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Sperm cells undergo maturation during their transit throughout the epididymis. This process takes place in region-specific manner in which sperm are battered by proteins secreted by epithelium lining the epididymal duct. Most of the genes that encode for the proteins involved in the sperm maturation remain uncharacterized. Previous studies showed that family of β-defensins preferentially eaxpressed in male reproductive tracts and play an important role in both innate immunity and sperm fertility. In this study we characterized Defb20 to gain insight on its role in sperm maturation. Bioinformatic tools were used to analyzed functional domains and signal peptide. qRT-PCR analyses were used to analyzed tissue distribution, dependency on androgen and testicular factors and developmental-regulated expression analysis. Defb20 sequence contains important domains such as N-myristoilation and kinase binding sites which are putatively involved in the protein activation and protein-plasma membrane interaction. Moreover, DEFB20 contains a signal peptide indicating characteristic of secretory proteins. Defb20 was expressed exclusively in the epididymis with the highest expression in the caput region and was down-regulated by gonadectomy. Defb20 was also regulated by testicular factors in which the expression was down-regulated after efferent duct ligation (EDL). The dependency on the androgen was further confirmed by postnatal expression analysis in which Defb20 began to express at day-20 postnatal indicating specific stage of expression after initial development of the testis. In conclusion, Defb20 have a potential to be involved in the epididymal sperm maturation process.

Background: Sperm maturation takes place through contact between sperm and proteins produced in the epididymal lumen. CD52 had been characterised in the sperm; however, the expression and its regulation in the epididymis are mostly unknown. Aim: This study aimed to analyse the expression and regulation of CD52 in the mouse epididymis. Setting and Design: Experimental design was used in this study. Materials and Methods: Epididymis tissues from mice strain Deutch Democratic Yokohama were used as sources of total RNA. Bioinformatic tool was used to predict signal peptides. Quantitative real-time reverse transcription–polymerase chain reaction was used to analyse tissue distribution, androgen, testicular factors dependency and postnatal development. Statistical Analysis: One-way analysis of variance was used to analyse differences between treatment and control untreated group. P < 0.05 was determined as a significant difference. Results: CD52 amino acid sequence contains a signal peptide, indicating it is a secretory protein. CD52 exhibited region-specific expression in the epididymis, with the highest level being in the cauda. CD52 expression was regulated by androgen indicated by a significant downregulation at day 1 and day 3 following a castration (P < 0.05). Dependency on androgen was confirmed by injection of exogenous testosterone which prevented downregulation by 50%. Moreover, lumicrine factors also influenced CD52 expression indicated by ligation of efferent duct which also reduced expression at day 1 to day 5 following the ligation (P < 0.05). CD52 expression was developmentally regulated. This was shown by increase in the level of expression starting at day 15 postnatally. Conclusion: CD52 shows characteristics of genes involved in sperm maturation in the epididymis.

Androgen has an important role in regulation epididymal sperm maturation Epididymal sperm maturation is occurs via interactions between sperm and proteins secreted by epididymal genes that are specifically expressed in a region-specific manner. Mice CD52 exhibited expression in all regions in epididymis. However, the regulation of its expression by androgen is needed to be known as a gene candidate that involved in sperm maturation. Objective: This study was aimed to analyze regulation of mice CD52 expression by androgen and a bioinformatics analysis was performed to predict functional domains of CD52. Method s : bioinformatics analysis using Motif Scan. For experiment, mice divided into 5 groups. The groups were control was injected ethanol 0,1 mL/day and others groups were 3, 5, 10, and 15 day used a androgen receptor antagonist, flutamide that dilute in ethanol at a dose of 50 mg/kg/day. And then quantitative real-time RT-PCR was used to analyse expression of CD52 and it’s normalized to Beta actin. Result: CD52 contained a two potential phosphorylation sites for protein kinase C and casein kinase II, N-myristoylation, and N-glycosylation that showed CD52 is secretory protein and predict it’s attached to sperm membrane. Expression of CD52 was decrease about 93% than control. Conclusion: CD52 expression depend on androgen. Keywords : androgen; epididymis; sperm maturation; expression of CD52

