| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Testis |
Department of Science for Laboratory Animal Experimentation, Research Institute for Microbial Diseases, Osaka University, Osaka 565-0871, Japan
 |
ABSTRACT |
|---|
 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
sperm, sperm motility and transport, spermatid, spermatogenesis, testis
|
INTRODUCTION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
The cytoskeletal element actin is presumably an important player in spermiogenesis [2–6], and actin-linked molecules [7–13] are important in shaping the sperm form, as has been noted in other systems [14]. In the mammalian testis, cytoplasmic actins are expressed at all stages of germ cell differentiation. In the mouse testis, the actin transcripts are 2.1 and 1.5 kilobases (kb). The 2.1-kb actin mRNA codes for the cytoplasmic ß- and 
-actins and is found throughout spermatogenesis, whereas the 1.5-kb actin mRNA is detected initially in haploid germ cells, where it codes for the testicular smooth muscle -actin, and is controlled developmentally by an element in the 3'-untranslated region [6]. The existence of developmental regulation of actin expression indicates that actin is involved in the differentiation of haploid germ cells.
Experiments using polyclonal and monoclonal antibodies have shown that actin (F-actin) is concentrated mainly in the subacrosomal space, which lies adjacent to the postacrosomal region, and in the intercellular bridges [1, 4, 9] and plays an important role in sperm function [15–17]. However, during spermiogenesis, the actins are localized in haploid germ cells in a species-specific fashion, which makes it difficult to elucidate their roles [2, 3]. Recently, actin-related proteins [18, 19] and actin-capping proteins [7, 11] have been implicated in sperm morphogenesis and sperm function [9, 17, 19]. These observations do not necessarily confirm a fundamental role for actin in cytoskeletal formation in spermatozoa, but they do point to the importance of actins and actin-related proteins in spermiogenesis.
In this study, we identified and characterized two novel cDNAs that encode the testicular germ cell-specific actins T-ACTIN 1 and T-ACTIN 2. These two unique actins were expressed only in haploid germ cells in a stage-specific manner during spermiogenesis. These results suggest that T-ACTINs play an important role in sperm function and in the specific formation of spermatozoa during spermiogenesis.
|
MATERIALS AND METHODS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
Total RNA was extracted from the testes of 17-day-old and 35-day-old C57BL/6 mice using the guanidine isothiocyanate-CsTFA method [20]. The preparation of the cDNA library has been described elsewhere [21]. The cDNA fragments were inserted directionally between the NotI (dephosphorylated) and BglII sites of the pAP3neo vector (Takara, Siga, Japan), and the plasmids were electroporated into Escherichia coli MC1061A. The complexity of the 35-day-old mouse testis cDNA library was 4 x 106 colony-forming units (cfu). The subtracted cDNA library was generated by subtracting the mRNA of a 17-day-old mouse testis from the cDNA library of a 35-day-old testis [22]. The plasmid DNA of clones, which were selected randomly from the subtracted cDNA library, was screened by Northern blot analysis using testis mRNA that was extracted at different developmental stages between Days 17 and 35. We designated these testis-specific cDNA clones transcription increased in spermiogenesis (TISP) [22].
Cloning of Actin-Like Genes
The TISPs prepared from testis-specific subtracted cDNA library were examined for similarities using the GenBank, EMBL, and DDBJ databases. The clones named TISP108 and TISP129 in the library showed DNA sequence homologies with actin genes.
To obtain full-length cDNAs, the pAP3neo mouse testis cDNA library [21] was screened, and the complete cDNAs of TISP108 and TISP129 were obtained under high-stringency conditions of hybridization. The E. coli MC1061A cells that carried the adult testis library were diluted for seeding at 2 x 105 cfu on a nitrocellulose filter, which was then placed on a Luria–Bertani medium (LB) plate. After incubation at 37°C, the colonies were transferred to two replicate nylon filters and lysed by sequentially soaking at room temperature in the following solutions: 0.5 N NaOH plus 1.5 M NaCl for 5 min, 0.5 M Tris-HCl (pH 7.4) plus 1.5 M NaCl for 5 min, and 2x saline sodium citrate (SSC) for 5 min. After baking at 80°C for 2 h, the filters were washed to remove the bacterial debris. Two 32P-labeled probes were prepared using the BcaBEST Random Primer Kit (Takara) and the TISP108 and TISP112 cDNA fragments that corresponded to the actin-like mRNAs. The filters were hybridized with the probe at 65°C for 20 h in the hybridization solution (4x SSC, 10x Denhardt solution, 0.1% SDS, 100 µg/ml denatured sonicated salmon sperm DNA). The positive clones were purified, and the cDNA inserts were sequenced.
