SciELO - Scientific Electronic Library Online

 
vol.34 número3-4Transcriptome analysis and crop improvement: (A review)Problemas biofilosóficos y epistemológicos en el estudio de los seres vivos: En diálogo con el pensamiento de Humberto Maturana Romesín índice de autoresíndice de assuntospesquisa de artigos
Home Pagelista alfabética de periódicos  

Serviços Personalizados

Journal

Artigo

Indicadores

Links relacionados

Compartilhar


Biological Research

versão impressa ISSN 0716-9760

Biol. Res. v.34 n.3-4 Santiago  2001

http://dx.doi.org/10.4067/S0716-97602001000300004 

Human epididymal proteins and sperm function during
fertilization: un update

ANDREA LASSERRE 1,2 ROMINA BARROZO 1,2,* JORGE G. TEZÓN 2 PATRICIA V. MIRANDA 2, MÓNICA H. VAZQUEZ-LEVIN2

1 These authors have equally contributed to the work
2 Instituto de Biología y Medicina Experimental (IBYME)
Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET)
Vuelta de Obligado 2490 (1428) Capital Federal. Argentina.
* Departamento de Ciencias Biológicas. Facultad de Ciencias Exactas y Naturales.
(Biol Res 2001; 34 3-4: 165-178)

Correspondence to: Mónica H. Vazquez-Levin, Ph.D. Vuelta de Obligado 2490. 1428-BUENOS AIRES, ARGENTINA. Phone (54-11)47832869. Fax (54-11)47862564. Email: mhvaz@dna.uba.ar

Received : June 7, 2001. In Revised form : July 3, 2001. Accepted: July 9, 2001

INTRODUCTION

In contrast to the majority of invertebrates as well as vertebrates with external fertilization, most mammalian spermatozoa that have completed their morphogenesis in the testis are immotile and unable to interact with the egg. The acquisition of sperm fertilizing ability has been associated to metabolic and structural changes in the male gamete, particularly those occurring at the plasma membrane. These modifications take place during sperm transit through the epididymis in a complex process called sperm maturation (for reviews: Cooper, 1986; Robaire and Hermo, 1988). The sperm maturation process is followed by further changes occurring during ejaculation and in the female tract. The latter has been collectively called sperm capacitation (Yanagimachi, 1994).

The epididymis, a single convoluted duct, has been grossly divided into three major regions: caput, corpus, and cauda: they have been primarily distinguished by their epithelial cell morphology, their sperm fertilizing ability, and more recently, by their specific pattern of gene expression. The epididymis provides a luminal microenvironment for sperm maturation and storage under androgen control (Robaire and Hermo, 1988; Robaire and Viger, 1995; Tezón and Blaquier, 1981; Vazquez et al, 1986a,b, 1989). Numerous studies done in animals have suggested that specific secretory proteins produced in the epididymis associate to spermatozoa during transit through the organ and play a key role in the mammalian sperm maturation process by conferring the male gamete the ability to recognize the oocyte (Orgebin Crist and Fournier Delpech, 1982; Cuasnicú et al, 1984a,b,c, 1990; Gonzalez Echeverria et al, 1984; Moore 1981; Moore and Hartmann, 1986; Rochwerger et al, 1990). In humans, the existence of a sperm maturation process has been reported (Hinrichsen and Blaquier, 1980; Moore et al, 1983), and the synthesis and secretion of glycoproteins has been documented (Tezón et al, 1985a,b, 1987; Allen-Hoffmann and Mosher, 1987; Cooper et al, 1988; Ross et al, 1990; Moore et al, 1992; Miranda and Tezón, 1992; Boué et al, 1995, 1996; Miranda et al, 1995). Epididymal sperm-coating proteins may exert their effects on the male gamete already in the epididymis or become functional in the female tract. Some of the proteins produced by the epididymis, called sperm decapacitating factors, may associate to the cell surface and prevent premature sperm activation. Other epididymal proteins may act directly in the process of gamete recognition and interaction, by remaining on spermatozoa to mediate sperm binding to the zona pellucida and oocyte plasma membrane. Additionally, there are evidences suggesting an indirect participation of epididymal proteins upon fertilization (e.g. the alteration of the sperm surface by glycosylation, hydrolysis, lipid removal etc., see below). In addition to this role of the epididymis on sperm maturation, a system of regulated storage of spermatozoa in the distal region of the organ has been developed in mammals, ensuring that the stored cells are quiescent and unreactive (Bedford and Yanagimachi, 1991).

In the last ten years, the field of research on epididymal proteins and their function has advanced to the molecular level. There have been several reports characterizing epididymal proteins in different species. For some of them, the apparent molecular weight and their isoelectric point have been determined. In addition, immunolocalization with specific antibodies and, more recently, cDNA cloning and in situ hybridization have been done to determine the nucleotide sequences of the encoding genes as well as the amino acid composition of proteins of interest and their pattern of expression. Finally, cross-specie homology analysis has suggested their conservation as well as possible functions of these proteins. Rodents have been primarily the animal models in many studies. However, identification of homologous gene products in other mammals, particularly humans, has been difficult, in part by poor evolutionary conservation. An important recent development has been the use of specific cDNA libraries to examine gene expression in the human epididymis (see Kirchhoff, 1999; Légaré et al, 1999). However, the identity and function of epididymal secretory proteins in sperm maturation and the mechanisms by which their genes are controlled have yet not been completely elucidated.

This review is aimed at summarizing some aspects of the biochemical, molecular, and functional characterization recently completed in a group of human epididymal proteins, in the light of their potential role in the sperm maturation process, with special interest in their participation in the interaction with the zona pellucida, and in the sperm-egg fusion process.

 

EPIDIDYMAL PROTEINS POTENTIALLY INVOLVED IN SPERM-EGG INTERACTION

Interaction with the zona pellucida

Capacitated spermatozoa initially interact, in a specie-specific manner, with the zona pellucida (ZP), an extracellular coat that surrounds all mammalian eggs. This process, called primary binding, is mediated by ZP glycoconjugates that recognize sperm receptors located on the surface of the male gamete. Bound spermatozoa undergo the acrosome reaction, and initiate penetration of the ZP. Sperm penetration involves digestion of the ZP and vigorous sperm motion, while keeping the sperm associated to the matrix, via sperm receptors; this interaction has been named secondary binding (Yanagimachi, 1994; Wassarman, 1999; Brewis and Wong, 1999). Numerous candidates have been postulated as sperm receptors for primary and secondary binding. Several proteins of epididymal origin proposed for these roles are described in the following paragraphs, and summarized in Table 1.

P34H

P34H is a 34 KDa human epididymal sperm protein synthesized and secreted predominantly by the principal cells in the proximal and distal section of the corpus epididymis. The predicted amino acid sequence reveals a high similarity with members of the short chain dehydrogenase / reductase family proteins. Moreover, it shows 65% identity with P26h, the hamster counterpart (Légaré et al, 1999). P26h has been characterized at the molecular level (Gaudreault et al, 1999), and its participation in the sperm _ egg interaction process has been well studied (Bérubé and Sullivan, 1994; Bégin et al, 1995; Bérubé et al, 1996). Immunocytochemical analysis with a polyclonal anti-P26h antibody shows a faint staining in human spermatozoa from the caput, while the majority of the cells in the cauda epididymis are stained, with a signal restricted to the acrosomal cap. The staining remains in capacitated spermatozoa, but apparently is lost during the acrosome reaction (Boué et al, 1994,1996). The presence of anti-P26h significantly reduced the ability of human spermatozoa to bind to homologous ZP, without affecting sperm acrosome reaction and fusion to the oolema (Boué et al, 1994). Finally, staining with anti-P26h has allowed the identification of infertile patients showing reduced P34H levels (Boué and Sullivan, 1996). Altogether, these evidences would suggest that P34H is a human epididymal protein involved in sperm interaction with the ZP.

