Günther F. Meyer

Differentiation of the synaptonemal complex and the kinetochore in Locusta spermatocytes studied by whole mount electron microscopy

When Locusta migratoria spermatocytes are surface-spread on various salines, the axial element of leptotene and zygotene chromosomes, and the synaptonemal complex of pachytene chromosomes are well-preserved, although, in most instances, virtually denuded of chromatin. A complex association of chromosome ends with the nuclear membrane is apparent as early as leptotene, and, as pairing is initiated, the nuclear attachment points of the partner half-bivalents fuse, apparently incorporating additional membrane material between them. The meiotic kinetochore originates in association with the axial element during early prophase, and prior to synaptonemal complex formation and chromosome condensation.

Genetic Activities of The Y Chromosome in Drosophila During Spermatogenesis

Publisher Summary This chapter focuses on genetic activities of the Y chromosome in Drosophila during spermatogenesis. The Y chromosome of Drosophila contains a number of fertility factors which are all essential for the formation of normally functioning sperm. These factors are cell autonomous and act even during the diploid phase, so that X-bearing spermatids that do not possess a Y chromosome are able in normal males to differentiate into normal spermatozoa. In spermatocyte nuclei of Drosophila species, special structures occur, which are not found in the nuclei of other tissues. They are restricted to the growth phase of primary spermatocytes. The functional nature of the Y-chromosomal loops has been demonstrated in two independent ways: (1) in autoradiographs the loops appear labeled after incubation with tritiated uridine, (2) if the DNA-dependent synthesis of RNA is blocked by actinomycin, the loops disintegrate within a few hours. Irreversible damage such as inhibition of loop regeneration and breaks within loops are induced by X-rays. Histochemical studies reveal the presence of proteins, RNA, and DNA in the loops. The chapter presents the morphogenetic effects of Y deficiencies on spermiogenesis. If a segment of the Y chromosome containing any of the loop-forming sites is lacking, males are invariably sterile.

The nucleolus in primary spermatocytes of Drosophila hydei

Transcribing ribosomal RNA genes in primary spermatocyte nucleoli of Drosophila hydei have been visualized by electron microscopy using a microspreading technique. The length of the transcribing unit is in agreement with the length of the ribosomal RNA precursor as determined by acrylamide gel electrophoresis (2.6×106 daltons). The length of the non-transcribed spacer is approximately 1.0×106 daltons. The maximum number of active cistrons in wild type (XY) males only approaches one half the estimated number of ribosomal cistrons of the replicated diploid genome (∼300) of D. hydei and is found to vary between 120 and 320 cistrons in different developmental stages of spermatocytes. There is also some variation between different males. In a few cases a variation in the transcriptional activity of different cistrons has been observed. The synthesis of ribosomal RNA therefore seems to be regulated primarily by the activation or inaetivation of varying numbers of ribosomal cistrons. Groups of adjacent cistrons seem to be under coordinated control. Inhibition of RNA synthesis by actinomycin, in contrast, follows a random pattern. The frequent observation of a bipartite nucleolus indicates that the nucleolus organizer regions of the two sex chromosomes are both active in transcription. The number of active ribosomal eistrons found in some X-Y translocation stocks and in XO males deviates considerably from that expected on the basis of DNA/RNA hybridization data. We conclude that, in agreement with the observations of other authors, a mechanism adjusting the number of ribosomal cistrons may be operating in these cases.

Interzelluläre Brücken (Fusome) im Hoden und im Ei-Nährzellverband von Drosophila melanogaster

Summary1.A detailed description is given of the structure of intercellular bridges connecting sister spermatogonia and spermatocytes in the testis as well as oocytes and nurse cells in the ovary of Drosophila melanogaster.2.The differentiated type of intercellular bridge found in the ovary is called „fusom“, using a term introduced by Hirschler (1948, 1953). The fusoms differ from simple intercellular bridges by the fact that the opening is more or less closed by a specific sealing substance.3.The sealing material seems to be produced by the stiff osmiophilic rim bordering the opening of the fusom.4.Presumably the aperture of the fusom is controled by factors originating in the nurse cells, since both closed and opened fusoms are found to connect the same oocyte with different nurse cells.

Spermiogenese in normalen und Y-defizienten Männchen von Drosophila melanogaster und D. hydei