DNA Sequencing
Dideoxy-chain-termination sequencing reactions were performed with fluorescent dye-labeled primers and the Thermal Cycle Sequencing Kit (Applied Biosystems, Foster City, CA). The reaction products were analyzed using a Genetic Analyzer 3100 (Applied Biosystems). The GenBank, EMBL, DDBJ, Swiss-Prot, and PIR databases were searched for homology with the whole t-actin cDNAs or for the deduced amino acid sequences.
Northern Blot Analysis
The freshly removed organs of adult C57BL/6 mice were homogenized in RNAzol B (Tel-Test, Friendswood, TX). The germ cells and other somatic cells of the testis were prepared as described previously [23]. Tunica albuginea was removed from each testis, and seminiferous tubules were placed in 0.02 M Hepes and 0.01% collagenase and gently unraveled with forceps. The tubule suspension was left standing to precipitate tubule fragments. The supernatant containing separated cells was filtered through a nylon mesh (NBC Industries Co., Tokyo, Japan) and centrifuged at 600 x g for 10 min. The precipitant was used as a Leydig cell fraction. Remaining tubules were dispersed in PBS containing 1 mM EDTA to remove residual Leydig cells. Tubules were cut into small fragments with a blade, transferred to a conical tube, and washed by pipetting in PBS containing 1 mM EDTA. The conical tube was left standing, and the supernatant was filtrated through a nylon mesh and used as a germ cell fraction. The remaining sedimented tubules were vigorously pipetted a few times then left standing. The sedimented sample was used as a tubule fraction (containing mainly Sertoli cells).
Total RNA samples were extracted according to the manufacturer's (Tel–Test) recommendations and quantified by optical density measurements. RNA samples in 2.2 M formaldehyde were subjected to electrophoresis in a 1.1% agarose gel that contained 0.66 M formaldehyde. The RNA bands were transferred to a nitrocellulose filter in 20x SSC. Hybridization was performed by incubating the filter with a 32P-labeled cDNA probe of full-length t-actin 1 or 2, which was prepared using the BcaBEST Random Primer Kit (Takara), at 42°C for 16 h in a hybridization solution (4x SSC, 5x Denhardt, 0.2% SDS, 12 µg/ml denatured sonicated salmon sperm DNA, 50% formamide). The filters were washed twice in 0.3x SSC plus 0.1% SDS at 60°C. The signals were detected using an Image Analyzer (Fuji Film, Tokyo, Japan). After exposure, the same filters were rehybridized with the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as a control.
In situ Hybridization
Antisense digoxigenin (DIG)-labeled RNA was used for the in situ hybridization experiments. The testes were frozen in OTC embedding compound (Tissue-Tek; Sakura Finetechnical Co., Tokyo, Japan), and cryosections (7 µm thickness) were prepared on APS-coated Superfrost micro-glass slides (Matsunami Glass Ind., Osaka, Japan). The sections were dried and fixed in a solution of 4% paraformaldehyde, 0.5% glutaraldehyde, and 0.5 M sodium phosphate buffer (pH 7.4). The 357-base pair (bp) fragment that encompasses nucleotides (nt) 514–870 of t-actin 1 and the 176-bp fragment (nt 246–421) of t-actin 2 were cloned into the multiple cloning site of pBluescript II SK+ and used for in situ hybridization. The t-actin 1 and 2 constructs in pBluescript II SK+ (Stratagene, LaJolla, CA) were digested with the appropriate restriction enzymes and used as templates for T7 or T3 RNA polymerase, thereby generating the sense and antisense probes. The probes were labeled with DIG-labeled UTP (Boehringer Mannheim, Mannheim, Germany). In situ hybridization was performed as described previously [24, 25]. After hybridization, the bound probe was detected by incubating the slides with anti-DIG-Fab fragments that were conjugated with alkaline phosphatase (Boehringer Mannheim), followed by color development with 4-nitroblue tetrazolium chloride (NBT; Boehringer Mannheim) and 5-bromo-4-chloro-3-indolyl-phosphate (BCIP; Boehringer Mannheim). The sections were contrasted with 1% methyl green stain solution (Muto Pure Chemicals, Osaka, Japan) and examined under a microscope.
