Explore the vast range of titles available.

PIWI-interacting RNAs (piRNAs) are a class of small non-coding RNAs that associate with proteins of the PIWI clade of the Argonaute family. First identified in animal germ line cells, piRNAs have essential roles in germ line development. The first function of PIWI–piRNA complexes to be described was the silencing of transposable elements, which is crucial for maintaining the integrity of the germ line genome. Later studies provided new insights into the functions of PIWI–piRNA complexes by demonstrating that they regulate protein-coding genes. Recent studies of piRNA biology, including in new model organisms such as golden hamsters, have deepened our understanding of both piRNA biogenesis and piRNA function. In this Review, we discuss the most recent advances in our understanding of piRNA biogenesis, the molecular mechanisms of piRNA function and the emerging roles of piRNAs in germ line development mainly in flies and mice, and in infertility, cancer and neurological diseases in humans.PIWI-interacting RNAs (piRNAs) are small non-coding RNAs with essential roles in germ line development through silencing of transposable elements and in regulation of protein-coding genes. Recent studies have deepened our understanding of the biogenesis and function of piRNAs and their roles in infertility, cancer and neurological diseases in humans.

Membraneless compartments, also known as condensates, provide chemically distinct environments and thus spatially organize the cell. A well-studied example of condensates is P granules in the roundworm Caenorhabditis elegans that play an important role in the development of the germline. P granules are RNA-rich protein condensates that share the key properties of liquid droplets such as a spherical shape, the ability to fuse, and fast diffusion of their molecular components. An outstanding question is to what extent phase separation at thermodynamic equilibrium is appropriate to describe the formation of condensates in an active cellular environment. To address this question, we investigate the response of P granule condensates in living cells to temperature changes. We observe that P granules dissolve upon increasing the temperature and recondense upon lowering the temperature in a reversible manner. Strikingly, this temperature response can be captured by in vivo phase diagrams that are well described by a Flory–Huggins model at thermodynamic equilibrium. This finding is surprising due to active processes in a living cell. To address the impact of such active processes on intracellular phase separation, we discuss temperature heterogeneities. We show that, for typical estimates of the density of active processes, temperature represents a well-defined variable and that mesoscopic volume elements are at local thermodynamic equilibrium. Our findings provide strong evidence that P granule assembly and disassembly are governed by phase separation based on local thermal equilibria where the nonequilibrium nature of the cytoplasm is manifested on larger scales.

Intracranial germ cell tumors (iGCTs) are rare, histologically classified as extragonadal germ cell tumors, and predominantly affect children and adolescents. These tumors are commonly found in the midline of the brain, with a notable male predominance and varying geographic incidences. The origins of iGCTs have been debated, particularly whether they arise from abnormal migration of primordial germ cells (PGCs) or transformed embryonic stem cells (ESCs). In this study, we hypothesize the potential presence of PGC-like cells in the pituitary gland, and these cells may differentiate into iGCTs during the complex and frequent regulation of the hypothalamic-pituitary–gonadal axis (HPGA).We analyzed the expression of four germ cell markers—MVH, OCT4, C-kit, and PLZF—using immunohistochemistry, Western blotting, and real-time quantitative PCR in human pituitary tissues, pituitary tumors, and pituitary germ cell tumors. Our findings indicate significant expression of these markers in pituitary tissues, with the highest levels found in pituitary germ cell tumors. These results support the existence of PGC-like cells that within the normal pituitary gland. This study provides new insights into the cellular origins of iGCTs and suggests further investigation into the regulatory mechanisms that may lead to tumorigenesis.

