene during spermatogenesis. We show that a Minos element insertion in bb8 gene, a predicted glutamate dehydrogenase, causes recessive male sterility. We demonstrate bb8 mRNA enrichment in spermatids and the mitochondrial localisation of Bb8 protein during spermatogenesis. We report that megamitochondria develop in the homozygous mutant testes, in elongating spermatids. Ultrastructural analysis of the cross section of elongated spermatids shows enlarged mitochondria and the production of paracrystalline in both major and minor mitochondrial derivatives. Our results suggest that the Bb8 protein and presumably glutamate metabolism has a crucial role in the normal development and establishment of the identity of the mitochondrial derivatives during spermatid elongation.

A mitochondria-targeted colorimetric and two-photon fluorescent probe for biological SO2 derivatives in living cells

Two colorimetric and two-photon fluorescent probes (PCB and MPCB) for SO2 derivatives were designed and synthesized with rapid response, high selectivity, low detection limits, and large Stokes shifts. The probes displayed “turn on” fluorescence response to SO2 derivatives probably due to the nucleophilic addition to the electron-poor CN bond, which was supported by the DFT/TDDFT calculations. Compared with PCB, MPCB was obtained by incorporating a methyl pyridinium group, which endowed MPCB with the red-shifted spectra properties, better water-solubility, larger two-photon excitation action cross-section (δ′) and the important ability of locating in mitochondria. Bio-imaging study established that MPCB can be used to detect SO2 derivatives in living cells under two-photon excitation with large δ′ value (1551 × 0.125 = 193.88 GM), little cytotoxicity and good biocompatibility. Meanwhile, standard co-staining experiments of MPCB and Mito Tracker Red FM (co-localization coefficient: 0.89) revealed that MPCB can be used for the real-time sensing and bioimaging of SO2 derivatives in mitochondria.

Male reproductive system and sperm ultrastructure of Furcatopanorpa longihypovalva (Hua and Cai, 2009) (Mecoptera: Panorpidae) and its phylogenetic implication

The male reproductive system and sperm ultrastructure of the scorpionfly Furcatopanorpa longihypovalva (Hua & Cai) were investigated using light and transmission electron microscopy. A peculiar feature of the male reproductive system is the position of the epididymis, separately from the base of testis within the peritoneal sheath, unlike the species of other genera of Panorpidae pressed against the lateral base of testis. The mature spermatozoon is filiform, approximately 1600μm in length. It has largely maintained a mecopteran groundplan condition, with a bilayered acrosome, an elongate nucleus, a neck region, and a long flagellum. As for the sperm features, similarly to other species of Panorpidae, the nucleus has two lateral grooves and a helical appearance. The neck region shows a sheath-shaped centriolar adjunct, beneath the nucleus. The flagellum is helical and consists of a 9+2 axoneme, two mitochondrial derivatives of unequal size, one accessory body and two extra-axonemal accessory structures, as in the genus Panorpa of Panorpidae. The uniqueness of spermatozoa of F. longihypovalva lies in the specifically hammer-shaped mitochondrial derivative along most of the flagellum length. The phylogenetic position of the genus Furcatopanorpa in Panorpidae is briefly discussed based on our findings and literature in this respect.

The evolution of insect sperm − an unusual character system in a megadiverse group

Spermatozoa provide an unusual character system, with a limited number of components organized in a single cell. Similar spermatozoa occur in groups widely separated in the phylogenetic tree of Metazoa. Nevertheless, the character system contains phylogenetic information. Hexapoda have acquired spermatophores along with the switch from aquatic to terrestrial habitats, and related to this, a multitude of different sperm types. The aim of this study is a formal evaluation of the phylogenetic information content of spermatozoa. For the first time, sperm characters are coded for formal phylogenetic analyses. Different approaches are used and compared. Mainly due to a high level of homoplasy, the evaluation of sperm characters alone is insufficient for a reconstruction of the phylogeny of the group. Yet, a reliable reconstruction of the evolution of insect sperm is possible when character transformations are assessed using a phylogeny based on extensive molecular data. Important changes took place in the early evolution of Hexapoda. Sperm characters support some major clades (e.g. Hexapoda, Dicondylia, Polyneoptera, Psocodea), but important steps in the evolution are not reflected by transformations of spermatozoa, notably the rise of Pterygota or Holometabola. Important innovations are the formation of mitochondrial derivatives and the acquisition of accessory microtubules. Some features are conservative, whereas others evolved rapidly (e.g. presence or absence of the acrosome vesicle). Some groups are conservative in their sperm features (e.g. Odonata, Heteroptera), whereas the evolution of spermatozoa was distinctly accelerated in others (e.g. Ephemeroptera). The rate of evolution can change drastically in closely related groups. Profound changes in the morphologically uniform Zoraptera underline that sperm evolution can follow a pattern very different from the general somatic morphology. The mode of character reconstruction preferred here will be useful for the evaluation of specialized morphological character systems and strengthen the concept of evolutionary morphology.

