Cytoplasm Of Spermatids Research Articles

High-Sensitivity and High-Resolution In Situ Hybridization of Coding and Long Non-coding RNAs in Vertebrate Ovaries and Testes

BackgroundSubcellular localization of coding and non-coding RNAs has emerged as major regulatory mechanisms of gene expression in various cell types and many organisms. However, techniques that enable detection of the subcellular distribution of these RNAs with high sensitivity and high resolution remain limited, particularly in vertebrate adult tissues and organs. In this study, we examined the expression and localization of mRNAs encoding Pou5f1/Oct4, Mos, Cyclin B1 and Deleted in Azoospermia-like (Dazl) in zebrafish and mouse ovaries by combining tyramide signal amplification (TSA)-based in situ hybridization with paraffin sections which can preserve cell morphology of tissues and organs at subcellular levels. In addition, the distribution of a long non-coding RNA (lncRNA), lncRNA-HSVIII, in mouse testes was examined by the same method.ResultsThe mRNAs encoding Mos, Cyclin B1 and Dazl were found to assemble into distinct granules that were distributed in different subcellular regions of zebrafish and mouse oocytes, suggesting conserved and specific regulations of these mRNAs. The lncRNA-HSVIII was first detected in the nucleus of spermatocytes at prophase I of the meiotic cell cycle and was then found in the cytoplasm of round spermatids, revealing expression patterns of lncRNA during germ cell development. Collectively, the in situ hybridization method demonstrated in this study achieved the detection and comparison of precise distribution patterns of coding and non-coding RNAs at subcellular levels in single cells of adult tissues and organs.ConclusionsThis high-sensitivity and high-resolution in situ hybridization is applicable to many vertebrate species and to various tissues and organs and will be useful for studies on the subcellular regulation of gene expression at the level of RNA localization.

Endocrine regulation of sperm release.

Spermiation (sperm release) is the culmination of a spermatid's journey in the seminiferous epithelium. After a long association with the Sertoli cell, spermatids have to finally 'let go' of the support from Sertoli cells in order to be transported to the epididymis. Spermiation is a multistep process characterised by removal of excess spermatid cytoplasm, recycling of junctional adhesion molecules by endocytosis, extensive cytoskeletal remodelling and final spermatid disengagement. Successful execution of all these events requires coordinated regulation by endocrine and paracrine factors. This review focuses on the endocrine regulation of spermiation. With the aim of delineating how hormones control the various aspects of spermiation, this review provides an analysis of recent advances in research on the hormonal control of molecules associated with the spermiation machinery. Because spermiation is one of the most sensitive phases of spermatogenesis to variations in hormone levels, understanding their molecular control is imperative to advance our knowledge of the nuances of spermatogenesis and male fertility.

Testosterone regulates granzyme K expression in rat testes.

Testosterone depletion induces increased germ cell apoptosis in testes. However, limited studies exist on genes that regulate the germ cell apoptosis. Granzymes (GZM) are serine proteases that induce apoptosis in various tissues. Multiple granzymes, including GZMA, GZMB and GZMN, are present in testes. Th us, we investigated which granzyme may be testosterone responsive and possibly may have a role in germ cell apoptosis aft er testosterone depletion. Ethylene dimethane sulfonate (EDS), a toxicant that selectively ablates the Leydig cells, was injected into rats to withdraw the testosterone. The testosterone depletion effects after 7 days post-EDS were verified by replacing the testosterone exogenously into EDS-treated rats. Serum or testicular testosterone was measured by radioimmunoassay. Using qPCR, mRNAs of granzyme variants in testes were quantified. The germ cell apoptosis was identified by TUNEL assay and the localization of GZMK was by immunohistochemistry. EDS treatment eliminated the Leydig cells and depleted serum and testicular testosterone. At 7 days post-EDS, testis weights were reduced 18% with increased germ cell apoptosis plus elevation GZMK expression. GZMK was not associated with TUNEL-positive cells, but was localized to stripped cytoplasm of spermatids. In addition, apoptotic round spermatids were observed in the caput epididymis. GZMK expression in testes is testosterone dependent. GZMK is located adjacent to germ cells in seminiferous tubules and the presence of apoptotic round spermatids in the epididymis suggest its role in the degradation of microtubules in ectoplasmic specializations. Thus, overexpression of GZMK may indirectly regulate germ cell apoptosis by premature release of round spermatids from seminiferous tubule lumen.

