Targeting and fusion proteins during mammalian spermiogenesis.

Regulated exocytosis is controlled by internal and external signals. The molecular machinery controlling the sorting from the newly synthesized vesicles from the Golgi apparatus to the plasma membrane play a key role in the regulation of both the number and spatial location of the vesicles. In this context the mammalian acrosome is a unique vesicle since it is the only secretory vesicle attached to the nucleus. In this work we have studied the membrane trafficking between the Golgi apparatus and the acrosome during mammalian spermiogenesis. During bovine spermiogenesis, Golgi antigens (mannosidase II) were detected in the acrosome until the late cap-phase spermatids, but are not found in testicular spermatozoa (maturation-phase spermatids). This suggests that Golgiacrosome flow may be relatively unselective, with Golgi residents retrieved before spermination is complete. Surprisingly, rab7, a protein involved in lysosome/endosome trafficking was also found associated with the acrosomal vesicle during mouse spermiogenesis. Our results suggest that the acrosome biogenesis is associated with membrane flow from both the Golgi apparatus and the endosome/lysosome system in mammalian spermatids.     

Selenium in mammalian spermiogenesis

The role of selenium in male fertility is reviewed with special emphasis on selenoprotein P and phospholipid hydroperoxide glutathione peroxidase (GPx4) in spermiogenesis. Inverse genetics reveal that selenoprotein P is required for selenium supply to the testis. GPx4 is abundantly synthesized in spermatids. As a moonlighting protein it is transformed in the later stages of spermiogenesis from an active selenoperoxidase into a structural protein that becomes a constituent of the mitochondrial sheath of spermatozoa. The transformation is paralleled by loss of glutathione. Mechanistically, the process is an alternate substrate inactivation of GPx4 resulting from reactions of its selenenic form with thiols of GPx4 itself and other proteins. Circumstantial evidence and ongoing experimental genetics indicate that the mitochondrially expressed form of the GPx4 gene is the most relevant one in spermiogenesis, with the nuclear form being dispensable for fertility and the role of cytosolic GPx4 remaining unclear. Clinical data reveal a strong association of low sperm GPx4 with infertility. Thus, impaired GPx4 biosynthesis, due to selenium deficiency or to genetic defects in gpx4 itself or in proteins involved in Se distribution and selenoprotein biosynthesis, causes male infertility, but can also be an epiphenomenon due to any perturbation of testicular function.     

Spermatogenesis and spermiogenesis in Ascaris lumbricoides Var. suum

Reorganization of the prophase I nucleus marks the beginning of the first meiotic division. A pair of centrioles is present at each pole at metaphase I and mitochondria are not observed in the spindle area. A chromosomal pellicle, which resembles a kinetochore plate but has no apparent association with microtubules, surrounds each autosome at metaphase I and II. The sex body lags behind the autosomes at anaphase I and segregates differentially to one daughter cell. Mitochondria and a pair of centrioles are present in the spindle during the second meiotic division. Localized condensation of chromatin and fusion of the condensed chromatin of the secondary spermatocyte telophase nucleus results in a compact spermatid nucleus. Loss of spermatid cytoplasm is effected by the ejection of a cytophore vesicle.     

Cytological aspects of oligochaete spermiogenesis

Spermiogenesis in Oligochaeta occurs with peculiar modalities, common to all the species studied in this group. Gonial cells, produced in the testes, drop into coelomatic cavities (usually seminal vesicles) where they undergo a series of mitotic divisions, without cytodieresis, and finally meiosis. In the seminal vesicles, a series of morulae is present, composed of 2, 4, 8, 18, 32, ... cells. The number of spermatids produced is variable, but is constant for each species. The process of spermiohistogenesis involves many steps, including: nuclear shaping, chromatin condensation, production of an acrosome, reduction of the number of mitochondria. The present knowledge of the mechanisms of cell differentiation, and the problem of the presence of two different sperm lines in some species is discussed.     