Antiserum Preparation
Two synthetic peptides, AC1 (MATKNSPSPKPMGT, residues 1–15 of T-ACTIN 1) and AC2 (DGPAKKASDQASMQT, residues 16–30 of T-ACTIN 2), which were designed from the deduced amino acid sequences of the novel actins, were purified, and each synthetic peptide was used to immunize two Japanese white rabbits (KAC, Kyoto, Japan). Polyclonal antisera were obtained by injection of each of the antigens, followed by three booster injections at 3-wk intervals, for a total of four injections. Each antiserum (AC1, AC2) reacted specifically with its antigen on Western blots. The most effective antiserum was used for each experiment.
Western Blot Analysis
Freshly prepared organs or cell fractions of adult C57BL/6 mice were homogenized on ice in a lysis buffer that contained 10 mM Tris-HCl (pH 7.6), 150 mM NaCl, 0.1% SDS, 0.1% sodium deoxycholate, 1% NP-40, and 0.5 ml/L protease inhibitor cocktail (Sigma, St. Louis, MO). The germ cells prepeared from adult mice testes were incubated with lysis buffer (10 mM Tris-HCl, pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.5% NP-40) on ice for 5 min. After centrifugation (9000 x g), supernatant and precipitant were used as cytoplasmic and nuclear fractions, respectively [23]. After centrifugation (4°C for 10 min, 15 000 x g) the protein concentration of each supernatant was estimated by the Bradford protein assay (Nacalai Tesque, Kyoto, Japan).
The extracts from the sperm, nuclear, and cytoplasmic fractions each contained between 30 and 50 µg protein, samples of which were subjected to SDS-PAGE following the procedure of Laemmli [26]. The protein bands were electroblotted onto polyvinylidene difluoride membrane filters (Millipore, Bedford, MA), and the filters were blocked with 5% nonfat milk and washed for 15 min with Tris-buffered saline (TBS; 50 mM Tris-HCl, pH 7.5, 150 mM NaCl) containing 0.05% Tween-20 (TBS-T). The filters were then incubated with the anti-T-ACTIN 1 rabbit antiserum (AC-1; 1:1500 dilution), rabbit serum before immunization of synthetic peptide for T-ACTIN 1 (1:1500 dilution), the anti-T-ACTIN 2 rabbit antiserum (AC-2; 1:1000 dilution), or rabbit serum before immunization of synthetic peptide for T-ACTIN 2 (1:1000 dilution) in TBS at 4°C overnight. The filters were then washed in TBS-T for 5 min, rewashed three times, and incubated at 25°C for 1 h with anti-rabbit Igs conjugated to horseradish peroxidase (1:1000 dilution) (Amersham Pharmacia, Piscataway, NJ). After further washing, the reactive bands were visualized by development with the POD Immunostain Kit (Wako, Osaka, Japan).
Immunohistochemistry
Frozen sections (10 µm thickness) of the testes were dried onto micro-glass slides (Matsunami). Sperm from the epididymis of adult mouse were suspended in PBS. After 1 h, a few drops of sperm were spotted onto micro-glass slides. The sections were fixed with 100% methanol for 20 min at room temperature, incubated with the blocking solution (Nacalai Tesque), and then reacted with the polyclonal anti-T-ACTIN 1 antiserum (1:1500 dilution), the anti-T-ACTIN 2 antiserum (1:1000 dilution), or each preimmune rabbit serum (1:1000 dilution) for controls overnight at 4°C. The sections were then incubated with a 1:750 dilution of the fluorescein isothiocyanate (FITC)-labeled anti-rabbit Ig antibody (Amersham Pharmacia) for 2 h at room temperature. The sections were examined under a fluorescence microscope.