Sperm ultrastructure and spermatodesm morphology of the spittle bug Locris transversa (Thunberg 1822) (Hemiptera: Cercopidae)

The structure of the spermatozoon and spermatodesm of the spittlebug Locris transversa (Thunberg 1822) was investigated by light and transmission electron microscopy. Males produced only one size class of sperm, which was 93–106 μm long. During spermatogenesis groups of spermatozoa are arranged around, and attached by their acrosomes to, a small central extracellular matrix to form a ball-shaped spermatodesm. Spermatodesmata were found in the testis, vas deferens and seminal vesicle of males and spermatheca of females. The sperm are filiform, each consisting of a ~15-μm-long head containing an anteriorly positioned conical ~2-μm-long acrosome and ~13-μm-long nucleus, and a midpiece and tail with a 9+9+2 axoneme. The acrosome, which has two posterior extensions that lie along one side of the anterior region of the nucleus, contains longitudinally orientated microfilaments. The nucleus has two unequal anterior extensions, whereas posteriorly it is flattened laterally to accommodate a putative centriolar adjunct and anterior ends of the two mitochondrial derivatives. The basal body lies posterior to the nucleus. The mitochondrial derivatives are elongated and extend for almost the entire length of the tail. The tail has glycogen within its centre, and towards its terminal end large deposits of glycogen surround the mitochondrial derivatives and axoneme. Compared to other members of the Cicadomorpha, the spermatozoa and spermatodesms of cercopids so far studied have some structural features in common with cicadellids and others in common with cicadas; therefore, if sperm features are to be useful in phylogenetic studies of cicadomorphs, attention must be given to examining key taxa to establish which character states are plesiomorphic or apomorphic.

Anatomy and Ultrastructure of Male Reproductive System in Two LeafhoppersCicadella viridisandKolla paulula(Hemiptera: Auchenorrhyncha: Cicadellidae)

The morphology and ultrastructure of the male reproductive system were investigated in two species of sharpshooter leafhoppers, Cicadella viridis (L.) and Kolla paulula (Walker), using light microscopy and transmission electron microscopy. The gross morphology and ultrastructure of the male reproductive system in the two species is similar to those of other Cicadellidae and related Auchenorrhyncha. The two species differ in the number and shape of testicular follicles, the shape of the seminal vesicle, and the size and color of the accessory gland. The testicular follicle consists of the epithelium lying on a thick basal lamina surrounded by a connective sheath, and a wide lumen filled with germ cells. Large numbers of vesicles and granular materials are observed in the epithelial cell. The two seminal vesicles close-set under a common tunica are composed of the basal lamina, the thick muscular-connective sheath, a mono-layered epithelium, and the vesicular lumen. The vesicles and granules are extensive in the cytoplasm of the flattened epithelial cell, whose apical surface usually has numerous microvilli. The accessory gland is made up of thin muscular-connective sheath, the basal lamina, the epithelium, and the tubular lumen. The epithelial cell with secretory granules has abundant rough endoplasmic reticulum and small vesicles, and its cytoplasm abounds with arrays of three types of granules. Spermiogenesis and sperm ultrastructure of K. paulula was also observed, whose sperm can be distinguished from those of other Cicadellidae by the accessory bodies and mitochondrial derivatives with gap region.

Identification of the active site of human mitochondrial malonyl-coenzyme a decarboxylase: A combined computational study.

Malonyl-CoA decarboxylase (MCD) can control the level of malonyl-CoA in cell through the decarboxylation of malonyl-CoA to acetyl-CoA, and plays an essential role in regulating fatty acid metabolism, thus it is a potential target for drug discovery. However, the interactions of MCD with CoA derivatives are not well understood owing to unavailable crystal structure with a complete occupancy in the active site. To identify the active site of MCD, molecular docking and molecular dynamics simulations were performed to explore the interactions of human mitochondrial MCD (HmMCD) and CoA derivatives. The findings reveal that the active site of HmMCD indeed resides in the prominent groove which resembles that of CurA. However, the binding modes are slightly different from the one observed in CurA due to the occupancy of the side chain of Lys183 from the N-terminal helical domain instead of the adenine ring of CoA. The residues 300 - 305 play an essential role in maintaining the stability of complex mainly through hydrogen bond interactions with the pyrophosphate moiety of acetyl-CoA. Principle component analysis elucidates the conformational distribution and dominant concerted motions of HmMCD. MM_PBSA calculations present the crucial residues and the major driving force responsible for the binding of acetyl-CoA. These results provide useful information for understanding the interactions of HmMCD with CoA derivatives. Proteins 2016; 84:792-802. © 2016 Wiley Periodicals, Inc.