The expression characteristics of FAM71D and its association with sperm motility.

What are the features of FAM71D (Family with sequence similarity 71, member D) expression and is there an association between FAM71D expression and sperm motility? FAM71D, a novel protein exclusively expressed in the testis, is located in sperm flagella and is functionally involved in sperm motility. Some testis-specific proteins have been reported as potential diagnostic biomarkers to evaluate the spermatogenesis process and sperm quality. We have identified a novel testis-specific protein, FAM71D, through microarray data analysis, yet little is known about its expression and function. FAM71D mRNA and protein expression was quantified during mouse testis development. Its localization in germ cells was detected by dual-labeled immunostaining in testis sections and sperm smears. The clinical significance was assessed by comparing FAM71D expression in spermatozoa from normozoospermic controls and asthenozoospermic patients. Testes were dissected from C57BL/6 J male mice at postnatal ages of 1, 2, 3, 4, 6, 8 weeks and 6 months, and sperm was collected from cauda epididymides of adult mice by the swim-up method. Human spermatozoa were isolated from 100 human semen samples by density gradient Percoll centrifugation. RT-qPCR and western blot were performed to semi-quantify the expression of FAM71D in mouse testis, and in the ejaculated spermatozoa of normozoospermic controls and asthenozoospermic patients. Immunofluorescence staining was used to detect the localization of FAM71D. Co-immunoprecipitation assay was performed to evaluate the interaction between FAM71D and calmodulin. An antibody blocking assay was employed to assess the role of FAM71D in sperm motility. Our results showed that FAM71D was exclusively expressed in the testis in an age-dependent manner. FAM71D expression exhibited dynamic change in the cytoplasm of spermatids during spermiogenesis and was finally retained in sperm flagella. FAM71D could interact with calmodulin. Use of anti-FAM71D antibody on sperm significantly decreased sperm motility. Expression level of FAM71D was markedly reduced in the ejaculated spermataozoa of asthenozoospermic patients (P < 0.05), and this was correlated with sperm progressive motility (r = 0.7435, P < 0.0001). N/A. The sample size was limited and it is necessary to verify the correlation of FAM71D expression with sperm motility in larger cohorts. Furthermore, our results were descriptive and follow-up studies would be needed to elucidate the detailed role of FAM71D in sperm motility. This is the first systematic study to document the expression of endogenous FAM71D and a function for FAM71D in sperm motility. It provides new insights into our understanding of sperm motility regulation and causes of male infertility. This study was funded by the National Natural Science Foundation of China, Guangdong Natural Science Foundation and the Shenzhen Project of Science and Technology. The authors have no competing interests.

KH-type splicing regulatory protein is a new component of chromatoid body.

The chromatoid body (CB) is a specific cloud-like structure in the cytoplasm of haploid spermatids. Recent findings indicate that CB is identified as a male germ cell-specific RNA storage and processing center, but its function has remained elusive for decades. In somatic cells, KH-type splicing regulatory protein (KSRP) is involved in regulating gene expression and maturation of select microRNAs (miRNAs). However, the function of KSRP in spermatogenesis remains unclear. In this study, we showed that KSRP partly localizes in CB, as a component of CB. KSRP interacts with proteins (mouse VASA homolog (MVH), polyadenylate-binding protein 1 (PABP1) and polyadenylate-binding protein 2 (PABP2)), mRNAs (Tnp2 and Odf1) and microRNAs (microRNA-182) in mouse CB. Moreover, KSRP may regulate the integrity of CB via DDX5-miRNA-182 pathway. In addition, we found abnormal expressions of CB component in testes of Ksrp-knockout mice and of patients with hypospermatogenesis. Thus, our results provide mechanistic insight into the role of KSRP in spermatogenesis.