SummarySpermiogenesis of X/Y males in Drosophila has been reinvestigated by light and electron microscopy. A description of normal spermiogenesis in both D. melanogaster and D. hydei, as well as spermatogenesis in Y deficient males of both species is provided. In D. hydei, spermatogenesis of X/O males does not proceed beyond meiosis. In D. melanogaster, the number of sperm in cysts of X/O testes may be reduced to less than half the normal number. Two different developmental patterns can be distinguished in X/O testes: In most cases early spermatid differentiation is severely disturbed so that the nebenkern disintegrates into many mitochondrial derivatives, each of them being incompletely transformed into paracrystalline material. Cysts with these deviations do not develop further than elongated spermatid stage. Less frequently, differentiation of the spermatid is relatively normal with almost complete transformation of the nebenkern derivatives into paracrystalline bodies. Even in such sperm, the geometrical relations in sperm tail organelles appear disturbed and development does usually not reach the stage of isolated sperm. In addition, many structural malformations in the axial complex are found. It seems that X/O spermatids in D. melanogaster contain all the structural components of normal sperm but fail to organize them properly.Morphogenetic processes in the developing spermatid of Drosophila hydei have been shown to be dependant upon the presence of regions of the Y chromosome which form specific lampbrush loops during growth stage of spermatocytes. In contrast to X/O males where spermatogenesis does not proceed beyond meiotic prophase, males with Y fragments produce aberrant spermatids and/or immotile, aberrant sperm. In one case, however, the production of slightly motile sperm in males deficient for a certain loop forming locus has been observed. Different Y segments (loops) differ in their morphogenetic capacity, but in general the effect of the loops seems to be additive and all five Y chromosomal factors must act together for the production of motile sperm. Developmental aberrations appearing during spermiogenesis in partially Y deficient males were classified into early and late effects and analysed. It appears that all structural components of sperm organelles (flagellum, nebenkern derivatives etc.) are present in the spermatids of all such males. The components, however, frequently fail to be organized into ordered complexes of typical architecture. The degree of order attained varies between cysts and also between sperm in the same cyst. It ranges from severe disorganization to near normality in a variable number of sperm. In addition there is a pronounced general effect of the Y, and of Y fragments on the length of sperm. It is concluded that the Y chromosomal factors control the coordination of the various synthetic and morphogenetic processes leading to the formation of functional sperm without themselves contributing structural information on the molecular level.Zusammenfassung1.Die Spermiennormogenese von D. melanogaster und D. hydei wird beschrieben. Nach Bildung der Nebenkernderivate fusionieren die Spermatiden bei beiden Arten. Aus dem Nebenkern gehen zwei Derivate hervor, von denen im Verlaufe des Längenwachstums bei D. melanogaster eines und bei D. hydei beide in kristallines Material transformiert werden. Die kristallinen Nebenkernderivate beginnen unmittelbar hinter dem Kern und sind so lang wie die Geißel (bei D. hydei über 6,5 mm). Die syncytiale Phase wird bis zum Ende der Spermiogenese beibehalten. Eine Trennung in Einzelspermien erfolgt erst nach der völligen Ausdifferenzierung kurz vor Erreichen der Motilität. Das reife Spermium ist nadeiförmig. Schwanz und Mittelstück sind nicht zu trennen.2.In X/O-Männchen von D. melanogaster beruht die Sterilität auf einer Störung der Differenzierung der Spermatiden. Neben frühen Defekten, die schon zu Beginn der Spermatidenentwicklung eintreten (z. B. bei der Nebenkern- und Akrosomdifferenzierung), kommen späte Entwicklungsstörungen vor, von denen bereits gestreckte Spermatiden betroffen werden. Sämtliche Komponenten der Spermatiden, d. h. Form und Größe des Zellkerns, Zahl und Größe der Nebenkernderivate, Zahl und geordnete Organisation der Geißelkomponenten, können in X/O-Spermatiden entweder verändert sein oder auch ganz fehlen. Sie können in manchen Spermatiden aber auch normal sein. Bei Zerfall des Nebenkernes bzw. seiner Derivate werden alle Fragmente in kristallines Material transformiert. In den Cysten von X/O-Hoden unterbleibt fast immer eine Trennung der syncytial verbundenen Spermienkomplexe in Einzelspermien.3.In X/O-Männchen von D. hydei ist die Spermatogenese bereits vor der 1. Reifeteilung blockiert. In partiell Y-defizienten Tieren können in Abhängigkeit vom Schleifentyp der aktiven Y-Loci frühe und späte Effekte auf die Differenzierung der Spermatiden unterschieden werden. Jede einzelne Schleife (bzw. Schleifenkombination) vermag den meiotischen Block in X/O-Männchen zu überwinden und als maximale Entwicklungsleistung die Synthese aller wesentlichen Spermatidenorganelle zu induzieren. Die morphogenetische Kapazität einzelner Schleifen und Schleifenkombinationen ist geringfügig verschieden und wurde im einzelnen untersucht und verglichen. Eine rein quantitative Abhängigkeit zwischen der Aktivität des Y-Chromosoms und der Länge der Spermien scheint gesichert. Die unvollständig differenzierten Spermien in Y-defizienten Männchen sind kürzer, die fertilen Spermien von Männchen mit Y-Duplikationen dagegen länger als normal. Es wird angenommen, daß die auf dem Y-Chromosom lokalisierten Faktoren neben qualitativen Funktionen auch quantitative Effekte auf das Spermatidenwachstum haben. Ihre Aufgabe scheint darin zu bestehen, regulierend oder induzierend auf bestimmte morphogenetische Prozesse während kritischer Phasen der Spermatiden-Differenzierung einzuwirken. Die möglichen molekularen Mechanismen ihrer Wirkungsweise werden diskutiert.