ß-HEXOSAMINIDASE

N-acetylglucosaminidase (NAG, E.C. 3.2.1.30) its a glycosidase reponsible for the hydrolysis of non reducing terminal N-acetylglucosamine (GlcNAc) residues from ß-glycosidic boundaries in numerous glycoconjugates. In mammals, the enzyme NAG is named ß-hexosaminidase (Hex, E.C. 3.2.1.52), since it also hydrolyses N-acetylgalactosamine. The significance of this enzyme upon normal epididymal function has been suggested by the severe anomalies observed in Hex-knock out mice (Trasler et al, 1998; Adamali et al, 1999a, 1999b). Moreover, several experimental evidences would support its involvement in the participation of GlcNAc residues on human sperm-ZP interaction (Miranda et al, 1995, 1997,2000). It was found that incubation of capacitated spermatozoa with bovine serum albumin (BSA)GlcNAc (Brandelli et al., 1994) triggers the acrosome reaction, following a mechanism that resembles induction by homologous ZP (Brandelli et al, 1994, 1996). Moreover, preincubation of capacitated spermatozoa with GlcNAc was found to inhibit their ability to interact with the ZP (Miranda et al, 1997). The human epidydimal enzyme has been characterized at a biochemical level (Miranda et al, 1995). Recently, participation of Hex in early events of sperm-oocyte interaction has been suggested by studies showing that gamete incubation in the presence of human recombinant Hex or its substrate precludes the interaction of human spermatozoa with homologous ZP (Miranda et al, 2000). However, whether the epididymal enzyme is responsible for such role on the sperm-ZP interaction process remains to be demonstrated.

Table 1
Human sperm proteins of epididymal origin proposed to interact with the ZP.


Protein

MW
(KDa)

Tissue
Localization*

Localization
on spermatozoa
Identity Homology with
other species

P34H

34 E:CA.CO,CU
and others
Acrosomal cap of
ejaculated spermatozoa.
Decrease along the
capacitation.
Absent in acrosome
reacted cells

short chain
deshydrogena
sa/reductase

hamster: 65% to
P26h
pig: 71% to
carbonyl
reductase of lung

ß-Hex

65

E

glycosidase

4A8

110 E Entire head of
ejaculated cells.
Increased in
capacitated cells.
Unique No

1G12

15-25 E: CO.
T

Entire spermatozoa
without changes during
or after capacitation or
acrosome reaction
lymphocyte
antigen
No

SOB3

18-19 E: CU and
others
Ejaculated cells: neck
and tail.
Acrosome reacted cells:
neck, tail and acrosomal
region.
Antimicrobial
protein
No

*E: epididymis (CA: caput, CO: corpus; CU: cauda). T: testis

4A8

The monoclonal antibody (mAb) named mAb4A8 was selected for its ability to inhibit human sperm-egg binding and penetration in a dose-dependent manner. It recognizes an epitope in a human epididymal and sperm surface protein not fully characterized. The antibody was found to specifically stain epithelial cells of the human epididymis and the spermatozoa in the lumen. Non glycosylated seminal plasma showed a weak reactivity in a 110 KDa protein band, while antigenic polypeptides of 78, 56 and 44 KDa were identified in glycosylated seminal plasma proteins (Batova et al, 1998). No further studies have been reported upon its role in fertilization.

IG12

The mAbIG12 was selected for its ability to immobilize and agglutinate human spermatozoa. Independently of these effects, a blockage of in vitro fertilization was observed in the presence of the antibody. The inhibitory effect was evidenced in a diminished sperm binding and penetration into the ZP. Moreover, a decrease in the number of sperm penetrations in the ZP free-hamster egg sperm penetration assay was seen in the presence of mAb IG12. Immunostaining of ejaculated spermatozoa reveals a positive granular pattern over the entire surface of the cell. By immunohistochemistry, mAbIG12 strongly stained epithelial cells of the corpus epididymis. Moreover, mAbIG12 recognized a single band of under 15 KDa on deglycosylated sperm extracts (Komori et al, 1997). No other studies have been reported until the present time.

SOB3

A sperm protein named SOB3 was identified using the mAbLB5 developed towards human sperm proteins. mAbLB5 stained the neck and flagellum of most spermatozoa and the acrosome of 10-20 % of the cells. In Western immunoblot analysis, it recognized two bands of 18 and 19 KDa in spermatozoa and cauda epididymis protein extracts. Presence of the antibody in an in vitro binding assay was associated with a decrease in the number of sperm bound to the ZP. Some of the studies would suggest that SOB3 may become accessible once spermatozoa have completed the acrosome reaction, and consequently, an involvement in secondary binding to the ZP has been postulated (Martin Ruiz et al, 1998). Recently, cloning and sequencing of SOB3 has been reported, revealing a single copy gene with 98% of homology with prepro-FALL39 and 100% homology of CAP18, two human genes which encode an antimicrobial protein (Hammami-Hamza, 2001). The authors suggest two roles for SOB3: sperm protection against bacterial injury and secondary binding to the zona pellucida.

Sperm-egg fusion proteins

Acrosome reacted spermatozoa that have completed zona pellucida penetration, reach the perivitelline space, bind and fuse to the egg plasma membrane, release the genetic material and initiate zygote development. Sperm fusion to the ooplasma has been associated to fusion events described in viral particles. Protein complexes with both functions, binding and fusion, would be present in both the egg and the spermatozoa and would interact with the counterpart on the surface of the other gamete (Snell and White, 1996; Töpfer-Petersen, 1999). Sperm binding to the oolema has been proposed to involve interaction of sperm receptors located in the equatorial segment and postacrosomal region with carbohydrates from oligosaccharides located on the egg plasma membrane (Gabriele et al, 1998; Fusi et al, 1996). Several sperm adhesion molecules have been described, being the protein named fertilin the one best studied (Myles et al, 1994). Regarding the sperm-egg fusion process itself, the actual mechanism is yet unknown. As seen in studies done with virus, fusion proteins may have sequences of hydrophobic residues and mostly of alpha-helical structure (fusion peptides); interaction of the sperm fusion peptide with the egg plasma membrane could destabilize the membrane, following a process that would end with the formation of the fusion pore (Pecheur et al, 1999).Several proteins of epididymal origin have been proposed to participate in the sperm _ egg fusion process. Some of them are described in the following paragraphs and listed in Table 2.

ARP

Two different research groups have cloned a human epididymal secretory protein named ARP (AEG Related Protein) (Hayashi et al, 1996, Kratzschmar et al, 1996). This protein is the human counterpart of the well studied rat and mouse epididymal protein DE (named by Cameo and Blaquier, 1976; AEG, Acidic Epididymal Glycoprotein; Lea et al, 1978). DE has been characterized as a putative sperm receptor in the egg fusion process (Cuasnicú et al, 1984a,b,c; Rochwerger and Cuasnicú, 1992; Rochwerger et al, 1992; Perez-Martínez et al, 1995; Ellerman et al, 1998). Among others, ARP and DE belong to the CRISP family members that share a highly conserved cluster of cysteines near the carboxy-terminus. The epididymal origin and secretory nature of ARP, and its localization on the sperm head has suggested a participation for this protein in the fusion process. ARP has been detected on the human sperm head after the acrosome reaction (Cohen et al, 2000). Moreover, in the zona-free hamster egg sperm penetration assay, an anti-ARP antibody had an inhibitory effect upon the number of penetrating human spermatozoa. Using the recombinant ARP expressed in bacteria, human oocytes revealed in their surface the presence of complementary sites to ARP (Cohen et al, 2000). All these evidences strongly support a role of ARP in human sperm-egg fusion process.