The Streptococcus‐derived CRISPR/Cas9 system is being widely used to perform targeted gene modifications in plants. This customized endonuclease system has two components, the single‐guide RNA (sgRNA) for target DNA recognition and the CRISPR‐associated protein 9 (Cas9) for DNA cleavage. Ubiquitously expressed CRISPR/Cas9 systems (UC) generate targeted gene modifications with high efficiency but only those produced in reproductive cells are transmitted to the next generation. We report the design and characterization of a germ‐line‐specific Cas9 system (GSC) for Arabidopsis gene modification in male gametocytes, constructed using a SPOROCYTELESS (SPL) genomic expression cassette. Four loci in two endogenous genes were targeted by both systems for comparative analysis. Mutations generated by the GSC system were rare in T1 plants but were abundant (30%) in the T2 generation. The vast majority (70%) of the T2 mutant population generated using the UC system were chimeras while the newly developed GSC system produced only 29% chimeras, with 70% of the T2 mutants being heterozygous. Analysis of two loci in the T2 population showed that the abundance of heritable gene mutations was 37% higher in the GSC system compared to the UC system and the level of polymorphism of the mutations was also dramatically increased with the GSC system. Two additional systems based on germ‐line‐specific promoters (pDD45‐GT and pLAT52‐GT) were also tested, and one of them was capable of generating heritable homozygous T1 mutant plants. Our results suggest that future application of the described GSC system will facilitate the screening for targeted gene modifications, especially lethal mutations in the T2 population.

Germ cells (GCs) serve as indispensable carriers in both animals and plants, ensuring genetic continuity across generations. While it is generally acknowledged that the timing of germline segregation differs significantly between animals and plants, ongoing debates persist as new evidence continues to emerge. In this review, we delve into studies focusing on male germ cell specifications in plants, and we summarize the core gene regulatory circuits in germ cell specification, which show remarkable parallels to those governing meristem homeostasis. The similarity in germline establishment between animals and plants is also discussed.

Reproductive cell formation An RNA interference screen of 30 gene candidates has identified Lin28, a negative regulator of let-7 microRNA processing, as a potentially key regulator of primordial germ cell development, the process in the developing embryo that selects the cells destined to produce sperm and eggs. In addition, Lin28 levels are elevated in primary human germ cell tumours, suggesting that it may also be implicated in germ cell malignancy. In order to investigate the earliest molecular mechanisms of germ cell specification, mouse embryonic stem cells were differentiated into putative primordial germ cells (PGCs) in vitro . The use of inhibitory RNAs to then screen candidate genes for effects on the development of these cells demonstrates a genetic pathway for PGC specification involving Lin28 , a negative regulator of let-7 microRNA processing. The rarity and inaccessibility of the earliest primordial germ cells (PGCs) in the mouse embryo thwart efforts to investigate molecular mechanisms of germ-cell specification. stella (also called Dppa3 ) marks the rare founder population of the germ lineage 1 , 2 . Here we differentiate mouse embryonic stem cells carrying a stella transgenic reporter into putative PGCs in vitro . The Stella + cells possess a transcriptional profile similar to embryo-derived PGCs, and like their counterparts in vivo , lose imprints in a time-dependent manner. Using inhibitory RNAs to screen candidate genes for effects on the development of Stella + cells in vitro , we discovered that Lin28 , a negative regulator of let-7 microRNA processing 3 , 4 , 5 , 6 , is essential for proper PGC development. Furthermore, we show that Blimp1 (also called Prdm1 ), a let-7 target and a master regulator of PGC specification 7 , 8 , 9 , can rescue the effect of Lin28 deficiency during PGC development, thereby establishing a mechanism of action for Lin28 during PGC specification. Overexpression of Lin28 promotes formation of Stella + cells in vitro and PGCs in chimaeric embryos, and is associated with human germ-cell tumours. The differentiation of putative PGCs from embryonic stem cells in vitro recapitulates the early stages of gamete development in vivo , and provides an accessible system for discovering novel genes involved in germ-cell development and malignancy.

Growth and development of the Ceratopteris hermaphroditic gametophytes are dependent on cell proliferation in the marginal meristem, which when destroyed will regenerate at a new location on the body margin. We established a laser ablation method to destroy a single initial cell in the meristem. Ablation caused the cessation of cell proliferation accompanied by the disappearance of the expression of an auxin synthesis gene (CrTAA2) and a cell proliferation marker gene (CrWOXB). New meristem regeneration occurred within a predictable distance from the original two days post-ablation, signified by cell proliferation and the expression of CrTAA2. Treatment with the naturally occurring auxin indole-3-acetic acid (IAA), synthetic auxin 2,4-dichlorophenoxyacetic acid (2,4-D), or the transport inhibitor naphthylphthalamic acid (NPA) altered positioning of the original marginal meristem toward the apex of the gametophyte. IAA altered positioning of the regenerated meristem after damaging the original meristem. A model of auxin involvement in the positioning of the marginal meristem in Ceratopteris is presented to encompass these results.

