Viable Gametes Research Articles

Functional recovery of the germ line following splicing collapse.

Splicing introns from precursor-messenger RNA (pre-mRNA) transcripts is essential for translating functional proteins. Here, we report that the previously uncharacterized Caenorhabditis elegans protein MOG-7 acts as a pre-mRNA splicing factor. Depleting MOG-7 from the C. elegans germ line causes intron retention in most germline-expressed genes, impeding the germ cell cycle, and causing defects in nuclear morphology, germ cell identity and sterility. Despite the deleterious consequences caused by MOG-7 loss, the adult germ line can functionally recover to produce viable and fertile progeny when MOG-7 is restored. Germline recovery is dependent on a burst of apoptosis that likely clears defective germ cells, and viable gametes generated from the proliferation of germ cells in the progenitor zone. Together, these findings reveal that MOG-7 is essential for germ cell development, and that the germ line can functionally recover after a collapse in RNA splicing.

Downregulation of Barley Regulator of Telomere Elongation Helicase 1 Alters the Distribution of Meiotic Crossovers.

Programmed meiotic DNA double-strand breaks (DSBs), necessary for proper chromosomal segregation and viable gamete formation, are repaired by homologous recombination (HR) as crossovers (COs) or non-crossovers (NCOs). The mechanisms regulating the number and distribution of COs are still poorly understood. The regulator of telomere elongation helicase 1 (RTEL1) DNA helicase was previously shown to enforce the number of meiotic COs in Caenorhabditis elegans but its function in plants has been studied only in the vegetative phase. Here, we characterised barley RTEL1 gene structure and expression using RNA-seq data previously obtained from vegetative and reproductive organs and tissues. Using RNAi, we downregulated RTEL1 expression specifically in reproductive tissues and analysed its impact on recombination using a barley 50k iSelect SNP Array. Unlike in C. elegans, in a population segregating for RTEL1 downregulated by RNAi, high resolution genome-wide genetic analysis revealed a significant increase of COs at distal chromosomal regions of barley without a change in their total number. Our data reveal the important role of RTEL1 helicase in plant meiosis and control of recombination.

Lager Yeast Design Through Meiotic Segregation of a Saccharomyces cerevisiae × Saccharomyces eubayanus Hybrid.

Yeasts in the lager brewing group are closely related and consequently do not exhibit significant genetic variability. Here, an artificial Saccharomyces cerevisiae × Saccharomyces eubayanus tetraploid interspecies hybrid was created by rare mating, and its ability to sporulate and produce viable gametes was exploited to generate phenotypic diversity. Four spore clones obtained from a single ascus were isolated, and their brewing-relevant phenotypes were assessed. These F1 spore clones were found to differ with respect to fermentation performance under lager brewing conditions (15°C, 15 °Plato), production of volatile aroma compounds, flocculation potential and temperature tolerance. One spore clone, selected for its rapid fermentation and acetate ester production was sporulated to produce an F2 generation, again comprised of four spore clones from a single ascus. Again, phenotypic diversity was introduced. In two of these F2 clones, the fermentation performance was maintained and acetate ester production was improved relative to the F1 parent and the original hybrid strain. Strains also performed well in comparison to a commercial lager yeast strain. Spore clones varied in ploidy and chromosome copy numbers, and faster wort fermentation was observed in strains with a higher ploidy. An F2 spore clone was also subjected to 10 consecutive wort fermentations, and single cells were isolated from the resulting yeast slurry. These isolates also exhibited variable fermentation performance and chromosome copy numbers, highlighting the instability of polyploid interspecific hybrids. These results demonstrate the value of this natural approach to increase the phenotypic diversity of lager brewing yeast strains.

Cryopreservation of gametes: history and current state of a question

Development of reproductive medicine, namely, extracorporal fertilization, set the task of preservation and preserving of biological suitability of female and men's gametes for scientific community. Opening of a method of a vitrification showed its benefit before other methods of a cryopreservation and became the encouraging event in development of cryobank of gametes. In this work the history of emergence of a method of a cryopreservation is described; modern aspects of a cryopreservation of gametes; shortcomings and benefits of various techniques of selection of full-fledged gametes are reflected; some protocols of carrying out a cryo-preservation are provided; short characteristic of separately used cryoprotectors and their various combinations capable more effectively to protect the freezing objects from damage, than, each of cryoprotectors separately is given. It is noted that in modern literature special attention is paid to consideration of mechanisms of cryodamages and cryoprotection during the freezing and a vitrification. Characteristics of the changes happening in gametes after the procedure of freezing and thawing are discussed. Modern techniques of assessment of viability of gametes before and after a cryopreservation are given. It is shown that violation of compaction of chromatin and fragmentation of DNA in morphologically not changed spermiya has negative impact and on quality of embryos. The benefit of a structurally functional condition of oocytes of mammals at a cryopreservation of oocytes in structure is emphasized an oocyte-kumulyusnykh of complexes in comparison with the oocytes which ripened in the kultivatsionny environment.