TABLE 2

Human sperm proteins of epididymal origin proposed to participate in sperm fusion to the
oocyte.


Protein

MW
(KDa)
Tissue
Localization*
Localization
on spermatozoa
Identity Homology with
other species

ARP

30 E: CA,CO,CU postacrosomal
region of ejaculated
and acros reacted
spermatozoa
cystein rich
protein
rat: 38% to DE/AEG
mouse: 40.6% to CRISP1;
41.2% to Tpx1; 38.8% to
CRISP3 macaque: 85%
to mAEG

SOB2

17.5,18
and 19
E: CA, CO.
ovary, placenta
Ejaculated spermatozoa:
postacrosomal region
and neck
unique

rat: protein of 26 KDa
hamster: protein of 28KDa
rabbit: protein of 50 KDa

FLB1

94-100 E: CA, CO. epidermal
cytokeratins

macaque, rat, mouse
and rabbit (100 KDa)

gp20

20

E

Ejaculated spermatozoa:
head and sperm tail
Capacitated spermatozoa:
equatorial region
unique protein not found on
spermatozoa or tissue of
rodents

E-cad

120-130 E

sperm head of ejaculated,
capacitated and acrosome
reacted spermatozoa

CAM rat / mouse: 90%

Fibronectin

30-35 E: CA   RGD (Arg-
Gly-Asp)
Dog

*E: epididymis (CA: caput, CO: corpus; CU: cauda). T: testis

SOB2

The Sperm Oocyte Binding 2 protein (SOB2) has been purified using preparative electrophoresis of detergent extracts from human spermatozoa, after protein identification in Western immunoblotting with the mAbG12. The antibody specifically stained the epithelium and spermatozoa of the caput and corpus epididymis, but did not show any signal in testis and efferent ducts. The antibody did not have any inhibitory effect upon sperm motility, interaction with the ZP or the acrosome reaction. On the other side, mAbG12 Fab fragments were found to strongly inhibit binding and penetration of human spermatozoa to zona free-hamster oocytes (Boué et al, 1992; Lefèvre et al, 1997), suggesting a role of SOB2 in sperm-oocyte fusion.

FLB1

A protein referred as FLB1 has been identified using a mAbCA6 raised against human sperm proteins. The epididymal protein was reported to be composed of two chains of 47 KDa and similar pI (5.8, 5.9), and was found to be homologous to cytokeratins 1 and 10. It was localized on the equatorial region of mouse, rat and hamster cauda epididymal spermatozoa, and in human, macaque, and rabbit ejaculated spermatozoa. It would progressively coat spermatozoa during their transit through the epididymis (Boué et al, 1995). In the presence of the mAbCA6, binding of human spermatozoa to zona free

hamster and human eggs was inhibited, without having an effect upon sperm forward progressive motility or cell interaction with the ZP (Boué et al, 1995).

GP20

A sialylglycoprotein of ~20 KDa (gp20) was purified by preparative electrophoresis of detergent extracts from human spermatozoa, and utilized to develop a polyclonal antiserum. The anti-gp20 antiserum allowed localization of the protein in capacitated spermatozoa over the equatorial segment in 90% of the cells. In addition, the strong staining seen in principal cells of the epithelial epididymis, but not in testis sections suggested its epididymal origin. In the zona free hamster egg sperm penetration assay, presence of anti-gp20 antibodies precluded sperm binding and penetration to the oocyte, again suggesting a role for this protein in the sperm-egg fusion process (Focarelli et al, 1998). GP20 was found to have in the N-terminal sequence a 100% homology with that of the lymphocyte antigen CDw52 (Hale et al, 1990). An evaluation using two dimensional electrophoresis and MALDI revealed two components of the gp20 sperm antigen (Focarelli et al, 1999).

EPITHELIAL CADHERINS

Cadherins are a family of Cell Adhesion Molecules (CAM) known to specifically bind to other cadherins located on adjacent cells (Edelman, 1988; Takeichi, 1990). They are calcium dependent surface glycoproteins protected from proteolysis through stabilization by this ion, without which the molecule may undergo conformational changes making it susceptible to digestion (Takeichi, 1995). Cadherins have been involved in several signal transduction pathways through tyrosine kinases associated with adhesion sites, affecting association with the cytoskeleton (Salomon et al, 1992). To the present time, more than 40 different cadherin types have been identified. Among them, one of the major families is the epithelial cadherins (E-cad). Presence of E-cad in the human epididymal epithelium was demonstrated by immunohistochemistry (Anderson et al, 1994). In the rat, the mRNA encoding E-cad was found to be present and translated in the epididymis, differentially distributed in the tissue, and regulated by circulating androgens (Cyr et al, 1992). Recently, E-cad were reported to be present on the surface of both human gametes (Rufas et al, 2000). Characterization of the expression of human E-cadherin in the human male tract, and their role in the process of sperm interaction with the egg is currently under investigation in our laboratory.

FIBRONECTIN

The involvement of integrins in the interaction of spermatozoa with the oocyte plasma membrane has been supported by several findings. In particular, Bronson and Fusi (1990) first demonstrated that a ligand recognition motif for integrins, RGD (Arg-Gly-Asp), would participate in binding and penetration of human spermatozoa to zona free hamster oocytes. Moreover, RGD binding receptors were identified on the human egg plasma membrane (Fusi et al, 1992). Regarding human spermatozoa, it is known that testicular cells express ß3, ß5 and ß6 chains of ß1 integrin, and fibronectin, which is known to contain an RGD sequence (Schaller et al, 1993). Fibronectin has been identified as a epidydimal secretory protein and proposed as a marker of human sperm maturation (Miranda and Tezón, 1992).

Recently, using the strategy of differential epididymal cDNA library screening, novel specific human epididymal proteins containing four fibronectin type II (Fn2) modules were identified (Saalmann et al, 2001). In the report, homologous mRNAs were also found in several animal species. Moreover, a specific antipeptide antiserum recognized 30-35 kDa protein bands in extracts from human epididymal tissue, in fluid from the cauda, and in detergent extracts of ejaculated spermatozoa. In canines, a specific signal was also found in spermatozoa from the cauda epididymis, although no proteins were detected in protein extracts from caput spermatozoa. The role of this protein in sperm function has yet to be determined, though in bovine, binding of major proteins of the seminal plasma (BSP), which are similar but not homologous to this novel protein, would help sperm capacitation (Miller et al, 1990).