Ubiquitination-Deficient Mutations in Human Piwi Cause Male Infertility by Impairing Histone-to-Protamine Exchange during Spermiogenesis

Genetic studies have elucidated critical roles ofPiwiproteins in germline development in animals,but whether Piwi is an actual disease gene in human infertility remains unknown. We report germline mutations in human Piwi (Hiwi) in patients with azoospermia that prevent its ubiquitination and degradation. By modeling such mutationsin Piwi (Miwi) knockin mice, we demonstratethat the genetic defects are directly responsiblefor male infertility. Mechanistically, we show that MIWI binds the histone ubiquitin ligase RNF8 in a Piwi-interacting RNA(piRNA)-independentmanner, and MIWI stabilization sequesters RNF8 in the cytoplasm of late spermatids. The resulting aberrant sperm show histone retention, abnormal morphology, and severely compromised activity, which can be functionallyrescued via blocking RNF8-MIWI interactioninspermatids with an RNF8-N peptide. Collectively, our findings identify Piwi as a factorin human infertilityand reveal its role in regulating the histone-to-protamine exchange during spermiogenesis.

Expression pattern of HSFY in the mouse testis and epididymis with and without heat stress.

Heat shock factors (HSFs) are critical regulators of spermatogenesis. However, heat shock responses, the associated components and the underlying functional mechanisms remain to be elucidated. Here, we characterize the expression pattern of HSFY, a member of the HSF family in the testis and epididymis. Its expression in testis and epididymis was initially identified by western blots. Immunofluorescence staining demonstrated that HSFY was confined to the cytoplasm of late spermatocytes and spermatids in adult testes, gonocytes in newborn testes and undifferentiated spermatogonia in 7days post-parturition testes. In the epididymis, HSFY was predominantly expressed in principal cells. Furthermore, a single transient scrotal heat stress did not change HSFY protein expression in the testes or epididymis, either on the expressional level or in cellular localization. In summary, this study detected the expression pattern of HSFY in the testes and epididymis and demonstrated that its expression was not regulated by transient elevated temperature.

Sperm head shaping in ratites: New insights, yet more questions.

Head shaping in mammalian sperm is regulated by a number of factors including acrosome formation, nuclear condensation and the action of the microtubular manchette. A role has also been suggested for the attendant Sertoli cells and the perinuclear theca (PT). In comparison, relatively little information is available on this topic in birds and the presence of a PT per se has not been described in this vertebrate order. This study revealed that a similar combination of factors contributed to head shaping in the ostrich, emu and rhea, although the Sertoli cells seem to play a limited role in ratites. A fibro-granular structure analogous to the mammalian PT was identified, consisting of sub- and post-acrosomal components. The latter was characterized by stage-specific finger-like projections that appeared to emanate from the cytoplasmic face of the nuclear envelope. They were particularly obvious beneath the base of the acrosome, and closely aligned, but not connected to, the manchette microtubules. During the final stages of chromatin condensation and elongation of the sperm head the projections abruptly disappeared. They appear to play a role in stabilizing the shape of the sperm head during the caudal translocation of the spermatid cytoplasm.

KIF1-binding protein interacts with KIF3A in haploid male germ cells.

Male fertility relies on the production of functional spermatozoa. Spermatogenesis is a complex differentiation process that is characterized by meiosis and dramatic morphogenesis of haploid cells. Spermatogenesis involves active changes in the microtubular network to support meiotic divisions, cell polarization, the reshaping of the nucleus, and the formation of a flagellum. Previously, we have demonstrated that a microtubule-based anterograde transport motor protein KIF3A is required for the sperm tail formation and nuclear shaping during spermatogenesis. In this study, we show that KIF3A interacts with a KIF1-binding protein (KBP) in the mouse testis. We have characterized the expression and localization pattern of KBP during spermatogenesis and localized both KIF3A and KBP in the cytoplasm of round spermatids and manchette of elongating spermatids. Interestingly, KBP localized also in the late chromatoid body (CB) of elongating spermatids, whose function involves intracellular movement and association with the microtubular network. Altogether our results suggest a role for KBP in spermatid elongation and in the function of the late CB.

Testis follicles ultrastructure of three species of terrestrial isopods (Crustacea, Isopoda Oniscidea).