Reduced gamete viability associated to somatic genome rearrangements increases fruit set sensitivity to the environment in Tempranillo Blanco grapevine cultivar

Sensitivity of fruit set to the environment is a genetic feature of uncertain origin that limits production in some grapevine clones and cultivars. Here we studied the developmental causes and environmental conditions leading to decreased fruit yield associated to somatic loss of fruit colour in Tempranillo Blanco (TB) white-berried derivative cultivar. We first compared fruit set-related production traits between TB and its black-berried clonal ancestor, Tempranillo Tinto (TT) cultivar, both grown under the same field conditions. We identified that lower cluster weight in TB correlated with a genetic reduction in pollen viability and lower fruit and seed setting. Then, a combination of correlation and two-way-ANOVA (Analysis of Variance) procedures, along with a series of multivariate linear regression models were developed to examine the effect of genotype and pre-flowering environmental factors on the reproductive performance of TB, using data recorded in nine field plots. These analyses identified prolonged pre-flowering cold periods and abundant rainfalls at flowering time as the main conditions increasing the ratio of seedless to seeded berries in the cluster, whereas the low pollen viability and low number of seeds characteristic of TB barely varied across plots. Collectively, these findings indicate that decreased gamete viability caused by complex genome rearrangements is in the origin of increased susceptibility to fruit set disorders in grapevine. The case of TB shows that such dysfunctions can be selected in clonally propagated crops as a trade-off of novel interesting traits emerged after genome reshuffling.

Evaluation of the TMRW vapor phase cryostorage platform using reproductive specimens and in vitro extended human embryo culture

To assess the impact of shipment and storage of sperm, oocytes, and blastocysts in vapor phase nitrogen compared with static storage in liquid phase nitrogen. Prospective cohort-matched study. Multiple invitro fertilization laboratories in an invitro fertilization network. Fifty-eight human embryos, 32 human oocytes, 15 units of bovine semen. Vapor vs. liquid nitrogen. The postwarming survival of oocytes, sperm, and blastocysts, and the developmental potential of blastocysts during invitro extended culture. Custom-designed labware, for use with the TMRW platform, enables continuous temperature monitoring during shipment and/or storage in the vapor phase robotic storage system. The highest temperature recorded for specimens shipped to a domestic laboratory was -180.2 °C with a mean ± SD of -190.4 ± 0.5 °C during shipment and -181.1 ± 0.6 °C during storage. Likewise, specimens shipped internationally had a high of -180.2 °C with a mean ± SD of -193.5 ± 0.6 °C during shipment and -181.2 ± 0.7 °C during storage. Results from the extended culture assays have revealed no deleterious effect of shipment and storage in nitrogen vapor. The viability of mammalian gametes and embryos was equivalent between the vapor phase and liquid phase storage. The evaluated system did not have any deleterious effects on the postwarming survival of sperm, oocytes, and blastocysts. The postwarming developmental potential of human blastocysts during invitro extended culture was unaffected by storage and handling in the vapor phase nitrogen TMRW platform when compared with static liquid phase nitrogen storage. Our results suggest that the vapor phase cryostorage platform is a safe system to handle and store reproductive specimens for human assisted reproductive technology.

The toxic dinoflagellate Alexandrium minutum affects oyster gamete health and fertilization potential

