The molecular evolution of spermatogenesis across mammals.

The testis produces gametes through spermatogenesis and evolves rapidly at both the morphological and molecular level in mammals1-6, probably owing to the evolutionary pressure on males to be reproductively successful7. However, the molecular evolution of individual spermatogenic cell types across mammals remains largely uncharacterized. Here we report evolutionary analyses of single-nucleus transcriptome data for testes from 11 species that cover the three main mammalian lineages (eutherians, marsupials and monotremes) and birds (the evolutionary outgroup), and include seven primates. We find that the rapid evolution of the testis was driven by accelerated fixation rates of gene expression changes, amino acid substitutions and new genes in late spermatogenic stages, probably facilitated by reduced pleiotropic constraints, haploid selection and transcriptionally permissive chromatin. We identify temporal expression changes of individual genes across species and conserved expression programs controlling ancestral spermatogenic processes. Genes predominantly expressed in spermatogonia (germ cells fuelling spermatogenesis) and Sertoli (somatic support) cells accumulated on X chromosomes during evolution, presumably owing to male-beneficial selective forces. Further work identified transcriptomal differences between X- and Y-bearing spermatids and uncovered that meiotic sex-chromosome inactivation (MSCI) also occurs in monotremes and hence is common to mammalian sex-chromosome systems. Thus, the mechanism of meiotic silencing of unsynapsed chromatin, which underlies MSCI, is an ancestral mammalian feature. Our study illuminates the molecular evolution of spermatogenesis and associated selective forces, and provides a resource for investigating the biology of the testis across mammals.

PIWI-interacting RNAs and human testicular function.

Small noncoding RNAs (sncRNAs) are pieces of RNA with a length below 200 bp and represent a diverse group of RNAs having many different biological functions. The best described subtype is the microRNAs which primarily function in posttranscriptional gene regulation and appear essential for most physiological processes. Of particular interest for the germline is the PIWI-interacting RNAs (piRNAs) which are a class of sncRNA of 21-35 bp in length that are almost exclusively found in germ cells. Recently, it has become clear that piRNAs are essential for testicular function, and in this perspective, we outline the current knowledge of piRNAs in humans. Although piRNAs appear unique to germ cells, they have also been described in various somatic cancers and biofluids. Here, we discuss the potential function of piRNAs in somatic tissues and whether detection in biofluids may be used as a biomarker for testicular function. This article is categorized under: Reproductive System Diseases > Genetics/Genomics/Epigenetics Reproductive System Diseases > Molecular and Cellular Physiology.

Organophosphate ester flame retardants have antiandrogenic potential and affect other endocrine related endpoints in vitro and in silico.

Organophosphate ester flame retardants (OPFRs) are used to prevent ignition and spreading of fire. They are present in various human matrices suggesting adult, fetal, and neonate exposure. Endocrine related effects have been observed in vivo, but information at the molecular level is lacking for some OPFRs. Also, a better understanding of potential contribution from chemical substructures is needed. The aim of this study was to screen OPFRs for endocrine disruptive potential in vitro and in silico. We selected eleven substances to represent some OPFRs with 1) little information on endocrine activity and others to represent 2) varied chemical substructures. We used in vitro assays for androgen receptor (AR), aryl hydrocarbon receptor (AhR), and Nrf2 activity, effects on steroidogenesis, and transthyretin (TTR) binding, as well as in silico models covering estrogen, thyroid, and CYP3A4 induction related endpoints. Ten OPFRs affected AR and AhR activity, seven affected TTR binding, and five affected 17ß-estradiol levels. Several substances had IC50-values below 10 µM and exhibited efficacious effects. These included TPHP, CDP, TMPP, TIPPP, and EHDPP for AR antagonism, suggesting that the degree of arylation and the size of the substance can play a role for the activity. Chlorinated OPFRs had low/no effect on TTR binding. No clear trend was observed for AhR and steroidogenesis, but all arylated OPFRs were predicted to have alert for estrogen receptor binding in an in silico model with metabolism simulator included. Collectively, our data suggest that OPFRs have endocrine disruptive potential warranting further studies to enable human risk assessment.

FSHB and FSHR gene variants exert mild modulatory effect on reproductive hormone levels and testis size but not on semen quality: A study of 2020 men from the general Danish population.