Endocrine regulation of the mouse initial segment (IS) and distal caput epididymides was studied using genome-wide profiling of gene expression. Among the IS-enriched genes, 29% were significantly down-regulated 1 day after gonadectomy. Of those genes, dihydrotestosterone (DHT) supplementation was not sufficient to maintain their pregonadectomy level of expression in 70%. Of the caput-enriched genes, 16% were significantly down-regulated after gonadectomy, and of those genes, DHT supplementation did not maintain the initial level of expression in 28%. Identical data were obtained by clustering analyses performed for the expression data of epididymal genes. Furthermore, the microarray data revealed that 26 androgen receptor coregulators were expressed in the epididymis, of which several were confirmed by quantitative reverse transcriptase polymerase chain reaction analysis. This suggests putative involvement of these proteins in the segment-specific regulation of the epididymal genes. The pattern of epididymal gene expression in the novel proximal epididymis-specific androgen receptor knockout mouse ProxE-ARKO, with severe hypotrophy and hypoplasia of the caput epithelium, furthermore suggested that a subset of genes whose expression cannot be maintained by systemic androgen alone still require either direct lumicrine androgen action or a permissive effect of circulating testosterone. It is evident that testicular factors, one of which could be the high-concentration luminal androgen, are important for the expression of IS-enriched genes, whereas the expression of distal caput-enriched genes is typically regulated by systemic androgens.

We have performed genome-wide expression profiling of endocrine regulation of genes expressed in the mouse initial segment (IS) and distal caput of the epididymis by using Affymetrix microarrays. The data revealed that of the 15 020 genes expressed in the epididymis, 35% were enriched in one of the two regions studied, indicating that differential functions can be attributed to the IS and the more distal caput regions. The data, furthermore, showed that 27% of the genes expressed in the IS and/or distal caput epididymidis are under the regulation of testicular factors present in the duct fluid, while bloodborne androgens can regulate for 14% of them. This is in line with the high testis dependency of epididymal physiology. We then focused on genes with moderate or strong expression, showing strict segment enrichment and strong dependency on testicular factors. Analyses of the 59 genes, including upregulated and downregulated genes, fulfilling the criteria indicated that the expression of 18 (17 downregulated genes; 1 upregulated gene) of 19 gonadectomy-responsive genes enriched in the IS was not maintained by the androgen treatment, whereas the expression of all six downregulated genes enriched in the distal caput and the majority of those with no strict segment enrichment of expression (28 of 34; consisting of 23 downregulated and 5 upregulated genes) were maintained by androgens. Hence, it is evident that testicular factors other than androgens are important for the expression of IS-enriched genes, whereas the expression of distal caput-enriched genes is typically regulated by androgens. Identical data were obtained by independent clustering analyses performed for the expression data of 3626 epididymal genes. Several novel genes with putative involvement in epididymal sperm maturation, such as a disintegrin and metallopeptidase domain 28 (Adam28) and a solute carrier organic anion transporter family, member 4C1 (Slco4c1), were identified, indicating that this approach is successful for identifying novel epididymal genes.

Background Epididymal sperm maturation occurs via interactions between sperm and proteins secreted by the epididymal epithelium. Although this is an important process, the genes that encode the involved proteins remain largely uncharacterized. Previous studies have demonstrated that the genes involved in sperm maturation are regulated by androgen. Spag11a is an epididymal gene that is influenced by androgen. However, little is known about the putative role of this gene in the sperm maturation process. The objective of this study was to characterize Spag11a in the mouse epididymis. Methods In silico analyses were performed to predict signal peptides and functional domains. Spag11a expression was measured by quantitative real-time RT-PCR. Western blots and immunocytochemistry were performed to determine protein expression. Results SPAG11A is a member of the beta defensin protein family and constitutes a secretory protein. Spag11a was expressed exclusively in the epididymis. Moreover, it exhibited region-specific expression in the caput, which is typical for genes that are involved in creating a suitable microenvironment for sperm maturation. Mouse Spag11a was regulated by androgen. A significant decrease of Spag11a expression was observed at third day following a gonadectomy (P