Dinoflagellates from the globally distributed genus Alexandrium are known to produce both paralytic shellfish toxins (PST) and uncharacterized bioactive extracellular compounds (BEC) with allelopathic, ichthyotoxic, hemolytic and cytotoxic activities. In France, blooms of Alexandrium minutum appear generally during the spawning period of most bivalves. These blooms could therefore alter gametes and/or larval development of bivalves, causing severe issues for ecologically and economically important species, such as the Pacific oyster Crassostrea (=Magallana) gigas. The aim of this work was to test the effects of three strains of A. minutum producing either only PST, only BEC, or both PST and BEC upon oyster gametes, and potential consequences on fertilization success. Oocytes and spermatozoa were exposed in vitro for 2 h to a range of environmentally realistic A. minutum concentrations (10–2.5 × 104 cells mL−1). Following exposure, gamete viability and reactive oxygen species (ROS) production were assessed by flow cytometry, spermatozoa motility and fertilization capacities of both spermatozoa and oocytes were analysed by microscopy. Viability and fertilization capacity of spermatozoa and oocytes were drastically reduced following exposure to 2.5 × 104 cells mL−1 of A. minutum. The BEC-producing strain was the most potent strain decreasing spermatozoa motility, increasing ROS production of oocytes, and decreasing fertilization, from the concentration of 2.5 × 103 cells mL−1. This study highlights the significant cellular toxicity of the BEC produced by A. minutum on oyster gametes. Physical contact between gametes and motile thecate A. minutum cells may also contribute to alter oyster gamete integrity. These results suggest that oyster gametes exposure to A. minutum blooms could affect oyster fertility and reproduction success.

DEFECTIVE EMBRYO AND MERISTEMS genes are required for cell division and gamete viability in Arabidopsis.

The DEFECTIVE EMBRYO AND MERISTEMS 1 (DEM1) gene encodes a protein of unknown biochemical function required for meristem formation and seedling development in tomato, but it was unclear whether DEM1’s primary role was in cell division or alternatively, in defining the identity of meristematic cells. Genome sequence analysis indicates that flowering plants possess at least two DEM genes. Arabidopsis has two DEM genes, DEM1 and DEM2, which we show are expressed in developing embryos and meristems in a punctate pattern that is typical of genes involved in cell division. Homozygous dem1 dem2 double mutants were not recovered, and plants carrying a single functional DEM1 allele and no functional copies of DEM2, i.e. DEM1/dem1 dem2/dem2 plants, exhibit normal development through to the time of flowering but during male reproductive development, chromosomes fail to align on the metaphase plate at meiosis II and result in abnormal numbers of daughter cells following meiosis. Additionally, these plants show defects in both pollen and embryo sac development, and produce defective male and female gametes. In contrast, dem1/dem1 DEM2/dem2 plants showed normal levels of fertility, indicating that DEM2 plays a more important role than DEM1 in gamete viability. The increased importance of DEM2 in gamete viability correlated with higher mRNA levels of DEM2 compared to DEM1 in most tissues examined and particularly in the vegetative shoot apex, developing siliques, pollen and sperm. We also demonstrate that gamete viability depends not only on the number of functional DEM alleles inherited following meiosis, but also on the number of functional DEM alleles in the parent plant that undergoes meiosis. Furthermore, DEM1 interacts with RAS-RELATED NUCLEAR PROTEIN 1 (RAN1) in yeast two-hybrid and pull-down binding assays, and we show that fluorescent proteins fused to DEM1 and RAN1 co-localize transiently during male meiosis and pollen development. In eukaryotes, RAN is a highly conserved GTPase that plays key roles in cell cycle progression, spindle assembly during cell division, reformation of the nuclear envelope following cell division, and nucleocytoplasmic transport. Our results demonstrate that DEM proteins play an essential role in cell division in plants, most likely through an interaction with RAN1.

The Pif1 helicase is actively inhibited during meiotic recombination which restrains gene conversion tract length.

Meiotic recombination ensures proper chromosome segregation to form viable gametes and results in gene conversions events between homologs. Conversion tracts are shorter in meiosis than in mitotically dividing cells. This results at least in part from the binding of a complex, containing the Mer3 helicase and the MutLβ heterodimer, to meiotic recombination intermediates. The molecular actors inhibited by this complex are elusive. The Pif1 DNA helicase is known to stimulate DNA polymerase delta (Pol δ) -mediated DNA synthesis from D-loops, allowing long synthesis required for break-induced replication. We show that Pif1 is also recruited genome wide to meiotic DNA double-strand break (DSB) sites. We further show that Pif1, through its interaction with PCNA, is required for the long gene conversions observed in the absence of MutLβ recruitment to recombination sites. In vivo, Mer3 interacts with the PCNA clamp loader RFC, and in vitro, Mer3-MutLβ ensemble inhibits Pif1-stimulated D-loop extension by Pol δ and RFC-PCNA. Mechanistically, our results suggest that Mer3-MutLβ may compete with Pif1 for binding to RFC-PCNA. Taken together, our data show that Pif1’s activity that promotes meiotic DNA repair synthesis is restrained by the Mer3-MutLβ ensemble which in turn prevents long gene conversion tracts and possibly associated mutagenesis.