Embryonic germ cells develop rapidly to establish the foundation for future developmental trajectories, and in this process, they make critical lineage choices including the configuration of their unique identity and a decision on sex. Here, we use single-cell genomics patterns for the entire embryonic germline in Drosophila melanogaster along with the somatic gonadal precursors after embryonic gonad coalescence to investigate molecular mechanisms involved in the setting up and regulation of the germline program. Profiling of the early germline chromatin landscape revealed sex- and stage-specific features. In the male germline immediately after zygotic activation, the chromatin structure underwent a brief remodeling phase during which nucleosome density was lower and deconcentrated from promoter regions. These findings echoed enrichment analysis results of our genomics data in which top candidates were factors with the ability to mediate large-scale chromatin reorganization. Together, they point to the importance of chromatin regulation in the early germline and raise the possibility of a conserved epigenetic reprogramming-like process required for proper initiation of germline development.

During mammalian development the fertilized zygote and primordial germ cells lose their DNA methylation within one cell cycle leading to the concept of active DNA demethylation. Recent studies identified the TET hydroxylases as key enzymes responsible for active DNA demethylation, catalyzing the oxidation of 5-methylcytosine to 5-hydroxymethylcytosine. Further oxidation and activation of the base excision repair mechanism leads to replacement of a modified cytosine by an unmodified one. In this study, we analyzed the expression/activity of TET1-3 and screened for the presence of 5 mC oxidation products in adult human testis and in germ cell cancers. By analyzing human testis sections, we show that levels of 5-hydroxymethylcytosine, 5-formylcytosine and 5-carboxylcytosine are decreasing as spermatogenesis proceeds, while 5-methylcytosine levels remain constant. These data indicate that during spermatogenesis active DNA demethylation becomes downregulated leading to a conservation of the methylation marks in mature sperm. We demonstrate that all carcinoma in situ and the majority of seminomas are hypomethylated and hypohydroxymethylated compared to non-seminomas. Interestingly, 5-formylcytosine and 5-carboxylcytosine were detectable in all germ cell cancer entities analyzed, but levels did not correlate to the 5-methylcytosine or 5-hydroxymethylcytosine status. A meta-analysis of gene expression data of germ cell cancer tissues and corresponding cell lines demonstrates high expression of TET1 and the DNA glycosylase TDG, suggesting that germ cell cancers utilize the oxidation pathway for active DNA demethylation. During xenograft experiments, where seminoma-like TCam-2 cells transit to an embryonal carcinoma-like state DNMT3B and DNMT3L where strongly upregulated, which correlated to increasing 5-methylcytosine levels. Additionally, 5-hydroxymethylcytosine levels were elevated, demonstrating that de novo methylation and active demethylation accompanies this transition process. Finally, mutations of IDH1 (IDH1 (R132)) and IDH2 (IDH2 (R172)) leading to production of the TET inhibiting oncometabolite 2-hydroxyglutarate in germ cell cancer cell lines were not detected.

Germline stem cells are germ cells at an early developmental stage, so their development is key to ensuring human reproduction. There is increasing evidence that long noncoding RNA (lncRNA) and circular RNA (circRNA) play important roles in the development of germ cells. This data descriptor provides unique lncRNA and circRNA transcriptomic information for mouse germline stem cells. Using the Illumina HiSeqx 2000 system, a total of 511,836,732 raw reads were generated. High-quality transcripts, lncRNAs, and circRNAs were identificated and quantified using the reads, and more precise annotations of lncRNAs (especially 9357 novel lncRNAs) and circRNAs were performed in the germline stem cells. We then analyzed the transcript structures, genetic variants, and the interaction between circRNA and microRNA to provide the basis for subsequent functional experiments. This comprehensive dataset will help advance data sharing and deepen our understanding of mouse germline stem cells, providing a theoretical foundation for research on germ cell development and human reproduction, among others.Design Type(s)transcription profiling design • cell type comparison designMeasurement Type(s)transcription profiling assayTechnology Type(s)RNA sequencingFactor Type(s)technical replicateSample Characteristic(s)Mus musculus • testis • ovaryMachine-accessible metadata file describing the reported data (ISA-Tab format)