Potential Role In Spermatogenesis Research Articles

Whole exome sequencing analysis of 167 men with primary infertility

BackgroundSpermatogenic failure is one of the leading causes of male infertility and its genetic etiology has not yet been fully understood.MethodsThe study screened a cohort of patients (n = 167) with primary male infertility in contrast to 210 normally fertile men using whole exome sequencing (WES). The expression analysis of the candidate genes based on public single cell sequencing data was performed using the R language Seurat package.ResultsNo pathogenic copy number variations (CNVs) related to male infertility were identified using the the GATK-gCNV tool. Accordingly, variants of 17 known causative (five X-linked and twelve autosomal) genes, including ACTRT1, ADAD2, AR, BCORL1, CFAP47, CFAP54, DNAH17, DNAH6, DNAH7, DNAH8, DNAH9, FSIP2, MSH4, SLC9C1, TDRD9, TTC21A, and WNK3, were identified in 23 patients. Variants of 12 candidate (seven X-linked and five autosomal) genes were identified, among which CHTF18, DDB1, DNAH12, FANCB, GALNT3, OPHN1, SCML2, UPF3A, and ZMYM3 had altered fertility and semen characteristics in previously described knockout mouse models, whereas MAGEC1,RBMXL3, and ZNF185 were recurrently detected in patients with male factor infertility. The human testis single cell-sequencing database reveals that CHTF18, DDB1 and MAGEC1 are preferentially expressed in spermatogonial stem cells. DNAH12 and GALNT3 are found primarily in spermatocytes and early spermatids. UPF3A is present at a high level throughout spermatogenesis except in elongating spermatids. The testicular expression profiles of these candidate genes underlie their potential roles in spermatogenesis and the pathogenesis of male infertility.ConclusionWES is an effective tool in the genetic diagnosis of primary male infertility. Our findings provide useful information on precise treatment, genetic counseling, and birth defect prevention for male factor infertility.

Brain-derived neurotrophic factor and neurotrophic tyrosine receptor kinase-2 in Stallion Testes: Insights into Seasonal Changes and Potential Roles in Spermatogenesis

Brain-derived neurotrophic factor and neurotrophic tyrosine receptor kinase-2 in Stallion Testes: Insights into Seasonal Changes and Potential Roles in Spermatogenesis

The rapidly evolving X-linked MIR-506 family fine-tunes spermatogenesis to enhance sperm competition.

Despite rapid evolution across eutherian mammals, the X-linked MIR-506 family miRNAs are located in a region flanked by two highly conserved protein-coding genes (SLITRK2 and FMR1) on the X chromosome. Intriguingly, these miRNAs are predominantly expressed in the testis, suggesting a potential role in spermatogenesis and male fertility. Here, we report that the X-linked MIR-506 family miRNAs were derived from the MER91C DNA transposons. Selective inactivation of individual miRNAs or clusters caused no discernible defects, but simultaneous ablation of five clusters containing 19 members of the MIR-506 family led to reduced male fertility in mice. Despite normal sperm counts, motility, and morphology, the KO sperm were less competitive than wild-type sperm when subjected to a polyandrous mating scheme. Transcriptomic and bioinformatic analyses revealed that these X-linked MIR-506 family miRNAs, in addition to targeting a set of conserved genes, have more targets that are critical for spermatogenesis and embryonic development during evolution. Our data suggest that the MIR-506 family miRNAs function to enhance sperm competitiveness and reproductive fitness of the male by finetuning gene expression during spermatogenesis.

The rapidly evolving X-linked MIR-506 family fine-tunes spermatogenesis to enhance sperm competition

Despite rapid evolution across eutherian mammals, the X-linked MIR-506 family miRNAs are located in a region flanked by two highly conserved protein-coding genes (SLITRK2 and FMR1) on the X chromosome. Intriguingly, these miRNAs are predominantly expressed in the testis, suggesting a potential role in spermatogenesis and male fertility. Here, we report that the X-linked MIR-506 family miRNAs were derived from the MER91C DNA transposons. Selective inactivation of individual miRNAs or clusters caused no discernible defects, but simultaneous ablation of five clusters containing 19 members of the MIR-506 family led to reduced male fertility in mice. Despite normal sperm counts, motility, and morphology, the KO sperm were less competitive than wild-type sperm when subjected to a polyandrous mating scheme. Transcriptomic and bioinformatic analyses revealed that these X-linked MIR-506 family miRNAs, in addition to targeting a set of conserved genes, have more targets that are critical for spermatogenesis and embryonic development during evolution. Our data suggest that the MIR-506 family miRNAs function to enhance sperm competitiveness and reproductive fitness of the male by finetuning gene expression during spermatogenesis.