|
RESULTS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
To study the mechanism of male germ cell differentiation, especially at the haploid stage, we constructed a cDNA library that was enriched for cDNAs that were expressed specifically during spermiogenesis. We subtracting cDNA clones that were prepared from 17-day-old premeiotic testes from the cDNA clones of 35-day-old adult testes [22, 27], and using the subtracted cDNA library, we isolated several cDNA clones that were expressed specifically in haploid germ cells. Two of these clones, TISP108 and TISP129 [22], encoded proteins that had homology to actins. We used these cDNA fragments as templates to screen 2 x 105 clones of a pAP3neo mouse testis cDNA library [21] and found approximately 30 positive signals. We then selected five clones each for TISP108 and TISP129 and performed DNA sequencing. The five clones that were derived from each cDNA probe had the same sequence and encoded a single open reading frame (ORF). The complete nucleotide sequences and deduced amino acid (aa) sequences are shown in Figure 1. Each of the cDNA clones showed 55% identity with the previously reported cDNA of mouse actin. We designated these cDNAs as testis-specific actin (t-actin) 1 and t-actin 2. The ORFs of t-actin 1 and t-actin 2 consisted of 418 and 440 amino acids, starting at the first methionine at nt 28 or nt 64, respectively. The deduced amino acid sequences of t-actin 1 and t-actin 2 showed 45% and 43% identity, respectively, with the mouse actins, although the total number of amino acids was different. The T-ACTINs 1 and 2 proteins were highly homologous (56%) (Fig. 1B). Both cDNA sequences contained putative polyadenylation signals at nt 1368 and nt 1487 (Fig. 1A). The deduced amino acid sequences of the cDNAs revealed that T-ACTIN 1 and 2 contained the actins signature 1 (T-ACTIN 1, aa position 100–110; T-ACTIN 2, aa 122–132), the actins signature 2 (T-ACTIN 1, aa 400–408; T-ACTIN 2, aa 421–429), and the actin and actin-related protein (arp) signature (T-ACTIN 1, aa 151–163; T-ACTIN 2, aa 173–185) (Fig. 1B) (PROSITE accession PDOC00340). Both T-ACTINs 1 and 2 showed 45% identity with the consensus sequence for actins signature 1, whereas T-ACTINs 1 and 2 had 88% and 77% identity, respectively, with the sequence of actins signature 2. The two T-ACTINs had 76% identity with the consensus sequence of the actins and arp signature. Computer-assisted homology searches with the t-actin 1 (accession AB023062) and t-actin 2 (accession AB023063) nucleotide sequences revealed similarities to the mouse actin homologues (mACTl7b, AF113520, and mACTl7a, AF113519; respectively) and the human actin genes (ACTl7b, AF113527, and ACTl7a, AF113526, respectively), which have been implicated as candidate genes for familial dysautonomia or hereditary sensory and autonomic neuropathy type III, which is an autosomal recessive disorder that is almost exclusively limited to persons of Ashkenazi Jewish descent [28].
|
Expression of Testicular Actin mRNAs
Northern blot analyses showed that the t-actin mRNAs were expressed exclusively in the testis as major transcripts of 2.2 kb (t-actin 1) and 1.8 kb (t-actin 2). These transcripts were not detected in the brain, heart, intestine, kidney, liver, lung, muscle, ovary, or spleen. The t-actin mRNAs were also absent from the cryptorchid and mutant mouse testes (Fig. 2A), both of which lack differentiated germ cells [27]. Transcripts were absent from the neonatal to adolescent testis during male germ cell development in the first 23 days. Thereafter, positive signals appeared, and the signals became stronger up to adulthood (Fig. 2B). These mRNAs were transcribed in germ cells but not in supporting cells, such as Leydig and Sertoli cells (Fig. 2C). We performed in situ hybridization of the t-actin 1 and 2 mRNAs, to confirm and extend the results of the Northern blotting (Fig. 3). Hybridization signals for both t-actin-specific probes were observed in the seminiferous tubules of early spermatids at stages II/III–XII. The in situ hybridization experiments confirmed the Northern blot results and demonstrated the presence of the two mRNAs in round spermatids. In contrast, these transcripts were not found in either spermatogonia or spermatocytes at the early stages of germ cell differentiation or in elongated spermatids during the terminal stages of spermiogenesis. These results indicate that the timing of the expression of these two genes is similar in haploid spermatids between steps 2 and 13 (Fig. 3).