The aim of the research, carried out on three species of terrestrial isopods - Armadillidium granulatum, Halophiloscia hirsuta and Trichoniscus alexandrae - is to bring a first consistent contribution to the knowledge of the ultrastructural organization of the testis follicles. The testis follicles are seat of a remarkable dynamic activity of their cell components (somatic cells and germ cells) that results in a continuous variation, related to the trend of spermatogenesis, of their morphology, organization and of the relationships between the two cell populations. The somatic cells, known in literature as follicular cells, nurse cells or Sertoli cells, are arranged at the periphery of the follicle to form an epithelial layer of variable thickness resting on a thin basal lamina in turn surrounded by a discontinuous network of muscle cells. In A. granulatum and H. hirsuta, two types of Sertoli cells are present: a first type, the nurse cells, envelop the spermatids in cavities within their cytoplasm and through their secretion activity play a fundamental role in the formation of the spermatophores; moreover, they phagocytizes the residual cytoplasm of spermatids. A second type of Sertoli cells shows features that leave clearly identify its supporting role to the spermatophores in formation. In T. alexandrae, instead, only one type of Sertoli cells, the nurse cell, is present, whose features are widely superimposable to those observed in the other two species. Moreover, two septa of Sertoli cells depart from the periphery of the testis follicle to constitute an articulated compartmentalization of the follicle itself, probably targeted to realize at its inside a series of microenvironments functionally diversified in order to meets the needs of the different stages of the spermatogenic cycle.

Evolution of cadmium effects in the testis and sperm of the tropical fish Gymnotus carapo.

The present study investigated the testis and sperm morphology of the tropical fish Gymnotus carapo after exposure to increasing CdCl2 concentrations (5-40 μM) for 24 and 96 h. The treatments induced Cd accumulation in the testis and a decrease in the gonadosomatic index from a 10 μM. Cd induced alterations in testis since 24h; however the extension and severity of damages increased after 96 h in all tested concentrations. Marked variations in the cysts size, proliferation of the interstitial tissue, infiltration of inflammatory cells, necrosis, reduction of germ cells and sperm aggregation was observed in 96 h treated fishes. In this time, there was a complete absence of germ cells in the testis of fish treated with 40 μM. The ultrastructural analysis allowed for the visualization of the initial damages over germ cells, such as the presence of vacuoles in the cytoplasm of spermatogonia, spermatocytes, and spermatids. Exposed fish (20 μM for 24 and 96 h) had alterations in sperm number and morphology. These results are important for establishing a direct correlation between the Cd accumulation and incidence of damages and can help characterize the mechanism of Cd-induced pathogenesis in the male reproductive system.

DNAJB13, a type II HSP40 family member, localizes to the spermatids and spermatozoa during mouse spermatogenesis.

BackgroundHundreds of HSP40s derived from various species have been identified, of which several proteins are involved in spermatogenesis. DNAJB13 is a type II HSP40/DnaJ protein. In a previous study, we cloned mouse Dnajb13, which is up-regulated in cryptorchidism. To date, however, little is known about the localization and functions of DNAJB13 during spermatogenesis. This study intends to identify the expression pattern of DNAJB13 during mammalian spermatogenesis.ResultsRT-PCR and western blot revealed that the Dnajb13 gene and DNAJB13 protein were expressed not only in the testis but also in several other ciliated cell-containing tissues like the trachea, lung and oviduct. Quantitative PCR showed that the expression of Dnajb13 mRNA in testis was detectable as early as postnatal week 1, and sharply increased from postnatal week 3. Western blotting and immunohistochemistry determined that the DNAJB13 protein, which was located in the cytoplasm of spermatids and the sperm flagellum, was detectable from postnatal week 4.ConclusionsBased on the spatiotemporal expression observed in the cytoplasm of spermatids and sperm flagella, we suggest that DNAJB13 participates in spermiogenesis and the motility of mature spermatozoa.

CEACAM2-L on spermatids interacts with poliovirus receptor on Sertoli cells in mouse seminiferous epithelium.

The removal of excess cytoplasm from elongated spermatids by Sertoli cells is the last essential step in spermatogenesis. It requires cell-to-cell recognition between a Sertoli cell and an elongating spermatid through protein-protein interactions. CEACAM2-L, an adhesion molecule of the immunoglobulin superfamily (IgSF), is present at the plasma membrane covering the excess cytoplasm of elongated spermatids, and is possibly involved in the cell-to-cell recognition. In this study, we investigated the interaction between CEACAM2-L and Poliovirus receptor (PVR), which is also from the IgSF and is expressed by Sertoli cells. Immunohistochemical analysis showed that CEACAM2-L expressed on elongated spermatids was in close contact with PVR-positive cell processes of Sertoli cells. Immunoprecipitation experiments both in vivo and in vitro demonstrated a direct heterophilic interaction between CEACAM2-L and PVR. We show that the N-terminal Ig domain of CEACAM2-L was critical for its interaction with PVR. In addition, we found that CEACAM2-L formed heterophilic trans-tetramers with PVR in transfected COS-7 cells. From these data, we propose that Sertoli cells recognize the excess cytoplasm of elongated spermatids through the PVR-CEACAM2-L interaction in mouse testis.