Transcriptome Analysis Reveals Spermatogenesis-Related CircRNAs and LncRNAs in Goat Spermatozoa.

Mammalian spermatozoa comprises both coding and non-coding RNAs, which are traditionally believed to be a residual of spermatogenesis. The differential expression level of spermatozoal RNAs is also observed between fertile and infertile human, thereby anticipated as potential molecular marker of male fertility. This study investigated the transcriptome profile of goat (Capra hircus) spermatozoa. The sperm transcriptome was analyzed by three different methods viz. RLM-RACE, long-read RNA sequencing (RNAseq) in Nanopore™ platform, and short-read RNAseq in Illumina™ platform. The Illumina™ sequencing discovered 16,604 transcripts with 357 mRNAs having FPKM (fragments per kilobase per million mapped reads) of more than five. The spermatozoal RNA suite included mRNA (94%), rRNA (3%), miscRNA (1%), circRNA (1%), miRNA (1%), etc. This study also predicted circRNAs (127), lncRNAs (655), and imprinted genes (160) that have potential role in male reproduction. The gene ontology analysis revealed the involvement of spermatozoal RNA in regulating male meiosis (TET3, STAT5B), capacitation (ACRBP, CATSPER4), sperm motility (GAS8, TEKT2), spermatogenesis (ADAMTS2, CREB3L4), etc. The spermatozoal RNA were also associated with different biological pathways viz. Wnt signaling pathway, cAMP signaling pathway, AMPK signaling pathway, and MAPK signaling pathways having potential role in spermatogenesis. Overall, this study enlightened the suite of spRNA transcripts in goat and their relevance in male fertility for diagnostic approach.

Expression Analysis of IZUMO1 Gene during Testicular Development of Datong Yak (Bos Grunniens).

Simple SummaryIZUMO1 (IZUMO sperm-egg fusion) is a crucial member of the immunoglobulin superfamily, and it plays a vital part in egg sperm interaction, fusion, and in spermatogenesis. Previous studies identified the fundamental role of the IZUMO1 gene in meiosis of bovine spermatogenesis and spermatogenic cell development and fertility. The current study confirmed the significant expression of the IZUMO1 gene with testicular development and its potential role in fertilization of Datong yak. This research explained the biological role of IZUMO1 in yak reproduction; therefore in the future it might provide a novel prospect to apply data on reproduction and to understand the further regulatory mechanisms of the IZUMO1 gene during spermatogenesis of different mammalian species.The IZUMO1 gene has promising benefits for the national development of novel non-hormonal contraceptives and in the treatments of fertility. Understanding the function of IZUMO1, its mRNA, and protein expression is critical to gain insight into spermatogenesis and promote sperm-egg fusion during reproduction of Datong yak. Therefore, we estimated the IZUMO1 gene expression in different ages of Datong yak by using semi quantitative PCR, qPCR, and western blotting. The results of the qPCR, semi-quantitative PCR and western blotting revealed that the expression level of IZUMO1 mRNA was significantly (p < 0.05) higher in the testis of 30 months and 6 years old followed by 18 and 6 months old Datong yak, respectively. We also predicted secondary and tertiary protein structure of IZUMO1 by using bioinformatics software that the revealed presence of a signal peptide, Izumo domain, immunoglobulin (Ig) like domain, and transmembrane region. Moreover, immunostaining analysis also elucidated that IZUMO1 was more prominent in the testis of 30 months and 6 years old yak, which represented that the IZUMO1 gene expression might be higher during the peak breeding ages (6 to 7 years) of the yak, and play a potential role in spermatogenesis, fertility, and testicular development.