Chromatin dynamics during spermiogenesis

The function of sperm is to safely transport the haploid paternal genome to the egg containing the maternal genome. The subsequent fertilization leads to transmission of a new unique diploid genome to the next generation. Before the sperm can set out on its adventurous journey, remarkable arrangements need to be made during the post-meiotic stages of spermatogenesis. Haploid spermatids undergo extensive morphological changes, including a striking reorganization and compaction of their chromatin. Thereby, the nucleosomal, histone-based structure is nearly completely substituted by a protamine-based structure. This replacement is likely facilitated by incorporation of histone variants, post-translational histone modifications, chromatin-remodeling complexes, as well as transient DNA strand breaks. The consequences of mutations have revealed that a protamine-based chromatin is essential for fertility in mice but not in Drosophila. Nevertheless, loss of protamines in Drosophila increases the sensitivity to X-rays and thus supports the hypothesis that protamines are necessary to protect the paternal genome. Pharmaceutical approaches have provided the first mechanistic insights and have shown that hyperacetylation of histones just before their displacement is vital for progress in chromatin reorganization but is clearly not the sole inducer. In this review, we highlight the current knowledge on post-meiotic chromatin reorganization and reveal for the first time intriguing parallels in this process in Drosophila and mammals. We conclude with a model that illustrates the possible mechanisms that lead from a histone-based chromatin to a mainly protamine-based structure during spermatid differentiation. This article is part of a Special Issue entitled: Chromatin and epigenetic regulation of animal development.     

The role of histones in chromatin remodelling during mammalian spermiogenesis.

One of the most dramatic chromatin remodelling processes takes place during mammalian spermatogenesis. Indeed, during the postmeiotic maturation of male haploid germ cells, or spermiogenesis, histones are replaced by small basic proteins, which in mammals are transition proteins and protamines. However, nothing is known of the mechanisms controlling the process of histone replacement. Two hints from the literature could help to shed light on the underlying molecular events: one is the massive synthesis of histone variants, including testis-specific members, and the second is a stage specific post-translational modification of histones. A new testis-specific 'histone code' can therefore be generated combining both histone variants and histone post-translational modifications. This review will detail these two phenomena and discuss possible functional significance of the global chromatin alterations occurring prior to histone replacement during spermiogenesis.     

Membranous tubes in pseudoscorpion spermiogenesis

The highly complicated differentiation of the spermatid in the pseudoscorpion Diplotemnus sp. is accomplished without the presence of microtubules. Instead membranous tubes measuring approximately 50 nm in diameter and closely associated with endoplasmic reticulum are found from early to mid spermatids. The lumen of the tube is devoid of electron dense contents but a fluffy material is attached to the cytoplasmic side. They run straight or slightly bent and are in open connection with the cell membrane. First appearing near the cell bridge of the interconnected spermatids they form a bundle in the longitudinal axis during a transitory phase of elongation. When the cell rounds off again the tubules together with the endoplasmic reticulum disappear. The arrangement of the tubes and their presence during abortive elongation of the spermatid suggest a supportive function commonly attributed to microtubules. Moreover, the open connection with the cell membrane and their close association with the endoplasmic reticulum may indicate their participation also in transport.     

Membrane flow during nematode spermiogenesis

Two distinct types of surface membrane rearrangement occur during the differentiation of Caenorhabditis elegans spermatids into amoeboid spermatozoa. The first, detected by the behavior of latex beads attached to the surface, is a nondirected, intermittent movement of discrete portions of the membrane. This movement starts when spermatids are stimulated to differentiate and stops when a pseudopod is formed. The second type of movement is a directed, continual flow of membrane components from the tip of the pseudopod to its base. Both membrane glycoproteins and fluorescent phospholipids inserted in the membrane flow backward at the same rate, approximately 4 micrometers/min, although their lateral diffusion coefficients in the membrane differ by at least a factor of 5. These observations suggest that pseudopodial membrane movement is due to bulk flow of membrane components away from the tip of the pseudopod.     