Table 3
Other human epididymal gene products


Protein

MW
KDa)
Tissue
localization*
Localization on
spermatozoa
Identity/
proposed function
Homology with
other species

HE1

25-27 E: CO, CU Weak binding
to reacted
spermatozoa
decapacitating factor,
maintaining the
cholesterol content
of male gametes
chimpanzee: EPI-1
96% pig: EP4

HE2

10 E:CO Acrosomal region unique Absent in rat, mouse,
cat, pig and marmoset
HE3

14.9

E: CA Weak binding unique absent in rat, dog,
horse, mouse and ram
pig: mRNA of HE3

HE4

10 E: CO, CU Weak binding decapacitating factor
"four disulfide core"
bovine: HE4-like
absent in rat-mouse

HE5/CD52

18-25 E: CU Ejaculated
spermatozoa:
tail, postacrosomal
region
CD52 antigen (99%)
protection from
immune attack
rat: SMEmG
mouse: B7 dog: CE5
Callitrix: mRNA
(from epididymis)

HE6

110 E glycoprotein receptor
signal transduction
mechanism

insect: DHR 29%
rat, mouse, dog,
cat, horse

Clusterin

70-73 E bovine: ß-chain of
clusterin 60%
rat, mouse and ram
Gp83

83

E

Semenogelin

71-76 E:CA,CO,CU Whole cell secretory protein
gel formation of
ejaculated semen
rodents: seminal
vesicle secretory
protein-IV
guinea pig: GPI

Ubiquitin

8.5 E and others Head and tail
of ejaculated
Marker of defective
spermatozoa
Ungulates, rodents,
primates

  *E: epididymis (CA: caput, CO: corpus; CU: cauda). T: testis

Other human epididymis _ specific gene products (Table 3)

HUMAN EPIDIDYMAL cDNA FAMILIES HE1-HE6

Using a strategy of differential secreening of human epididymal cDNA libraries, a set of six major secretory proteins of human epididymal epithelial cells, named HE1-HE6, were identified (Kirchhoff et al, 1990). With the exception of HE5, the others are novel (human) gene products, not previously reported in animal studies. A brief description of HE1, HE2, HE3, HE4, HE5, and HE6 molecular characterization is presented in the following paragraphs.

HE1: HE1 is a major secretory glycoprotein detected as a ~ 20 KDa protein, present within the epithelium and lumen of the corpus epididymis, and accumulated in the lumen of cauda epididymis and vas deferens. The mRNA encoding HE1 was found to be the most abundant gene product obtained using the strategy of differential screening. HE1 is well conserved among mammals (in chimpanzee, named EPI-1 (Perry et al, 1995; Fröhlich and Young, 1996) and boar, named EP4 (Parry et al, 1992) suggesting a common functional role in the mammalian epididymis. Moreover, HE1 N-terminus is almost identical to the same amino acid sequence of ram lipid transfer proteins, suggesting for HE1 a role in maintaining the high cholesterol content of spermatozoa during epididymal transit and storage (Kirchhoff et al, 1996).

HE2: The cDNA clone encoding HE2 was identified and characterized to encode a small secretory protein, specifically produced by the caput epididymal epithelium. Antibodies raised towards a recombinant HE2 produced in bacteria showed specific staining over the equatorial region of human ejaculated spermatozoa, suggesting its putative participation in sperm-oocyte interaction (Osterhoff et al, 1994). However, is it still not clear whether HE2 has a role in the sperm-egg interaction process, considering the lack of a significant effect of the HE2 antisera tested in the outcome of ZP binding test and the hamster egg sperm penetration assay (Kirchhoff, 1999).

HE3: Similarly to HE2, a major epididymal mRNA named HE3 encodes a Human Epididymal Protein 3, and predicts a small secretory glycoprotein (Mr 14900). HE3 is expressed in a highly regionalized manner in the epididymis. The epididymal caput is the region where it is abundant. Analysis of human genomic DNA has shown that HE3 presents three independent related genes, a, ß, g. However only a, b seem to be expressed by the human epididymis, while HE3-g probably is a nonfunctional pseudogene (Kirchhoff et al, 1994). Like HE2, HE3 is poorly conserved among mammals. The lack of homology with other known protein sequences makes difficult any speculation on its potential role in sperm maturation.

HE4: the cDNA sequence of a major human epididymis gene product called HE4 was described to encode a small (approximately 10 KDa) acidic secretory protein. The position of the half-cysteins suggested HE4 to be a two-domain member of the "four-disulfide core" or Whey Acidic Protein (WAP)-domain proteins, examples of which are HUSI-I/SLPI and guinea pig caltrin seminal plasma proteinase inhibitors (Kirchhoff et al, 1991). Anti HE4 antibodies give an specific signal in epididymal epithelium and duct lumen, as well as in the surface of ejaculated human spermatozoa. Dissociation of HE4 from the spermatozoa during capacitation would suggest a role as a decapacitating factor (Kirchhoff, 1998). A high nucleotide homology (76.8%) to HE4 was found in the rabbit protein BE-20, which was structurally related to extracellular proteinase inhibitors and only found in the epididymis (Fan et al, 1999).

HE5: The gene product HE5 was found to be abundant in the epithelial cells of the distal epididymis and deferent duct, in spermatozoa of the tubule, and in blood lymphocytes (Pera et al, 1996; Kirchhoff and Hale, 1996; Yeung et al, 1997). It is identical (99 % sequence homology) to the CD52 antigen which is expressed on the cell surface of human lymphocytes (Kirchhoff et al, 1993). HE5/CD52 transcripts are abundant only in the epididymis and vas deferens epithelial cells and in blood lymphocytes. Kirchhoff and Hale (1996) have proposed for HE5 a role in protection of spermatozoa from immune attack during maturation, storage and fertilization. The epididymal protein identified as gp20 has been found to be a homologue of HE5 (see above).

HE6: A novel gene product called HE6 was found to be expressed within the epithelial cells lining the human epididymal duct, mainly in the caput region. HE6 was found to be highly conserved among mammals. It shows homology with the seven-transmembrane-domain (Tm7) receptor superfamily, typical of the majority of G-protein_coupled cell-surface hormone receptors. Within the Tm7 family, HE6 shows low but significant homology to the secretin/VIP family. The predicted N-terminal extension was found to be longer than that of this family, and was similar to the highly glycosylated mucin-like surface molecules (Osterhoff et al, 1997). However, its possible involvement in the signal transduction mechanism remains to be determined.

CLUSTERIN

Clusterin or sulphated glycoprotein-2 (SGP-2) is an abundant glycoprotein apparently produced by the testis, epididiymis and seminal vesicles (O´Bryan et al, 1994a). In the rat, the androgen regulation of SGP-2 and its sperm association in the epididymis suggests the presence of specific protein functions in this tissue and others where it is also expressed (Sylvester et al, 1991). Hermo et al (1991) have proposed that the testicular SGP-2 is released from spermatozoa when it leaves the seminiferous tubule and is removed by the epithelium of the rete testis and efferent ducts. Subsequently, SGP-2 is replaced by the protein secreted by the caput epididymis, suggesting a role in sperm maturation. Immunocytochemical studies on human spermatozoa revealed surface coating with the 80KDa conventional heterodimeric clusterin, while normal cells showed over the acrosomal cap a different form of clusterin which is reactive to a anticlusterin a-chain antibody (O´Bryan et al, 1994b). Participation of clusterin in the sperm-egg interaction process has yet not been reported.

GP83 and GP39

Using Wheat Germ Agglutinin (WGA), Concanavalin-A (Con-A) and (PNA) lectins, human epididymal proteins GP83 and GP39, named after their Mr, were identified. In tissue homogenates and epididymal fluids, WGA and Con-A recognized GP83 and GP39 while only GP39 was detected when using PNA. Moreover, an specific signal for GP83 and GP39 was obtained in sperm membrane extracts recovered from the corpus and cauda regions when using WGA (Liu et al, 2000). In a subsequent study (Sun et al, 2000), GP83 was purified from human seminal fluid and partially characterized. An specific antibody localized the protein in the corpus and cauda epididymis, specifically in the supranuclear region and cell membrane of principal cells as well in the luminal content. Immunolocalization of GP83 in spermatozoa showed an specific staining over the acrosome of ejaculated and capacitated cells, and a signal over the equatorial region in acrosome reacted spermatozoa, suggesting redistribution of the antigen. Participation of GP83 in the sperm _ egg interaction process has yet not been reported.