LncRNA4667 is Dispensable for Spermatogenesis and Fertility in Mice

Objective: Spermatogenesis is a complex process which is of vital importance for sexual reproduction. In many studies of spermatogenesis, the mRNAs, protein-coding genes, as well as small noncoding RNAs (ncRNAs) have been well characterized. However, there remain numerous questions despite previously characterized molecular mechanisms. Long ncRNAs (lncRNAs) are a relatively new addition to our knowledge of ncRNAs. Limited studies have examined the function of lncRNAs in spermatogenesis. Therefore, we selected a testis-specific lncRNA, lncRNA4667, to analyze its potential role in spermatogenesis and male fertility. Methods: In situ hybridization and quantitative reverse transcription polymerase chain reaction analyses were used to confirm testis-specific expression of lncRNA4667. LncRNA4667 knockout mice were generated using CRISPR/Cas9 technology. Histology, sperm counts, sperm motility, body parameters, and fertility were compared between wild-type and knockout mice (n = 8/group). Results: Expression analysis showed that lncRNA4667 was testis specific and localized to round spermatids in seminiferous tubules of adult mouse testes. Mice homozygous for a null mutation of lncRNA4667 displayed normal spermatogenesis and fertility compared with wild-type mice. Conclusions: These data indicate that lncRNA4667 is dispensable for spermatogenesis and fertility in mice, and the localization of lncRNA4667 makes it a useful marker for the identification of round spermatids in mice.

Age-associated expression of vitamin D receptor and vitamin D-metabolizing enzymes in the male reproductive tract and sperm of Hu sheep

The cellular response to 1,25-dihydroxyvitamin D3 (Vit D3; biologically active form of Vitamin D) is complex and depends not only on Vitamin D receptor (VDR) expression but also on cellular uptake of circulating Vit D3 and the presence and activity of Vitamin D-metabolizing enzyme. This study evaluated the expression of VDR and Vitamin D-metabolizing enzymes in the ram reproductive tract at different developmental stages and in spermatozoa. Nearly all cell types in the testes and epithelial cells of the caput, corpus, and cauda expressed VDR, CYP27B1, and CYP24A1 proteins. The mRNA and protein expression of CYP2R1, CYP27A1, and CYP27B1 in the testes and cauda increased significantly with increasing age (P < 0.05). However, epididymal VDR mRNA and protein expression showed no significant difference (P < 0.05) between adult (9- and 24-month-old) and prepubertal (3-month-old) rams. Furthermore, VDR and CYP24A1 were mainly concentrated in the mid-piece of ejaculated or cauda epididymis spermatozoa or both. Additionally, VDR and CYP27B1 mRNA and protein expression levels were significantly higher in ejaculated spermatozoa than in cauda epididymal spermatozoa (P < 0.05). Moreover, VDR and CYP24A1 expression was significantly higher in high-motility than in low-motility spermatozoa (P < 0.05). The diverse expression patterns of VDR and Vitamin D-metabolizing enzymes in the ram reproductive tract at different developmental stages and spermatozoa suggest it plays a potential role in spermatogenesis.

Dynamic expression of long noncoding RNAs reveals their potential roles in spermatogenesis and fertility.

Mammalian reproduction requires that males and females produce functional haploid germ cells through complex cellular differentiation processes known as spermatogenesis and oogenesis, respectively. While numerous studies have functionally characterized protein-coding genes and small noncoding RNAs (microRNAs and piRNAs) that are essential for gametogenesis, the roles of regulatory long noncoding RNAs (lncRNAs) are yet to be fully characterized. Previously, we and others have demonstrated that intergenic regions of the mammalian genome encode thousands of long noncoding RNAs, and many studies have now demonstrated their critical roles in key biological processes. Thus, we postulated that some lncRNAs may also impact mammalian spermatogenesis and fertility. In this study, we identified a dynamic expression pattern of lncRNAs during murine spermatogenesis. Importantly, we identified a subset of lncRNAs and very few mRNAs that appear to escape meiotic sex chromosome inactivation, an epigenetic process that leads to the silencing of the X- and Y-chromosomes at the pachytene stage of meiosis. Further, some of these lncRNAs and mRNAs show a strong testis expression pattern suggesting that they may play key roles in spermatogenesis. Lastly, we generated a mouse knockout of one X-linked lncRNA, Tslrn1 (testis-specific long noncoding RNA 1), and found that males carrying a Tslrn1 deletion displayed normal fertility but a significant reduction in spermatozoa. Our findings demonstrate that dysregulation of specific mammalian lncRNAs is a novel mechanism of low sperm count or infertility, thus potentially providing new biomarkers and therapeutic strategies.

Association of G/T(rs222859) polymorphism in Exon 1 of YBX2 gene with azoospermia, among Iranian infertile males.