|
|
Expression of Testicular ACTIN Proteins
Western blot analyses with anti-T-ACTIN 1 and 2 rabbit antisera showed reactive bands of Mr 43 100 (T-ACTIN 1) and Mr 47 200 (T-ACTIN 2) in the testicular extracts but not in the other organs of the mouse (Fig. 4A). However, no signal was detected by Western blot analysis with each preimmunization rabbit serum (data not shown). These findings are consistent with the results of the Northern analysis (Fig. 2A). However, only T-ACTIN 2 showed a positive signal in sperm extracts. The two testicular actins were first detected in 3-wk-old testes, and the signal increased gradually with age and the development of the adult testis (Fig. 4B). Although the two proteins were detected in both the cytoplasm and nuclei of germ cells, the main subcellular locations of these two proteins were quite different; T-ACTIN 1 was primarily located in the cytoplasm, and T-ACTIN 2 was primarily located in the nuclei of testicular germ cells (Fig. 4C). Thus, these results indicate that both the transcription and translation of the t-actin 1 and 2 genes occur exclusively in haploid male germ cells, and the timing of gene expression is regulated precisely during the development of male germ cells, although the subcellular locations of these proteins are different.
|
Immunohistochemical Analyses
Immunohistochemical staining of the testicular cross sections showed that T-ACTIN 1 expression occurred during the late stages of spermatogenesis, i.e., in spermatids at developmental steps 2–16, and was found mainly in the spermatid cytoplasm (Fig. 5, A–D). However, T-ACTIN 1 was not detected in mature sperm (Fig. 4A; immunostaining data not shown). Because amorphous positive immunostaining appears at the centers of the step VIII seminiferous tubules, it seems that T-ACTIN 1 is discarded at the end of spermiogenesis (Fig. 5D). In contrast, T-ACTIN 2 was detected mainly in the nuclei of spermatids at developmental step 2 and in mature sperm (Fig. 6, A–D) and in the principal piece of the sperm flagellum (Fig. 7). Because amorphous positive immunostaining was seen at the centers of the step VIII seminiferous tubules, it appears that some of the T-ACTIN 2 is also discarded at the end of spermiogenesis. These observations are in agreement with the results of the Western blot analysis and indicate that T-ACTINs 1 and 2 are novel differentiation-associated proteins whose expression levels peak upon the appearance of the haploid spermatids. The subcellular localizations of T-ACTINs 1 and 2 were different; T-ACTIN 1 was present mainly in the cytoplasm of the round and elongated spermatids (Fig. 5), whereas T-ACTIN 2 was found in the nuclei of the spermatids (Fig. 6) and in the head and tail of the mature sperm (Fig. 7).
|
|
|
|
DISCUSSION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
We identified two novel haploid germ cell-specific actins that are involved in spermiogenesis. T-ACTIN 1 is located in the spermatid cytoplasm and is discarded at the end of spermiogenesis (Figs. 4C and 5, E–G), whereas T-ACTIN 2 is found in the spermatid nucleus (Figs. 4C and 6, E–G) and in the sperm head and tail (Fig. 7). Both T-ACTINs are produced concomitantly with spermiogenesis (Figs. 2B, 3, 4B, 5, and 6). Although the principal physiological locations of these two molecules were quite different (Figs. 4C, 5, E–G, 6, E–G, and 7), some of these proteins may associate with each other to form filamentous actin. Recently, we identified a novel actin-capping protein (CP-
3) in both mice and humans, which was expressed specifically in haploid spermatids [7, 21]. The formation of filamentous F-actin may be regulated by various proteins, such as thymosin-ß10 [13], ß-chimaerin [12], scinderin [10], and actin-capping proteins [7, 11]. These control proteins or CP-3 may act as regulatory or capping proteins for T-ACTINs 1 and 2. The actin-capping protein is attached to the barbed end of F-actin and is important in the control of actin filament elongation, because it acts as an actin-capping heterodimer. The presence of CP-ß3, which may be the partner of CP-3, was demonstrated recently in the testis [11].