SEMENOGELINS

The semenogelins are predominant secretory proteins from the human seminal vesicles responsible for the gel formation of freshly ejaculated semen (Lilja and Laurell, 1984; 1985). In addition, the mRNA encoding SgII was detected, although in lower levels, in the caudal epididymal epithelium (Lilja et al, 1992; Bjartell et al, 1996). Moreover, SgII was localized in the posterior part of the head, midpiece, and tail of ejaculated spermatozoa (Bjartell et al, 1996). The role of SgII in the cauda region of the epididymis and on the spermatozoa remains to be established.

UBIQUITIN

Ubiquitin, a 8.5 KDa protein, is a universal marker of proteolysis (Ciechanover, 1994). Recently, Sutovsky et al (2000, 2001a) demonstrated that ubiquitin secreted by the epididymal epithelium binds to the surface of defective spermatozoa, followed by phagocytosis of most ubiquitinated spermatozoa by epididymal epithelial cells. These findings of cell-surface ubiquitination in defective spermatozoa provides a possible mechanism for sperm quality control in mammals. Moreover, ubiquitination can be utilized as a new marker for semen abnormalities. In a recent report (Sutovsky et al, 2001b), a "sperm-ubiquitin tag immunoassay (SUTI)" was described as a valuable new tool for infertility diagnosis and prediction of IVF success in subfertile men diagnosed with idiopathic infertility.

CONCLUSIONS

This review has briefly shown some findings reported by several investigators on a set of human epididymal proteins identified and partially characterized in the recent years. The results suggest that the epididymis produces and secretes numerous proteins that would associate to the spermatozoa while they are transiting through the organ. These components would dramatically affect sperm functionality, allowing the male gamete to recognize the oocyte. However, much still is waiting to be done to comprehend this phenomenon. Implementation of genomics and proteomics will help, in the near future, to further characterize some already identified proteins, as well as to describe novel epididymal components, anticipating great advances in the elucidation of the sperm maturation process in humans.

ACKNOWLEDGEMENTS

This work was supported by grants from the World Health Organization (grant # 97175), the Agencia Nacional de Promoción de Ciencia y Tecnología (PICT97 00207), and the Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET) of Argentina (PIP 4404).

REFERENCES

ADAMALI HI, SOMANI IH, HUANG JQ, MAHURAN D, GRAVEL RA, TRASLER JM, HERMO LI (1999a) Abnormalities in cells of the testis, efferent ducts, and epididymis in juvenile and adult mice with ß-Hexosaminidase A and B deficiency. J Androl 20 (6): 778-802.         [ Links ]

ADAMALI HI, SOMANI IH, HUANG JQ, MAHURAN D, GRAVEL RA, TRASLER JM, HERMO LI (1999a) Characterization and development of the regional- and cellular-specific abnormalities in the epididymis of mice with ß-Hexosaminidase A deficiency. J Androl 20: 803-824.         [ Links ]

ALLEN-HOFFMANN BL, MOSHER DF (1987) Matrix assembly sites for exogenous fibronection are decreased on human fibroblast after treatment with agents with increase intracellular cAMP. J Biol Chem 262: 14361-14365.         [ Links ]

ANDERSON A, KLAUSS E, SKAKKEBEK NE (1994) Expression and localization of N-and E-cadherin in the human testis and epididymis. Int J Androl 17: 174-180.         [ Links ]

BATOVA IN, IVANOVA MD, MOLLOVA MV, KYURKCHIEV SD (1998) Human sperm surface glycoprotein involved in sperm-zona pellucida interaction. Int J Androl 21: 141-153.         [ Links ]

BEDFORD JM (1967) Effect of duct ligation on the fertilising capacity of spermatozoa in the epididymis. J Exp Zool 166: 271-281.         [ Links ]

BEDFORD M, YANAGIMACHI R (1991) Epididymal storage at abdominal temperature reduces the time required for capacitation of hamster spermatozoa. J Reprod Fertil 91: 403-10.         [ Links ]

BEGIN S, BERUBE B, BOUE F, SULLIVAN R (1995) Comparative immunoreactivity of mouse and hamster sperm proteins recognized by an anti-p26h hamster sperm protein. Mol Reprod Dev 41: 249-256.         [ Links ]

BERUBE B, LEFEVRE L, COUTU L, SULLIVAN R (1996) Regulation of the epididymal synthesis of P26h, a hamster sperm protein. J Androl 17: 104-110.         [ Links ]

BERUBE B, SULLIVAN R (1994) Inhibition of in vivo fertilization by active immunization of male hamsters against a 26-kDa sperm glycoprotein. Biol Reprod 51: 1255-1263.         [ Links ]

BJARTELL A, MALM J, MOLLER C, GUNNARSSON M, LUNDWALL A, LILJA A (1996) Distribution and tissue expression of semenogelin I and II in man as demonstrated by in situ hybrydization and immunocytochemistry. J Androl 17:17-26.         [ Links ]

BOUE F, BERUBE B, DE LARMINADE E, GAGNON C, SULLIVAN R (1994) Human sperm-zona pellucida interaction is inhibited by an antiserum against a hamster sperm protein. Biol Reprod 51: 577-587.         [ Links ]

BOUE F, BLAIS J, SULLIVAN R (1996) Surface localization of P34H, an epididymal protein, during maturation, capacitation, and acrosome reaction of human spermatozoa. Biol Reprod 54: 1009-1017.         [ Links ]

BOUE F, DUQUENNE C, LASSALLE B, LEFEVRE A, FINAZ C (1995) FLB1, a human protein of epididymal origin that is involved in the sperm-oocyte recognition process. Biol Reprod 52: 267-278.         [ Links ]

BOUE F, LASSALLE B, DUQUENNE C LEFEVRE A, FINAZ C (1992) Human sperm proteins from testicular and epididymal origin that participate in fertilization: modulation of sperm binding to zona-free hamster oocytes, using monoclonal antibodies. Mol Reprod Dev 33: 470-480.         [ Links ]

BOUE F, SULLIVAN R (1996) Cases of human infertility are associated with the absence of P34H, an epididymal sperm antigen. Biol Reprod 54: 1018-1024.         [ Links ]

BRANDELLI A, MIRANDA PV, AÑON-VAZQUEZ MG, MARIN-BRIGGILER CI, SANJURJO C, GONZALEZ-ECHEVERRÍA F, BLAQUIER JA, TEZON JG (1995) A new predictive test for in vitro fertilization based on the induction of sperm acrosome reaction by N-acetylglucosamine-neoglycoprotein. Hum Reprod 10: 1751-1756.         [ Links ]

BRANDELLI A, MIRANDA PV, TEZON JG (1994) Participation of glycosylated residues in the human sperm acrosome reaction: Possible role of N-acetylglucosaminidase. Biochim Biophys Acta Mol Cell Res 1220: 299-304.         [ Links ]

BRANDELLIi A, MIRANDA PV, TEZON JG (1996) Voltage-dependent calcium channels and Gi regulatory protein mediate the human sperm acrosomal exocytosis induced by N-acetylglucosaminyl/mannosyl neoglycoproteins. J Androl 17: 522-529.