Dynamics of spermiogenesis in Drosophila melanogaster

Summary A morphogenetic process that transforms spermatids from a syncytial state to a state in which each spermatid is invested in its own membrane, is initiated at the head region of the spermatid bundle and traverses through the entire length of the bundle in the testis of Drosophila melanogaster. This process not only eliminates the syncytial bridges between spermatids but also removes unneeded organelles and the excess parts of the nuclear membrane, nucleoplasm and cytoplasm. It also brings about structural modifications to flagellar elements. The propagation of this process is seen as the caudal movement of a fusiform swelling of the spermatid bundle, 100 or more in length. Spermatids are individualized in the basal half of the swelling, whereas they remain syncytial in the apical half. The swelling increases its volume as it accumulates cytoplasmic debris while traversing the sperm bundle, from about 15 in maximum diameter in the basal testicular region to as large as 30 at the apical end where it becomes a bag of wastes. A variation of the process in a mutant stock which is known to inactivate up to half of the products of meiosis is briefly described. The morphological change of interspermatid bridges prior to the individualization is also reported.This work was supported by grants from the National Institutes of Health (USPHS-HD 03015 and GM-15971) and a contract from the Atomic Energy Commission, AT(04-3)-34, P.A. 150.Graduate training grant USPHS GM 00702.     

Hormonal regulation of spermatogenesis and spermiogenesis

Normal testicular function is dependent upon hormones acting through endocrine and paracrine pathways both in vivo and in vitro. Sertoli cells provide factors necessary for the successful progression of spermatogonia into spermatozoa. Sertoli cells have receptors for follicle stimulating hormone (FSH) and testosterone which are the main hormonal regulators of spermatogenesis. Hormones such as testosterone, FSH and luteinizing hormone (LH) are known to influence the germ cell fate. Their removal induces germ cell apoptosis. Proteins of the Bcl-2 family provide one signaling pathway which appears to be essential for male germ cell homeostasis. In addition to paracrine signals, germ cells also depend upon signals derived from Sertoli by direct membrane contact. Somatostatin is a regulatory peptide playing a role in the regulation of the proliferation of the male gametes. Activin A, follistatin and FSH play a role in germ cell maturation during the period when gonocytes resume mitosis to form the spermatogonial stem cells and differentiating germ cell populations. In vitro cultures systems have provided evidence that spermatogonia in advance stage of differentiation have specific regulatory mechanisms that control their fate. This review article provides an overview of the literature concerning the hormonal pathways regulating spermatogenesis.     

Ethanol-phosphotungstic acid and bismuth staining of spermatid nucleoli in mouse spermiogenesis.

The nucleoli of developing mouse spermatids were examined with ethanol-phosphotungstic acid (E-PTA) staining, and also with bismuth staining following formaldehyde fixation (FA-Bi staining) and glutaraldehyde fixation (GA-Bi staining). Only the cortical zone of the nucleolar dense fibrillar component (DFC) in the round spermatids was stained with E-PTA, while the inner area remained either faintly (early Golgi-phase spermatids) or completely unstained (cap-phase spermatids). Incubation of the fixed testis with dithiothreitol before E-PTA staining resulted in homogeneously intense staining of the DFC. The facts suggest that numerous E-PTA-positive basic proteins were present in the DFC, but disulfide crosslinks formed in the DFC proteins prevent penetration of PTA into the DFC interior. The DFC was stained with bismuth after FA-Bi and GA-Bi staining until the disappearance of the nucleoli occurring in acrosome-phase spermatids. The fibrillar center, homogeneously stained using E-PTA, FA-Bi, and GA-Bi methods was present in the nucleoli of Golgi-phase and early cap-phase spermatids, but disappeared in the nucleoli of late cap-phase spermatids. These results are discussed based on the previous studies dealing with the ribosomal RNA synthesis in mouse spermiogenesis.