The expression pattern of PARP-1 and PARP-2 in the developing and adult mouse testis

Although the importance of the PARP family members in the adult testis has already been acknowledged, their expression in the developing testis has not been addressed. We performed immunohistochemistry by using PARP-1 and PARP-2 antibodies on the developing mouse testis at embryonic day (E) 15.5, E17.5, postnatal day (PN) 0, PN3, PN9, PN20 and adult. Our results showed that at embryonic and early postnatal days, the expression of PARP-1 was in the nuclei of gonocytes and spermatogonia. PARP-1 was positive in interstitial cells with nuclear localization at all studied ages. At embryonic and early postnatal days, the expression of PARP-2 was in the cytoplasm of gonocytes and spermatogonia. During the progress of spermatogenesis, PARP-2 was localized in the cytoplasm of pre-leptotene spermatocytes on PN9, in the cytoplasm of pachytene spermatocytes on PN15 and in the cytoplasm of round spermatids on PN20. In the adult, PARP-2 staining can still be observed in the cytoplasm of spermatogonia, but to a much lesser degree than in the round and elongating spermatids. For all the studied ages, PARP-2 was positive in Sertoli cells and interstitial cells with cytoplasmic localization. Our results indicate that PARP proteins are present in germ and somatic cells during testis development in mice.

Differential expression of aromatase, estrogen receptor alpha and 17β-HSD associated with the processes of total testicular regression and recrudescence in the bat Myotis nigricans (Chiroptera: Vespertilionidae)

Despite the worldwide distribution and many unique reproductive adaptations that bats present, many aspects of their reproductive hormonal regulation have not been adequately studied, especially in species that presented patterns of total testicular regression. Thus, this study aimed to evaluate the testicular expression of 17β-HSD type 1, aromatase and ERα in the bat Myotis nigricans, during the four periods of its reproductive cycle. Immunoreactivity for ERα was detected only in the cytoplasm of elongated spermatids and in the nuclei of spermatogonia and Sertoli cells. Expression of aromatase was observed in round and elongated spermatids and in Sertoli and Leydig cells. Immunoreactivity for 17β-HSD was restricted to the cytoplasm of Leydig cells. The three expression patterns varied significantly during the four periods of the reproductive cycle. Expression of ERα and aromatase in spermatids was continuous, while expression of ERα in spermatogonia occurred only in initial types (Ap). Expression of ERα and aromatase in Sertoli cells varied, with expression only in periods of spermatogenetic activities; and the same variation was observed for the expression of aromatase and 17β-HSD in Leydig cells. We, therefore, propose that the processes of total testicular regression and posterior recrudescence suffered by M. nigricans from September to January in the northwest of the São Paulo State of Brazil, are directly regulated by testosterone and estrogen. This occurs via the production of testosterone by 17β-HSD, its conversion into estrogen by aromatase, and activation/deactivation of Sertoli cells’ AR and spermatogonia’s ERα.

Expression and identification of a novel gene Spata34 in mouse spermatogenic cells

Leucine-rich repeat (LRR) containing proteins play an essential role in signal transduction, cell adhesion, cell development, DNA repair and RNA processing. Here we cloned a novel gene, Spata34, encoding a LRR containing protein of 415 aa. Spata34 gene consisted of 9 exons and 8 introns and mapped to chromosome 3qA3. Spata34 is conserved across species in evolution. The Spata34 gene was expressed at various levels, faintly before first weeks postpartum and strongly from 2weeks postpartum in adult testes. Western blot analysis showed that Spata34 protein was specially expressed in mouse testis. Immunohistochemical analysis revealed that Spata34 protein was most abundant in the cytoplasm of round spermatids and elongating spermatids within seminiferous tubules of the adult testis. Overexpression of Spata34 in COS7 cells inhibited the transcriptional activity of AP-1, p53 and p21 which suggested that Spata34 protein may act as a transcriptional repressor in p53 and p21 pathway.