Animal model studies have shown that MSY2 gene has a potential role in spermatogenesis. Some mutations on this gene have been proposed to be associated with human male infertility. In this study, polymorphisms of exon 1 of YBX2 gene have been investigated. A total of 276 men were evaluated. They included 96 men with normal spermatogenesis, 60 men with nonobstructive azoospermia, 60 men with oligospermia and 60 men with asthenospermia. We extracted DNA from blood and testis tissues of samples, and analysed polymorphisms of exon 1 by sequencing method. Moreover, YBX2 gene expression was studied by real-time PCR on blood and testis tissue of samples. Sequencing results showed that among the studied polymorphisms, frequency of TT genotype in rs222859 polymorphism was significantly higher in azoospermic patients compared to control group (P<0.001). Azoospermic men exhibited significant underexpression of YBX2 gene in blood and testis samples in comparison with controls, oligosperm and asthenosperm samples (P<0.001), but there was no significant difference in gene expression of YBX2 gene in blood and testis tissues of azoospermic men, with and without mutation (P>0.05). According to our results, the alterations of this gene might be involved in azoospermia among Iranian population.

MicroRNA profiling in three main stages during porcine spermatogenesis.

Spermatogenesis is an intricate biological event wherein an undifferentiated spermatogonium develops into mature sperms. MicroRNAs are a type of single strand small non-coding RNA molecule and are implicated in the regulation of many crucial pathways during cell proliferation, apoptosis, and differentiation. Here, we present a comprehensive comparison of miRNA expression profiling in three main stages during porcine spermatogenesis using high-throughput sequencing. We built three small RNA libraries for the testis, the epididymis and the ejaculated sperm from a Landrace boar, and in total obtained 3821 precursor hairpins encoding for 4761 mature miRNAs, of which 23 are miRNA*. Notably, 940 precursor miRNAs produced both the 5'- and 3'- strands as sister pairs, indicating the distinctive expression patterns of germ cell miRNAs. Additionally, 418 out of 710 co-expressed miRNAs were identified as being differentially expressed between libraries (P < 0.001). Apart from the sexual specific X chromosome, many miRNAs were found to be located on chromosome 12, which may play potential roles in spermatogenesis according to the result of synteny analysis with human and mouse. The Gene Ontology and KEGG pathway analysis revealed that the target genes of co-expressed miRNAs were highly involved in the cell cycle process, metal ion binding, modification of plasma membrane, and the p53 signal pathway.

X chromosome-linked CNVs in male infertility: discovery of overall duplication load and recurrent, patient-specific gains with potential clinical relevance.

IntroductionSpermatogenesis is a highly complex process involving several thousand genes, only a minority of which have been studied in infertile men. In a previous study, we identified a number of Copy Number Variants (CNVs) by high-resolution array-Comparative Genomic Hybridization (a-CGH) analysis of the X chromosome, including 16 patient-specific X chromosome-linked gains. Of these, five gains (DUP1A, DUP5, DUP20, DUP26 and DUP40) were selected for further analysis to evaluate their clinical significance.Materials and MethodsThe copy number state of the five selected loci was analyzed by quantitative-PCR on a total of 276 idiopathic infertile patients and 327 controls in a conventional case-control setting (199 subjects belonged to the previous a-CGH study). For one interesting locus (intersecting DUP1A) additional 338 subjects were analyzed.Results and DiscussionAll gains were confirmed as patient-specific and the difference in duplication load between patients and controls is significant (p = 1.65×10−4). Two of the CNVs are private variants, whereas 3 are found recurrently in patients and none of the controls. These CNVs include, or are in close proximity to, genes with testis-specific expression. DUP1A, mapping to the PAR1, is found at the highest frequency (1.4%) that was significantly different from controls (0%) (p = 0.047 after Bonferroni correction). Two mechanisms are proposed by which DUP1A may cause spermatogenic failure: i) by affecting the correct regulation of a gene with potential role in spermatogenesis; ii) by disturbing recombination between PAR1 regions during meiosis. This study allowed the identification of novel spermatogenesis candidate genes linked to the 5 CNVs and the discovery of the first recurrent, X-linked gain with potential clinical relevance.