Sperm morphology of Muscidifurax uniraptor (Hymenoptera: Chalcidoidea: Pteromalidae)

Sperm morphology of the parasitoid Muscidifurax uniraptor was investigated under light and transmission electron microscopy. M. uniraptor sperm are filiform, spiraled, approximately 150 μm in length, with a distinctive head, hooded by an extracellular sheath and a flagellum. This extracellular layer, from which many filaments radiate, measures approximately 90 nm in thickness and covers a small acrosome and the anterior nuclear region. The acrosome is composed of an acrosomal vesicle and a perforatorium with its base inserted in the nuclear tip. The nucleus is filled with homogeneously compacted chromatin. The centriolar adjunct extends towards the anterior portion in a spiral around the nucleus for 3.5 μm in length. The two mitochondrial derivatives begin exactly at the centriole adjunct base and, in cross-section, have a circular shape with equal areas that are smaller than the axoneme diameter. It is coiled, with 9 + 9 + 2 microtubules and begins from the centriole, just below the nuclear base. The axoneme is connected to the mitochondrial derivatives by two small irregularly shaped masses. Between the derivatives and the axoneme, the ‘center-flagellar material’ is observed. Overall, these characteristics are recognized in other Chalcidoidea, especially in the eurytomids, but together they form a set of species-specific data.

A Fluorescent Probe for Ratiometric Imaging of SO2 Derivatives in Mitochondria of Living Cells

When SO2 is inhaled through the respiratory tract into the body, it can be easily hydrated to sulfite (SO32–) and bisulfate (HSO3–) ions. They are widely used as essential preservatives for foods, beverages, and pharmaceutical products. Exposure to high doses of bisulfate induces a large number of respiratory diseases relevant to lung cancer, cardiovascular diseases, and many neurological disorders, as they involve in various physiology and pathological processes in mitochondria cell apparatus. In this work, a new fluorescent probe CY-SO2 for bisulfate/sulfite based on hemicyanine dye is reported, which can used in solution detection with 56-fold fluorescence ratio (F467 nm/F580 nm) enhancement, fast response (completed within 90 s) and excellent sensitivity (DL 2.67 nM). The effect have been exhibited in real sugar samples tests. CY-SO2 also displayed fluorescent imaging of breast cancer cells (MCF-7) successfully, demonstrating that the probe is a novel mitochondria-targeted ratiometric fluorescent prob...

First multigene analysis of Archamoebae (Amoebozoa: Conosa) robustly reveals its phylogeny and shows that Entamoebidae represents a deep lineage of the group

Archamoebae is an understudied group of anaerobic free-living or endobiotic protists that constitutes the major anaerobic lineage of the supergroup Amoebozoa. Hitherto, the phylogeny of Archamoebae was based solely on SSU rRNA and actin genes, which did not resolve relationships among the main lineages of the group. Because of this uncertainty, several different scenarios had been proposed for the phylogeny of the Archamoebae. In this study, we present the first multigene phylogenetic analysis that includes members of Pelomyxidae, and Rhizomastixidae. The analysis clearly shows that Mastigamoebidae, Pelomyxidae and Rhizomastixidae form a clade of mostly free-living, amoeboid flagellates, here called Pelobiontida. The predominantly endobiotic and aflagellated Entamoebidae represents a separate, deep-branching lineage, Entamoebida. Therefore, two unique evolutionary events, horizontal transfer of the nitrogen fixation system from bacteria and transfer of the sulfate activation pathway to mitochondrial derivatives, predate the radiation of recent lineages of Archamoebae. The endobiotic lifestyle has arisen at least three times independently during the evolution of the group. We also present new ultrastructural data that clarifies the primary divergence among the family Mastigamoebidae which had previously been inferred from phylogenetic analyses based on SSU rDNA.

Ultrastructural features of spermatozoa and their phylogenetic application in Zaprionus (Diptera, Drosophilidae)

ABSTRACTThe genus Zaprionus consists of approximately 60 species of drosophilids that are native to the Afrotropical region. The phylogenetic position of Zaprionus within the Drosophilidae family is still unresolved. In the present study, ultrastructural features of spermatozoa of 6 species of Zaprionus as well as the species Drosophila willistoni and Scaptodrosophila latifasciaeformis were analyzed. The ultrastructure revealed that the species have the same flagellar ultrastructure. Two mitochondrial derivatives, one larger than the other, close to the axoneme were present, primarily in D. willistoni (subgenus Sophophora). Except for Z. davidi and Z. tuberculatus, the analyzed species had paracrystalline material in both mitochondrial derivatives. Moreover, the testes showed 64 spermatozoa per bundle in all of the species. In the cluster analysis, 6 Zaprionus species were grouped closely, but there were some incongruent positions in the cladogram. The results indicated that sperm ultrastructure is an important tool for elucidating the phylogeny and taxonomy of insects.