Localization of proteins involved in endocytosis at tubulobulbar complexes in rat testes

Tubulobulbar Complexes (TBCs) are actin-rich structures formed between Sertoli-cells and spermatids at the time of sperm release. The main functions of the TBCs are to remove excess spermatid cytoplasm and acrosomal contents, internalize and recycle junctional complexes by endocytosis prior to spermiation. However, in addition to recycling some of the molecules undergo lysosomal degradation. The molecular machinery involved in endocytosis at the TBCs is not well understood. To bridge this gap localization of various proteins, involved at various steps of endocytosis studied in other systems, was demonstrated in TBCs using testicular fragmented material or sections by immunoblotting and immunofluroscence. The presence of key molecules like Vamp-2, syntaxin and Lamp-2 indicates occurrence of lysosomal degradation in addition to junctional recycling at the TBCs present at the time of sperm release. TBCs are endocytic devices functioning to recycle junctional molecules or remove spermatid cytoplasm that were present between spermatids and Sertoli-cells all through the process of spermatid maturation and in turn regulate male fertility.

The cytoplasmic droplet may be indicative of sperm motility and normal spermiogenesis

Although the cytoplasm of spermatids is removed at the end of spermiogenesis, a tiny portion is usually retained in the sperm flagellum, which is termed the cytoplasmic droplet (CD) in mammals. CDs are believed to play a role in sperm volume adaptation. However, we have noticed that epididymal spermatozoa that display initial (flagellation in situ) and progressive motility mostly possess CDs, whereas spermatozoa without CDs are rarely motile, suggesting that CDs have a role in motility development during sperm epididymal maturation. In the present study, we analyzed the relationship between the presence or absence of CDs, motility development and positional changes of CDs during sperm epididymal maturation in mice and monkeys. We also examined CDs on spermatozoa of three knockout mouse lines with late spermiogenic defects. Our data suggest that the CD is a normal organelle transiently present exclusively on epididymal spermatozoa, and normal CD morphology and location are associated with normal motility development during epididymal maturation of spermatozoa. Abnormal CD formation, e.g., a complete lack of CDs or ectopic CDs, is indicative of defective spermiogenesis. If CDs are essential for sperm motility development, then CDs may represent an ideal drug target for the development of non-hormonal male contraceptives.

Three Testis-Specific Paralogous Serine Proteases Play Different Roles in Murine Spermatogenesis and Are Involved in Germ Cell Survival During Meiosis1

Spermatogenesis is a complex process that generates spermatozoa; its molecular mechanisms are not completely understood. Here we focused on the functions of three testis-specific serine proteases: Prss42/Tessp-2, Prss43/Tessp-3, and Prss44/Tessp-4. These protease genes, which constitute a gene cluster on chromosome 9F2-F3, were presumed to be paralogs and were expressed only in the testis. By investigating their mRNA distribution, we found that all three genes were expressed in primary and secondary spermatocytes. However, interestingly, the translated proteins were produced at different locations. Prss42/Tessp-2 was found in the membranes and cytoplasm of secondary spermatocytes and spermatids, whereas Prss43/Tessp-3 was present only in the membranes of spermatocytes and spermatids. Prss44/Tessp-4 was detected in the cytoplasm of spermatocytes and spermatids. To assess the roles of these proteases in spermatogenesis, we used organ culture of mouse testis fragments. Adding antibodies against Prss42/Tessp-2 and Prss43/Tessp-3 resulted in meiotic arrest at the stage when each protease was beginning to be translated. Furthermore, the number of apoptotic cells dramatically increased after the addition of these antibodies. These results strongly suggest that the three paralogous Prss/Tessp proteases play different roles in spermatogenesis and that Prss42/Tessp-2 and Prss43/Tessp-3 are required for germ cell survival during meiosis.

Polyubiquitination and Proteasome Signals in Tubulobulbar Complexes of Rat Late Spermatids

To illustrate the involvement of tubulobulbar complexes (TBC) in ubiquitin-proteasome degradation of unnecessary proteins in the head cytoplasm of late spermatids, the localization of polyubiquitin and proteasome was studied by immunofluorescence and immunoelectron microscopy. Polyubiquitin localized to TBC and proteasome subunit α to dense materials surrounding the TBC in the cytoplasm of Sertoli cell enwrapping sickle-shaped spermatid heads. The results suggest that the TBC is a structural device for ubiquin-proteasome degradation of unnecessary proteins in the cytoplasm of spermatid head during rapid reduction of the head cytoplasm and nuclear compaction of late spermatids.