Characterization of Cherax quadricarinatus prohibitin and its potential role in spermatogenesis

Prohibitin (PHB) proteins have diverse functions, such as cellular signaling, transcriptional control and mitochondrial biogenesis. In this study, we characterized PHB gene and its protein expression in Cherax quadricarinatus. PHB cDNA comprises 1472 nucleotides with an open reading frame of 828bp, which encodes 275 amino acid residues. The highest transcript levels were found during the spermatogonial developmental phase, with the lowest levels detected during the resting phase in the reproductive cycle. Western blot analysis revealed that PHB is an approximately 30kDa protein, and occurs in a number of unexpected isoforms, ranging from 30kDa to greater than 180kDa in the testes of different developmental phases, which may be the ubiquitinated substrates. The strongest immunolabeling signal was found in spermatogonia, with lower levels of staining in secondary spermatocytes, and weak or absent expression in mature sperm. Immunogold electron microscopy results confirmed the localization of PHB in the inner mitochondrial membranes. The results showed that PHB is a substrate protein for spermatogenesis, with a potential reproductive function involving sperm ubiquitination in invertebrates.

Structural variation of the human genome: mechanisms, assays, and role in male infertility

Genomic disorders are defined as diseases caused by rearrangements of the genome incited by a genomic architecture that conveys instability. Y-chromosome related dysfunctions such as male infertility are frequently associated with gross DNA rearrangements resulting from its peculiar genomic architecture. The Y-chromosome has evolved into a highly specialized chromosome to perform male functions, mainly spermatogenesis. Direct and inverted repeats, some of them palindromes with highly identical nucleotide sequences that can form DNA cruciform structures, characterize the genomic structure of the Y-chromosome long arm. Some particular Y chromosome genomic deletions can cause spermatogenic failure likely because of removal of one or more transcriptional units with a potential role in spermatogenesis. We describe mechanisms underlying the formation of human genomic rearrangements on autosomes and review Y-chromosome deletions associated with male infertility.

Gene discovery and comparative analysis of X-degenerate genes from the domestic cat Y chromosome

Mammalian sex chromosomes are the remnants of an ancient autosomal pair present in the ancestral mammalian karyotype. As a consequence of random decay and chromosome rearrangements over evolutionary time, Y chromosome gene repertoires differ between eutherian lineages. To investigate the gene repertoire and transcriptional analysis of the domestic cat Y chromosome, and their potential roles in spermatogenesis, we obtained full-length cDNA sequences for all known Y genes and their X chromosome gametologues and used those sequences to create a BAC-based physical map of the X-degenerate region. Our results indicate the domestic cat Y chromosome has retained most X-degenerate genes that were present on the ancestral eutherian Y chromosome. Transcriptional analysis revealed that most feline X-degenerate genes have retained housekeeping functions shared by their X chromosome partners and have not been specialized for testis-specific functions. Physical mapping data indicate that the cat SRY gene is present as multiple functional copies and that very little of the felid Y chromosome may be single copy. X-Y gene divergence time estimates obtained using Bayesian methods confirm an early origin of Stratum 1 genes prior to the origin of therian mammals. We observed no statistical difference in the ages of Stratum 2 and Stratum 3 gene pairs, suggesting that eutherian and marsupial Stratum 2 genes may have been independently retained in each lineage.

Sperm-associated antigen 1 is expressed early in pancreatic tumorigenesis and promotes motility of cancer cells

Sperm-associated antigen 1 (SPAG1) was recently identified in a rare form of infertility where anti-SPAG1 antibodies derived from the serum of an infertile woman were reported to cause sperm agglutination. Except for its expression and potential role in spermatogenesis, the function of SPAG1 is completely unknown. The unexpected finding of high levels of SPAG1 expression in pancreatic adenocarcinoma compared to normal pancreatic tissue in our previous cDNA array experiments prompted us to look in more detail at the expression and role of this gene in a panel of normal and malignant human tissues as well as in a larger series of pancreatic cancer specimens. We have generated an SPAG1-specific monoclonal antibody and showed high levels of SPAG1 protein in testis and in a large proportion of pancreatic ductal adenocarcinomas (PDAC). In the latter, SPAG1 expression was predominantly cytoplasmic and confined to malignant cells. Furthermore, the extent and intensity of SPAG1 expression was shown to be associated with stage and tumour nodal status, while analysis of precursor lesions, pancreatic intraepithelial neoplasias (PanINs), demonstrated its increased immunoreactivity with increasing PanIN grade, suggesting that SPAG1 is a novel marker of PDAC progression. Immunocytochemical analysis demonstrated colocalization of SPAG1 with microtubules, and their association was confirmed by co-immunoprecipitation; subsequent motility assays further substantiated a potential role of SPAG1 in cancer cell motility. Combined with the finding of its early expression in PDAC development, our data suggest that SPAG1 could contribute to the early spread and poor prognosis of pancreatic adenocarcinoma.