Although the precise role of the actin cytoskeleton in sperm morphogenesis is unclear, filamentous actin becomes concentrated in the developing subacrosomal space of round spermatids and subsequently extends beneath the acrosome in the form of the spermatid elongate. This distribution pattern suggests that actin stabilizes the position and/or shape of the acrosome with respect to the nucleus [4, 9, 18, 19]. The potential roles of actin include anchorage of the acrosome or shaping of the acrosome head by the nuclear membrane. Therefore, actins may have an important role in determining the final shape of the mature sperm head, in acrosome formation, and in the acrosome reaction. Actin and actin-binding proteins may also participate in membrane remodeling and intracellular signaling during epididymal maturation and in the acrosome reaction [4, 15, 17]. Cytochalasins B and D and anti-actin antibodies inhibited the zona pellucida-induced acrosome reaction, whereas the inhibition of actin depolymerization with phalloidin had no effect, which indicates that actin polymerization is probably involved in the acrosome reaction in human sperm [16]. Aberrant expression of the actin-capping proteins, arps, or T-ACTINs in combination with defects in the associated molecules that control actin filaments may underlie many of the abnormalities in sperm morphology that are observed in infertile semen.
|
ACKNOWLEDGMENTS |
|---|
|
FOOTNOTES |
|---|
Received: 25 January 2003.
First decision: 17 February 2003.
Accepted: 26 March 2003.
|
REFERENCES |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
-actin in postmeiotic male germ cells of mice. Mol Cell Biol 1989 9:1875-1881This article has been cited by other articles:
|
|
 |
|
  K. Tokuhiro, M. Hirose, Y. Miyagawa, A. Tsujimura, S. Irie, A. Isotani, M. Okabe, Y. Toyama, C. Ito, K. Toshimori, et al. Meichroacidin Containing the Membrane Occupation and Recognition Nexus Motif Is Essential for Spermatozoa Morphogenesis J. Biol. Chem., July 4, 2008; 283(27): 19039 - 19048. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. Tokuhiro, Y. Miyagawa, S. Yamada, M. Hirose, H. Ohta, Y. Nishimune, and H. Tanaka The 193-Base Pair Gsg2 (Haspin) Promoter Region Regulates Germ Cell-Specific Expression Bidirectionally and Synchronously Biol Reprod, March 1, 2007; 76(3): 407 - 414. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Y. Nishimune and H. Tanaka Infertility Caused by Polymorphisms or Mutations in Spermatogenesis-Specific Genes J Androl, May 1, 2006; 27(3): 326 - 334. [Full Text] [PDF]
|
|
|
|
 |
|
 H. Obermann, I. Raabe, M. Balvers, B. Brunswig, W. Schulze, and C. Kirchhoff Novel testis-expressed profilin IV associated with acrosome biogenesis and spermatid elongation Mol. Hum. Reprod., January 1, 2005; 11(1): 53 - 64. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Kitamura, N. Iguchi, Y. Kaneko, H. Tanaka, and Y. Nishimune Characterization of a Novel Postacrosomal Perinuclear Theca-Specific Protein, CYPT1 Biol Reprod, December 1, 2004; 71(6): 1927 - 1935. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. D. Mruk and C. Y. Cheng Sertoli-Sertoli and Sertoli-Germ Cell Interactions and Their Significance in Germ Cell Movement in the Seminiferous Epithelium during Spermatogenesis Endocr. Rev., October 1, 2004; 25(5): 747 - 806. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Kitamura, H. Tanaka, and Y. Nishimune Haprin, a Novel Haploid Germ Cell-specific RING Finger Protein Involved in the Acrosome Reaction J. Biol. Chem., November 7, 2003; 278(45): 44417 - 44423. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