BREWIS IA, WONG CH (1999) Gamete recognition: sperm proteins that interact with the egg zona pellucida. Reviews in Reproduction 4: 135-142.         [ Links ]

BRONSON RA, FUSI FM (1990) Sperm-oolemal interaction: role of the Arg-Gly-Asp (RGD) adhesion peptide. Fertil Steril 54: 527-529.         [ Links ]

CAMEO MS, BLAQUIER JA. (1976) Androgen-controlled specific proteins in rat epididymis. J Endocrinol 69: 47-55.         [ Links ]

CIECHANOVER A (1994) The ubiquitin-proteasome proteolytic pathway. Cell 79:13-21.         [ Links ]

COHEN D, ELLERMAN D, BUSSO D, PIAZZA A, YOUNG E, CUASNICU PS (2000) Participación de la proteína epididimaria ARP y sus sitios complementarios en el ovocito en el proceso de fusión de gametas humanas. Medicina 60: 762.         [ Links ]

COOPER TG (1986) The epididymis: sperm maturation and fertilisation. In: The male gamete: from basic science to clinical applications. C.Gagnon (ed)., pp 268-272 Cache River Press, Viena IL USA.         [ Links ]

COOPER TG, YEUNG CH, NASHLAN D, NIESCHLAG E (1988) Epididymal markers in human infertility. J Androl 9: 91-102.         [ Links ]

CUASNICU PS, CONESA D, ROCHWERGER L (1990) Potential contraceptive use of an epididymal protein that participate in fertilization. In: Alexander NJ, Griffin D, Spieler JM, Waites GMH (ed). Gamete interaction. Prospects for immunocontraception. New York. John Wiley Liss & Sons, Inc. pp. 143-153.         [ Links ]

CUASNICU PS, GONZALEZ ECHEVERRIA MF, PIAZZA A, BLAQUIER JA (1984a) Addition of androgens to cultures hamster epididymis increases zona recognition by immature spermatozoa. J Reprod Fertil 70: 541-547.         [ Links ]

CUASNICU PS, GONZALEZ ECHEVERRIA MF, PIAZZA A, CAMEO M, BLAQUIER JA (1984b) Antibodies against epididymal glycoproteins block fertilizing ability in rat. J Reprod Fert, 72: 467-471.         [ Links ]

CUASNICU PS, GONZALEZ ECHEVERRIA MF, PIAZZA A, PINEIRO L, BLAQUIER JA (1984c) Epididymal proteins mimic the androgenic effect on zona pellucida recognition by immature hamster spermatozoa. J Reprod Fertil 71: 427-431.         [ Links ]

CYR DG, HERMO L, BLASHUK, AND ROBAIRE B (1992) Distribution and regulation of epithelial cadherin messenger ribonucleic acid and immunocytochemical localization of epithelial cadherin in the rat epididymis. Endocrinology 130: 353-363.         [ Links ]

EDELMAN GM (1988) Morphoregulatory molecules. Biochemistry 27:3533-3543.         [ Links ]

ELLERMAN DA, BRANTUA VS, PEREZ MARTINEZ S, COHEN DJ, CONCESA D, CUASNICU PS (1998). Potential contraceptive use of epididymal proteins: immunization of male rats with epididymal rpotein DE inhibits sperm fusion ability. Biol Reprod 59: 1029-1036.         [ Links ]

FAN HY, MIAO SY, WANG LF (1999). Expression an characterization of an epididymis-specific gene. Arch Androl 42: 63-69.         [ Links ]

FOCARELLI R, DELLA GOIVAMPAOLA C, SERAGLIA R, BRETTONI C, SABATINI L, PESCAGLINI M, ROSATI F (1999) Biochemical and MALDI analysis of the human sperm antigen gp20, homologue of leukocyte CD52. Biochem Biophys Res Commun 258: 639-643.         [ Links ]

FOCARELLI R, GIUFFRIDA A, CAPPARELLI S, SCIBONA M, MENCHINI FABRIS F, FRANCAVILLA F, FRANCAVILLA S, DELLA GIOVAMPOLA C, ROSATI F (1998) Specific localization in the equatorial region of gp20, a 20 kDa sialylglycoprotein of the capacitated human spermatozoon acquired during epididymal transit which is necessary to penetrate zona-free hamster eggs. Mol Human Reprod 4:119-125.         [ Links ]

FRÖHLICH O, YOUNG LG (1996) Molecular cloning and characterization of EPI-1, the major protein in chimpanzee (Pan troglodytes) cauda epidydimal fluid. Biol Reprod 54: 857-864.         [ Links ]

FUSI FM, VIGNALI M, BUSACCA M, BRONSON RA (1992) Evidence for the presence of an integrin cell adhesion receptor on the oolema of unfertilized human oocytes. Mol Reprod Dev 31: 215-222.         [ Links ]

FUSI FM, MONTESANO M, BERNOCCHI N, FERRERA F, VILL A, BRONSON RA (1996) P-selectin is expressed on the oolema of human and hamster oocytes following sperm adhesion and is also detected on the equatorial segment of acrosome-reacted human spermatozoa. Mol Hum Reprod 2:341-347.         [ Links ]

GABRIELE A, D'ANDREA GD, CORDESCHI G, PROPERZI G, GIAMMATTEO M, DE STEFANO C, ROMANO R, FRANCAVILLA F, FRANCAVILLA S (1998) Carbohydrate binding activity in human spermatozoa: localization, specificity, and involvement in sperm-egg fusion. Mol Hum Reprod 543-553.         [ Links ]

GAUDREAULT C, LEGARE C, BERUBE B, SULLIVAN R (1999) Hamster sperm protein, P26h: a member of the short-chain dehydrogenase/resuctase superfamily. Biol Reprod 61: 264-273.         [ Links ]

GONZALEZ ECHEVERRIA MF, CUASNICU PS, PIAZZA A, PINEIRO L, BLAQUIER JA (1984) Addition of an androgen-free epididymal extract increases the ability of immature hamster spermatozoa to fertilize in vivo and in vitro. J Reprod Fertil 71: 433-437.         [ Links ]

HALE G, XIA MQ, TIGHE HP, DYER MJS, WALDMANN H (1990) The CAMPATH-1 antigen (CDw52). Tissue antigens 35: 118-127.         [ Links ]

HAMMAMI-HAMZA S, DOUSSAU M, BERNARD J, ROGIER E, DUQUENNE C, RICHARD Y, LEFEVRE A, FINAZ C. 2001. Cloning and sequencing of SOB3, a human gene coding for a sperm protein homologous to an antimicrobial protein and protentially involved in zona pellucida binding. Mol Human Reprod 7: 625-632.         [ Links ]

HAYASHI M, FUSHIMOTO S, TAKANO H, USHIKI T, ABE K, ISHIKURA H, YOSHIDA M, KIRCHOFF C, ISHIBASHI T, KASAHARA M (1996) Characterization of a human glycoprotein with potential role in sperm-egg fusion: cDNA cloning, immunohistochemical localization, and chromosomal assignment of the gene (AEGL1). Genomics 32: 367-374.         [ Links ]

HERMO L, WRIGHT J, OKO R, MORALES CR (1991) Role of epithelial cells of the male excurrent duct system of the rat in the endocytosis or secretion of sulfated glycoprotein-2 (clusterin). Biol Reprod 44: 1113-1131.         [ Links ]

HINRICHSEN M, BLAQUIER JA (1980) Evidence supporting the existence of sperm maturation in the human epididymis. J Reprod Fertil 60 291-294.         [ Links ]

KIRCHHOFF C (1998) Molecular characterization of epididymal proteins. Reviews Reprod 3: 86-95.         [ Links ]

KIRCHHOFF C (1999) Gene expression in the epididymis. Int Rev Cytol 188: 133-202.         [ Links ]

KIRCHHOFF C, HABBEN I, IVELL R, KRULL N (1992) A major human epididymis specific cDNA encodes a protein with sequence homology to extracellular proteinase inhibitors. Biol Reprod 45: 350-357.         [ Links ]

KIRCHHOFF C, HALE G (1996). Cell-to-cell transfer of glycosylphosphatidylinositol-anchored membrane proteins during sperm maturation. Mol Hum Reprod 2: 177-184.         [ Links ]

KIRCHHOFF C, KRULL N, PERA I, IVELLl R (1993). A major mRNA of the human epididymal procipal cells, HE5, encodes the leucocyte differentiation CDw52 antigen peptide backbone. Mol Reprod Dev 34: 8-15.