A spermatogenesis-related gene expression profile in human spermatozoa and its potential clinical applications.

Spermatogenesis is an essential stage in the human reproductive process. In a previously study aiming to determine which genes might be involved in spermatogenesis, we compared the gene expression profiles of adult and fetal testes by hybridizing cDNA probes prepared from adult and fetal testes to membranes dotted with gene clones derived from a commercial human testis library. We identified 266 differentially expressed genes that showed higher expression levels in adult testes, indicating their potential roles in spermatogenesis. In the present study, we applied the same cDNA microarray technique to the analysis of gene expression in the spermatozoa of normal fertile men and found 149 genes that were expressed at higher levels in adult testis. A further study of five sperm motility-related genes selected from this profile by real-time PCR revealed that there was significant difference in the expression levels of two genes ( TPX-1, testis-specific protein 1 and LDHC, lactate dehydrogenase C, transcript variant 1) between normal ( n=29) and motility impaired ( n=24) semen samples, indicating that these genes are involved in sperm function. Our results demonstrated that spermatogenesis-related gene profiling could help to assess sperm quality in humans, and further study of these genes will help us to elucidate the mechanisms involved in spermatogenesis and diseases relating to human infertility.

Screening of the SPO11 and EIF5A2 genes in a population of azoospermic and severely oligozoospermic men

Screening of the SPO11 and EIF5A2 genes in a population of azoospermic and severely oligozoospermic men

Suppression of Estrogen Receptor-mediated Transcription and Cell Growth by Interaction with TR2 Orphan Receptor

The transcriptional activity of the estrogen receptor (ER) is known to be highly modulated by the character and amount of coregulator proteins present in the cells. TR2 orphan receptor (TR2), a member of the nuclear receptor superfamily without identified ligands, is found to be expressed in the breast cancer cell lines and to function as a repressor to suppress ER-mediated transcriptional activity. Utilizing an interaction blocker, ER-6 (amino acids 312-340), responsible for TR2 interaction, the suppression of ER by TR2 could be reversed, suggesting that this suppression is conferred by the direct protein-protein interaction. Administration of antisense TR2, resulting in an enhancement of ER transcriptional activity in MCF7 cells, indicates that endogenous TR2 normally suppresses ER-mediated signaling. To gain insights into the molecular mechanism by which TR2 suppresses ER, we found that TR2 could interrupt ER DNA binding via formation of an ER-TR2 heterodimer that disrupted the ER homodimerization. The suppression of ER transcription by TR2 consequently caused the inhibition of estrogen-induced cell growth and G(1)/S transition in estrogen-dependent breast cancer cells. Taken together in addition to the potential roles in spermatogenesis and neurogenesis, our data provide a novel biological function of TR2 that may exert an important repressor in regulating ER activity in mammary glands.

Identification of four novel ADAMs with potential roles in spermatogenesis and fertilization.

The ADAM (A Disintegrin And Metalloprotease) family is known to have important roles in various developmental systems, e.g., myogenesis and neurogenesis. In this study, we searched for ADAMs that may function in spermatogenesis or fertilization, and have cloned and sequenced four new mouse ADAM cDNAs: ADAM 24, ADAM 25, ADAM 26 and ADAM 27. The deduced amino acid sequences show that all four contain the complete domain organization common to ADAM family members. Messenger RNA for each of the four ADAMs was found only in the testis. The conserved zinc-dependent metalloprotease active site HEXGHXXGXXHD was found in the metalloprotease domain of three of the novel ADAMs, suggesting that they are testis-specific proteases, to which we give the alternative names: testase 1, ADAM 24; testase 2, ADAM 25; and testase 3, ADAM 26. Using RNA extracted from testes of pre-pubertal males of increasing age (8-40days), we found that adult levels of transcription, assessed in Northern blots, are reached by day 20 (ADAM 27), day 25 (ADAMs 24 and 25) and in the range day 25-50 (ADAM 26). These results suggest that each ADAM is transcribed in spermatogenic cells in a regulated pattern at a specific developmental stage.