A TP-FRET-based two-photon fluorescent probe for ratiometric visualization of endogenous sulfur dioxide derivatives in mitochondria of living cells and tissues.

A ratiometric two-photon fluorescent probe for SO2 derivatives was first proposed based on acedan-merocyanine dyads via a TP-FRET strategy. It was successfully applied to visualization of the fluctuations of enzymatically generated SO2 derivatives in the mitochondria of HepG2 cells and rat liver tissues using two-photon fluorescence microscopy imaging.

A peculiar new virus-spermatozoon association in the bug Raphigaster nebulosa (Poda) (Heteroptera-Insecta)

The sperm of the heteropteran bug Raphigaster nebulosa (Poda) are of two types, differing in length and size of their flagella. The thicker sperm are shorter than the thinner ones and have large mitochondrial derivatives. The presence of virus particles associated with the plasma membrane of thinner sperm is described for the first time; thicker sperm are immune to virus infection. The fact that virus particles are present on thinner sperm only initiates considerations on the transmission of virus.

The sperm structure and spermiogenesis of the heteropteran Coptosoma scutellatum (Geoffroy) with emphasis on the development of the centriole adjunct

The sperm structure and spermiogenesis of the morphologically peculiar heteropteran Coptosoma scutellatum has been examined with electron microscopy. The sperm has a short monolayer acrosome, a cylindrical nucleus, a 9+9+2 axoneme and two mitochondrial derivatives with crystallized matrix. It shows the synapomorphies typical of the group, consisting of (a) two crystallized areas in the mitochondrial derivatives; (b) the presence of two bridges connecting the intertubular material of the flagellar axoneme to the flattened cisterns adhering to the inner sides of the mitochondrial derivatives; (c) the absence of accessory bodies. The most interesting feature is the presence, in the early spermatid, of an extended centriole adjunct material with the shape of a large ribbon-like structure. This material surrounds the posterior nuclear region and extends anteriorly along the nucleus. It is a microtubule-organizing-center (MTOC) producing the many microtubules surrounding the sperm components At the end of spermiogenesis, the centriole adjunct material modifies into two kidney-like structures accompanying the nuclear posterior end and surrounding the modified centriole.

Giant spermatozoa of Diasemopsis (Diopsidae, Diptera) – Structural, ultrastructural and functional aspects

The spermatozoa of Diasemopsis comoroensis and Diasemopsis meigenii differ from the conventional brachyceran type in several respects. Not only are they very long but they are also extraordinarily wide, especially at the very end of the tail. The latter is effected by two cellular components: oversized mitochondrial derivatives and a prominent central band, which is a peculiar structure not known from any other spermatozoa. Based on its position with respect to the other organelles and its origin during early spermiogenesis, the central band is interpreted as a derivative of the centriolar adjunct material. Like the axoneme, the mitochondrial derivatives and the central band extend through the entire length of the spermatozoon tail. The spermatozoon tail is helical and can be coiled up into a tight cone-shaped spiral with a peculiar corkscrew shaped end portion. The potential adaptive significance of these features and their coevolution with the morphology of the female's multi-chambered ventral receptacle are discussed, as are sperm competition and cryptic female choice.

Testis Morphology and Dichotomic Spermatogenesis in the Sugarcane Borer Diatraea saccharalis (F.) (Lepidoptera: Crambidae).

In the present study, we describe the testis morphology and spermatogenesis of the sugarcane borer, one of the most destructive pests of sugarcane, using light, transmission, and scanning electron microscopies. Diatraea saccharalis (F.) larvae have paired, kidney-shaped testes that are limited externally and divided into four follicles by tunica cells. The adults have two fused whitish and spherical testes that are divided into eight follicles and are composed of spermatogonia, spermatocyte, spermatid, and sperm bundles. The spermatogonia and spermatocyte cysts are located at the apical extremity of the follicle, while the spermatid cysts and sperm bundles are located at the basal extremity of the follicle. Inside of the cysts, which are formed by the somatic cells, the germ line cells develop synchronously and are interconnected by cytoplasmic bridges. Each cyst contains only apyrene or eupyrene, and none of the cysts contain both of the cells. The inner part of the apyrene cyst consists of an electron-dense cap as opposed to a nucleus. The eupyrene sperm contains a nucleus, acrosome, and reticular and lacinate appendages. Both of the sperm types have a "9 + 9 + 2" axoneme and two mitochondrial derivatives at the tail region. The spermatogenesis process and spermatozoa morphology are similar to that of other Lepidoptera.