KIRCHHOFF C, OSTERHOFF C, HABBEN I, IVELL R (1990). Cloning and analysis of mRNAs expressed specifically in the human epididymis. Int J Androl 13: 155-167.         [ Links ]

KIRCHHOFF C, OSTERHOFF C, YOUNG L (1996) Molecular cloning and characterization of HE1, a major secretory protein on the human epididymis. Biol Reprod 54: 847-856.         [ Links ]

KIRCHHOFF C, PERA I, RUST W, IVELL R (1994) Major human epididymis-specific gene product, HE3, is the first representative of a novel gene family. Mol Rep Dev 37: 130-137.         [ Links ]

KOMORI S, KAMEDA K, SAKATA A, HASEGAWA A, TOJI H, TSUJI Y, SHIBAHARA H, KOYAMA K, ISOJIMA S (1997) Characterization of fertilization-blocking monoclonal antibody IG12with human sperm immobilizing activity. Clin Exp Immunol 109: 547-554.         [ Links ]

KRATZSCHMAR J, HAENDLER B, EBERSPAECHER U, ROOSTERMAN D, DONNER P, SCHLEUNING WD (1996). The human cysteine-rich secretory protein (CRISP) family. Primary structure and tissue distribution of CRISP-1, CRISP-2 and CRISP-3. Eur J Biochem 236: 827-836.         [ Links ]

LEA OA, PERTRUSZ P, FRENCH FS. (1978) Purification and localization of acidic epididymal glycoprotein (AEG): a sperm coating protein secreted by the rat epididymis. Int J of Androl Suppl 2: 592-607.         [ Links ]

LEFEVRE A, MARTIN RUIZ C, CHOKOMIAN S, DUQUENNE C, FINAZ C (1997) Characterization and isolation of SOB2, a human sperm protein with a potential role in oocyte membrane binding. Mol Human Reprod 3: 507-516.         [ Links ]

LEGARE C, GAUDREAULT C, ST-JACQUES S, SULLIVAN R (1999) P34H sperm protein is preferentially expressed by the human corpus epididymis. Endocrinology 140: 3318-3327.         [ Links ]

LILJA H, LAURELL C-B (1984) Liquefaction of coagulated human semen. Scan J Clin Lab Invest 44: 447-452.         [ Links ]

LILJA H, LAURELL C-B (1985) The predominant protein in human seminal coagulate. Scan J Clin Lab Invest 45: 635-641.         [ Links ]

LILJA H, LAURELL C-B (1992) Molecular cloning of epididymal and seminal vesicular transcripts encoding a semenogelin-related protein. Proc Nat Acad Sci USA 89: 4559-4563.         [ Links ]

LIU HW, LIN YC, CHAO CF, CHANG SY, SUN GH (2000) GP-83 and GP-39, two glycoproteins secreted by human epididymis are conjugated to spermatozoa during maturation. Mol Hum Reprod 6: 422-428.         [ Links ]

MARTIN RUIZ C, DUQUENNE C, TRETON D, LEFEVRE A, FINAZ C (1998) SOB3, a human sperm protein involved in zona pellucida binding: physiological and biochemical analysis, purification. Molec Reprod Dev 49: 286-297.         [ Links ]

MILLER DJ, WINER MA, AX RL (1990) Heparin-binding proteins from seminal plasma bind to bovine spermatozoa and modulate capacitation by heparin. Biol Reprod 42: 899-915.         [ Links ]

MIRANDA PV, BRANDELLI A, TEZON JG (1995) Characterization of ß-N-acetylglucosaminidase from human epididymis. Int J Androl 18: 263-270.         [ Links ]

MIRANDA PV, GONZALEZ-ECHEVERRÍA F, BLAQUIER JA, MAHURAN DJ, TEZON JG (2000) Evidence for the participation of ß-hexosaminidase in human sperm-zona pellucida interaction in vitro. Mol Hum Reprod 6: 699-706.         [ Links ]

MIRANDA PV, GONZALEZ-ECHEVERRIA F, MARIN-BRIGGILER CI, BRANDELLI A, BLAQUIER JA, TEZON JG (1997) Glycosydic residues involved in human sperm-zona pellucida binding in vitro. Mol Hum Reprod 3: 399-404.         [ Links ]

MIRANDA PV, TEZON JG (1992) Characterization of fibronectin as a marker for human epididymal sperm maturation. Mol Reprod Dev 33: 443-450.         [ Links ]

MOORE HD, HARTMAN T, PRYOR J (1983) Development of the oocyte-penetrating capacity of spermatozoa in the human epididymis. Int J Androl 6: 310-318.         [ Links ]

MOORE HDM (1981) Glycoprotein secretions of the epididymis in the rabbit and hamster: localization on epididymal spermatozoa and the effect of specific antibodies on fertilization in vivo. J Exp Zool 215: 77-85.         [ Links ]

MOORE HDM, CURRY MR, PENFOLD LM, PRYOR JP (1992) The culture of human epididymal epithelium and in vitro maturation of epididymal spermatozoa. Fertil Steril 58: 76-783.         [ Links ]

MOORE HDM, HARTMAN TD (1986) In vitro development of the fertilizing ability of hamster epididymal spermatozoa following co-culture with epithelium from proximal cauda epididymis. J Reprod Fertil 78: 347-352.         [ Links ]

MYLES DG, KIMMEL LH, BLOBEL CP, WHITE JM, PRIMAKOFF P (1994) Identification and binding sited in the desintegrin domain of fertilin required for sperm-egg fusion. Proc Natl Acad Sci USA 1: 4195-4198.         [ Links ]

O´BRYAN MK, MALLIDIS C, MURPHY BF, BAKER HWG (1994a) Immunohistological localization of clusterin in the male genital tract in humans and marmosets. Biol Reprod 50: 502-509.         [ Links ]

O´BRYAN MK, MURPHY BF, LIU DY, CLARKE GN, BAKER HWG (1994b) The use of anticlusterin monoclonal antibodies for the combined assessment of human sperm morphology and acrosome integrity. Human Reprod 9: 1490-1496.         [ Links ]

ORGEBIN CRIST MC, FOURNIER-DELPECH S (1982) Sperm-egg interaction. Evidence for maturational changes during epididymal transit. J Androl 3: 429-433.         [ Links ]

OSTERHOFF C, IVELL R, KIRCHHOFF C (1997) Cloning of a human epididymis-specific mRNA, HE6, encoding a novel member of the seven transmembrane-domain receptor superfamily. DNA and Cell Biol 16: 379-389.         [ Links ]

OSTERHOFF C, KIRCHHOFF C, KRULL N, IVELL R (1994) Molecular cloning and characterization of a novel human sperm antigen (HE2) specifically expressed in the proximal epididymis. Biol Reprod 50: 516-525.         [ Links ]

PARRY RV, BARKER PJ, JONES R (1992) Characterization of low Mr zona pellucida binding proteins from boar spermatozoa and seminal plasma. Mol Rep Dev 33: 108-115.         [ Links ]

PECHEUR EI, SAINTE-MARIE J, BIENVENUES A, HOCKSTRA D (1999) Peptides and membrane fusion: towards the understanding of molecular mechanism of protein-induced fusion. J Membr Biol 167: 1-17.         [ Links ]

PERA I, IVELL R, KIRCHHOFF C (1996) Body temperature (37°C) specifically down-regulates the mRNA for the major sperm surface antigen CD52 in epididymal cell culture. Endocrinology 137: 4451-4459.         [ Links ]