Chromatin elimination in the hypotrichous ciliate Stylonychia mytilus

Chromatin elimination in the hypotrichous ciliate Stylonychia mytilus was studied by means of electron microscopy using a microspreading procedure. In the polytene chromosomes of the macronuclear anlagen three organization patterns are observed: Bands of various size composed of 300 Å chromatin fibers, large blocks of 300 Å nucleofilaments which probably represent the “heterochromatic” regions of the chromosome and axial 120 Å filaments. Those DNA sequences which become eliminated belong to the 300 Å fiber type. The eliminated chromatin occurs in the form of rings of variable size corresponding to a DNA content between 18 and 160 Kb while the axial 120 Å filaments appear to be preserved.

Struktur-Differenzierungen im Y-Chromosom von Drosophila hydei und ihre Beziehungen zu Gen-Aktivitäten

Summary1.In the loops of primary spermatocyte nuclei actinomycin causes characteristic and reproducible disintegrations of the structural elements composing the loops. The alterations become visible for the first time about 6 hours after injection of actinomycin and reach a maximum after about 24–30 hours.2.The structure of the nucleolus is also changed by actinomycin. It looses its cortical “ribosomal” layer and detaches from the nuclear membrane to which it is normally attached.3.All structural damage of the loops and the nucleolus caused by actinomycin is reversible. The regeneration of the loops starts approximately 40 hours after injection and is completed after about 100–120 hours.4.In males with two Y chromosomes which form loops of different morphology because of genetical differences the actinomycin effects similar alterations as in normal males. During the regeneration each Y chromosome autonomously determines its specific type of loops.5.The results are in good agreement with our hypothesis that the loops are modifications of the chromosome structure at sites of active genes.

The formation of polytene chromosomes during macronuclear development of the hypotrichous ciliate Stylonychia mytilus

The formation of polytene chromosomes during macronuclear development of the ciliate Stylonychia mytilus was examined in spread electron microscopical preparations. The chromatin organization of early macronuclear anlagen closely resembles the organization of micronuclear chromatin. In the course of polytenization 300 Å chromatin fibers become organized in loop-like structures laterally attached to a thinner axial fiber. It is suggested that this reorganization of chromatin during polytenization is a necessary event for the subsequent chromatin elimination.

Die parakristallinen Körper in den Spermienschwänzen von Drosophila

Summary1.The transformation of the two mitochondrial nebenkern derivatives, characteristic for D. melanogaster, hydei and bifurca, into rod-shaped, dense bodies is described.2.After application of the negative staining procedure with PTA to intact, non-motile sperm, the rod-shaped bodies reveal a macromolecular architecture of a regular paracrystalline pattern. They are regularly cross-striated with an average period of 260 Å in D. melanogaster, 320 Å in D. bifurca and 300 and 400 Å respectively in D. hydei. X/0 sperms in D. melanogaster occasionally show a period of 100 Å only. The limited variability of the periodicity can be explained either as preparational artifact or as result of a multiplication of a smaller fundamental period or both.3.The rod-shaped bodies of motile sperm show characteristic modifications of the paracrystalline pattern and periodicity. Spermatozoa can be divided into two groups with homomorphic or heteromorphic rod-shaped bodies.4.Vital staining with Janus green demonstrates the participation of respiratory proteins in the formation of these bodies. Extraction experiments for paramyosin and myosin gave negative results.5.These rod-shaped bodies may function as a reservoir of respiratory proteins and/or as stabilizing elements for the extremely elongated spermatozoa in the genus Drosophila (6,6 mm in D. hydei). The rod-shaped bodies of Drosophila represent an extreme, special case of the wide spread phenomenon of total or partial transformation of mitochondrial nebenkern derivatives into paracrystalline structures.

Some facts concerning the nature and formation of axial core structures in spermatids of Gryllus domesticus

Examination of living spermatid nuclei of Gryllus domesticus has revealed the presence of the same structures, the X chromosome, the round body and the axial core structures, which have been described from electron microscopic observations. The outer ribbons of the axial core structures and the round body are composed of 100 Å fibres indiscernible from and often continuous with the fibres composing the X chromosome. That the outer ribbons of the axial core structures and the round body are chromosomal is further substantiated by the results of cytochemical examinations of formaldehyde fixed material which show that the axial core structures and the round body contain RNA, DNA and basic protein. Neither acetic acid-ethanol nor cold ethanol fixation preserve the round body and the axial core structures suggesting that a protein may be responsible for maintenance of the central core structure. The central core structures are always found in close association with condensed chromatin in regions where the chromosome elements are about 1000 Å apart, suggesting that the relative state of condensation of the chromatin and the spacial relationship between condensed regions may be two of the chief factors concerned in central core formation. Maintainance of the condensed state of the chromatin, however, may in turn depend upon central core integrity.