BACKGROUND: Spermatogenesis depends on stimulation by follicle-stimulating hormone (FSH) which binds to FSH receptors (FSHR) on testicular Sertoli cells. Three FSH-related single-nucleotide polymorphisms (SNPs), FSHB -211G>T (rs10835638), FSHR -29G>A (rs1394205) and FSHR 2039A>G (rs6166) affect FSH action, and have been suggested to affect testicular function, but the evidence is uncertain. OBJECTIVE: To describe the associations between the three SNPs and testicular function in a large and well-characterised cohort of men from the general population. MATERIALS AND METHODS: A cross-sectional study of 2020 Danish men unselected regarding testicular function. Outcome variables were semen parameters, reproductive hormones and testis size. Genotyping was done by competitive allele-specific quantitative PCR. Differences in genotype frequencies were tested by chi-square test and associations between genotypes and outcomes were assessed by multivariate linear regressions. RESULTS: The SNPs affected serum FSH; carriers of the variant affecting FSH secretion (FSHB -211G>T) had lower FSH levels while carriers of variants affecting receptor expression (FSHR -29G>A) and receptor sensitivity (FSHR 2039A>G) had higher FSH levels. Carriers of FSHB -211G>T had lower calculated free testosterone/LH ratio. Although both FSHB -211G>T and FSHR 2039A>G were associated with smaller testis size, no clear association was detected in relation to any semen parameters, except a lower total number of morphologically normal spermatozoa in the heterozygous carriers of the FSHB -211G>T DISCUSSION AND CONCLUSION: The studied polymorphisms have only minor modulating influence on testis size and function in healthy men. We detected subtle effects of the three SNPs on FSH levels, but also effects of FSHB -211G>T on calculated free testosterone/LH ratio, compatible with altered Leydig cell function. Thus, the role of these FSH-related polymorphisms is complex and modest in men with normal testicular function, but the possible importance of FSH polymorphisms in men with impaired testicular function should be evaluated in future studies in more detail.

Intrauterine exposure to diethylhexyl phthalate disrupts gap junctions in the fetal rat testis.

Fetal exposure to certain phthalate esters can disrupt testis development in rodents and lead to male reproductive disorders, but with a causal link less certain in humans. Di(2-ethylhexyl) phthalate (DEHP) is one of the most common phthalates found in the environment and in rodents it is known to induce serious testis toxicity, as well as male reproductive disorders including cryptorchidism, hypospadias, impaired spermatogenesis and reduced fertility. In this study, we show that perinatal DEHP exposure disrupts gap junction localization in fetal and postnatal rat testis and correlate these findings to morphological changes. The protein Connexin 43 (CX43), normally expressed strongly in testicular gap junctions, was markedly downregulated in Leydig cells of DEHP-exposed fetal testes. In the postnatal testes, CX43 expression was recovered in the DEHP-exposed animals, even though Leydig cell clusters and malformed cords with intratubular Leydig cells were still present.

Transcriptome analysis of the adult human Klinefelter testis and cellularity-matched controls reveals disturbed differentiation of Sertoli- and Leydig cells.

The most common human sex chromosomal disorder is Klinefelter syndrome (KS; 47,XXY). Adult patients with KS display a diverse phenotype but are nearly always infertile, due to testicular degeneration at puberty. To identify mechanisms causing the selective destruction of the seminiferous epithelium, we performed RNA-sequencing of 24 fixed paraffin-embedded testicular tissue samples. Analysis of informative transcriptomes revealed 235 differentially expressed transcripts (DETs) in the adult KS testis showing enrichment of long non-coding RNAs, but surprisingly not of X-chromosomal transcripts. Comparison to 46,XY samples with complete spermatogenesis and Sertoli cell-only-syndrome allowed prediction of the cellular origin of 71 of the DETs. DACH2 and FAM9A were validated by immunohistochemistry and found to mark apparently undifferentiated somatic cell populations in the KS testes. Moreover, transcriptomes from fetal, pre-pubertal, and adult KS testes showed a limited overlap, indicating that different mechanisms are likely to operate at each developmental stage. Based on our data, we propose that testicular degeneration in men with KS is a consequence of germ cells loss initiated during early development in combination with disturbed maturation of Sertoli- and Leydig cells.