PEREZ MARTINEZ S, CONESA D, CUASNICU PS (1995) Potential contraceptive use of epididymal proteins: evidence for the participation of specific antibodies against rat epididymal protein DE in male and female fertility inhibition. J Reprod Immunol 29: 31-45.         [ Links ]

PERRY ACF, JONES R, HALL L (1995) The monkey ESP14.6 mRNA a novel transcript expressed at high levels in the epididymis. Gene 153: 291-291.         [ Links ]

ROBAIRE B, HERMO L (1988) Efferent ducts, epididymis, and vas deferens: structure, functios, and their regulation. In: Knobil E, Neill J (eds). The physiology of reproduction. New York: Raven Press, pp 999-1080.         [ Links ]

ROBAIRE B, VIGER RS (1995) Regulation of epididymal epithelial cell functions. Biol Reprod 52: 226-236.         [ Links ]

ROCHWERGER L, COHEN DJ, CUASNICU PS (1992) Mammalian sperm-egg fusion: the rat egg has complementary sites for a sperm protein that mediates gamete fusion. Dev Biol 153: 83-90.         [ Links ]

ROCHWERGER L, CUASNICU P, CONESA D (1990) Participation of a rat sperm epididymal glycoprotein in the acquisition of sperm-egg fusion ability during maturation. In: Alexander NJ, Griffin D, Spieler JM, Waites, GMH (ed). Gamete interaction. Prospects for immunocontraception. New York. John Wiley Liss & Sons, Inc. pp. 630-631.         [ Links ]

ROCHWERGER L, CUASNICU PS (1992). Reditribution of a rat sperm epididymal glycoprotein after in vivo and in vitro capacitation. Mol Reprod Dev 31: 34-41.         [ Links ]

ROSS P, KAN FWK, ANTAK P, VIGNEAULT N, CHAPDELAINE A, ROBERTS KD (1990) Protein synthesis and secretion in the human epididymis and immunoreactivity with sperm antibodies. Mol Reprod Dev 26: 16-23.         [ Links ]

RUFAS O, FISCH B, SHALGI R (2000) Expression of cadherin adhesion molecucles on human gametes. Molec Human Reprod 6: 163-169.         [ Links ]

SAALMANN A, MUNZ S, ELLERBROCK K, IVELL R, KIRCHHOFF C (2001) Novel sperm-binding proteins of epididymal origin contain four fibronectin type II-modules. Molec Reprod Dev 58: 88-100.         [ Links ]

SALOMON D, AYALON O, PATEL-KING R, HYNES RO, GEIGER B (1992) Extrajunctional distribution of N-cadherin in cultured human endothelial cells. J Cell Sci 102: 7-17.         [ Links ]

SCHALLER J, GLANDER HJ, DETHLOFF J (1993) Evidence of ß1 integrins and fibronectin on spermatogenic cells in human testis. Hum Reprod 8: 1873-1878.         [ Links ]

SNELL WJ, WHITE JM (1996) The molecules of mammalian fertilization. Cell 85:629-637.         [ Links ]

SUN GH, LIN YC, GUO YW, CHANG SY, LIU HW (2000) Purification of GP-83, a glycoprotein secreted by the human epididymis and conjugated to mature spermatozoa. Mol Hum Reprod 6: 429-434.         [ Links ]

SUTOVSKY P, MORENO R, RAMALHO-SANTOS J, DOMINKO T, SIMERLY C, SCHATTEN G. 2000. Ubiquitin-dependent mechanism for sperm quality control in mammalian epididymis. Biol Reprod, 62 (Suppl.1), 110.         [ Links ]

SUTOVSKY P, MORENO R, RAMALHO-SANTOS J, DOMINKO T, THOMPSON V, SCHATTEN G. 2001a. A putative, ubiquitin-dependent mechanism for the recognition of elimination of defective spermatozoa in the mammalian epididymis. J Cell Science 114: 1665-1675.         [ Links ]

SUTOVSKY P, TERADA Y, SCHATTEN G. 2001b. Ubiquitin-based sperm assay for the diagnosis of male factor infertility. Human Reprod 16: 250-258.         [ Links ]

SYLVESTER SR, MORALES C, OKO R, GRISWOLD MD (1991) Localization of sulfated glycoprotein-2 (clusterin) on spermatozoa and in the reproductive tract of the male rat. Biol Reprod 45:195-207.         [ Links ]

TAKEICHI M (1990) Cadherins: a molecular family important in selective cell-cell adhesion. Annu Rev Biochem 59: 237-252.         [ Links ]

TAKEICHI M (1995) Morphogenetic roles of classic cadherins. Curr Op Cell Biol 7: 619-627.         [ Links ]

TEZON JG and BLAQUIER JA (1981) The organ culture of human epididymal tubules and their response to androgens. Molec Cell Endocrinol 3: 233-242.         [ Links ]

TEZON JG, RAMELLA E, CAMEO M, VAZQUEZ MH, BLAQUIER JA (1985) Immunochemical localization of secretory antigens in the human epididymis and their association with spermatozoa. Biol Reprod 32: 591-597.         [ Links ]

TEZON JG, VAZQUEZ MH, LARMINAT M, PIÑEIRO L, PIAZZA A, SCORTICATTI C, BLAQUIER JA (1987) Further characterization of a model system for the study of human epididymal physiology and its relation to sperm maturation. Annual NY Acad Sci: 215-221.         [ Links ]

TEZON JG, VAZQUEZ MH, PIÑEIRO L, LARMINAT M, BLAQUIER JA (1985) Identification of androgen-induced proteins in human epididymis. Biol Reprod 32: 584-590.         [ Links ]

TOPFER-PETERSEN, E (1999) Molecules on the sperm´s route to fertilization. J Exp Zool 285: 259-266.         [ Links ]

TRASLER J, SABERI F, SOMANI IH, ADAMALI HI, HUANG JQ, FORTUNATO SR, RITTER G, AEBERSOLD R, GRAVEL RA, HERMO L (1998) Characterization of the testis and epididymis in mouse models of human Tay Sachs and Sandhoff diseases and partial determination of accumulated gangliosides. Endocrinology 139 (7): 3280-8.         [ Links ]

VAZQUEZ MH, LARMINAT M, BLAQUIER JA (1986a) Effect of androgens on androgen receptor in cultured human epididymis. J Endocrinol 111: 343-348.         [ Links ]

VAZQUEZ MH, DE LARMINAT M, SCORTICATI C, BLAQUIER JA (1989) The effect of in vitro androgen stimulation upon androgen metabolism and trophic parameters in cultured human epididymis. Andrologia 1: 9-17.         [ Links ]

VAZQUEZ MH, LARMINAT M, GURPIDE E, SCORTICATTI C, BLAQUIER JA (1986b) Effect of in vivo estrogen administration. Androgen metabolism in the human epididymis. J Steroid Biochem 25: 239-244.         [ Links ]

WASSARMAN PM (1999) Mammalian fertilization: molecular aspects of gamete adhesion, exocytosis, and fusion. Cell 96: 175-183.         [ Links ]

YANAGIMACHI, R. (1994) Mammalian fertilization. In: Knobil E., Neill JD (eds) The physiology of Reproduction. New York: Raven Press, 1994: 189-317.         [ Links ]

YEUNG CH, COOPER TG, NIESCHLAG E (1997). Human epididymal secreted protein CD52 on ejaculated spermatozoa: correlation with semen characteristics and the effect of its antibody. Mol Hum Reprod 3: 1045-1051         [ Links ]

Creative Commons License Todo o conteúdo deste periódico, exceto onde está identificado, está licenciado sob uma Licença Creative Commons