Southwestern Atlantic reef-building corals Mussismilia spp. are able to spawn while fully bleached

Bleaching events are becoming more frequent and intense worldwide. Southwestern Atlantic corals are considered stress-tolerant species that suffer less during bleaching episodes. Therefore, we investigated if the reef-building corals Mussismilia hispida and M. harttii are capable of spawning while fully bleached. We compared spermatozoa concentration and motility, egg diameter and embryo viability between seemingly healthy and bleached colonies for both species. Findings show that both species spawned viable gametes. Concentration and motility of newly-released spermatozoa were similar between healthy and bleached colonies for both species. Unlike M. hispida, a 10% reduction in size was found in egg diameter for M. harttii. Embryo viability for bleached M. hispida colonies did not decrease; for bleached M. harttii there was a reduction from 14.7 to 6.3%, but fertilization and larval development still took place. These findings describe the unprecedented episode of corals spawning viable gametes when entirely bleached, supporting the suggestion of greater resilience for Southwestern Atlantic corals.

Long-term dietary diludine (diethyl 1,4-dihydro-2,6-dimethyl-3,5-pyridinedicarboxylate) intake and gamete storage affect α-tocopherol status and reproductive performance in female rainbow trout, Oncorhynchus mykiss

Rainbow trout broodstocks were fed diets with varying levels of diludine [0 (D0; control), 0.25 (D1), 0.5 (D2) and 1 g kg−1 (D3)] during 2 consecutive reproduction seasons. Fish were started to be fed 3 months before spawning for the first reproduction season and they were kept on the same diets as well as an additional dietary group with 3 g kg−1 dietary diludine (D4) until the following reproduction season, providing short and long-term dietary treatments. Gametes storage at 6 °C for four and fifteen days prior to the fertilization in the first and second reproduction season, respectively, was also tested. Short-term feeding trial did not influence reproductive performance when the eggs were fertilized immediately after stripping, whereas survival at eyed-stage embryos significantly decreased in eggs, from fish fed diludine free diet, fertilized after 4-day storage. Dietary diludine level corresponded with increasing level of vitamin E in the eggs, which positively correlated with survival at eyed-stage embryos from the eggs with or without 15-day storage after long-term feeding. Fish fed the diet with highest level of diludine (D4) produced eggs higher in number but smaller in size in comparison to those from the other four experimental groups. In eggs fertilized after 15-day storage, survival rate was 20-fold higher in D4 diet fed fish in comparison to the other experimental groups. Our overall results indicate that dietary diludine has vitamin E sparing effect in the eggs and protects gamete viability during the storage when it is provided at 3 mg kg−1 dietary level, which result in lower lipid peroxidation and higher reproductive performance.

THE EFFECT OF DROUGHT ON WHEAT PLANTS AT DIFFERENT GROWTH STAGES

The climate conditions of Ukraineand particularly Odesa region are gradually changing: the average temperatures increase, the rainfall distribution becomes irregular and cases of extreme weather conditions, such as droughts, rainstorms, significant temperature fluctuations, become more frequent. It necessitates detailed studies of the effect of climatic factors on economically important plants, the mechanisms of the formation of their drought resistance. This knowledge will facilitate the selection process of universal varieties and improve the system of agrotechnical events. The purpose of this study is the systematisation of the information on the effects of moisture deficiency on biochemical processes, growth, development, and productivity of soft winter wheat ( Triticum aestivum L.), depending on the phenological phase during which drought occurs in the Southern Steppe zone of Ukraine, particularly in Odesa region. For this purpose, present publications on the impact of drought on wheat during different stages of development were analysed; agrometeorological conditions of Odesa region within the last 20 years were tracked; modern soft wheat varieties adapted to the arid conditions of the cultivation inSouthern Ukraine were marked. It was noted that moisture deficiency during the vegetative stage affects the biomass accumulation and formation of productive tillers, and during flowering and ripening, it reduces grain yield and deteriorates its quality. At the germination stage, drought leads to uneven sprouting, impairs mineral nutrition, inhibits the growth of aboveground parts. At the tillering stage, fewer productive shoots are formed, photosynthesis is suppressed, phenological phases are displaced – flowering and maturation are accelerated. A drought during heading leads to the reduction of the number of grains in spikes. Insufficient moisture supply during flowering impairs fertility and viability of gametes, and during grain filling, it inhibits endosperm cell division and adversely affects grain quality. In general, the impact of drought on biochemical and physiological processes of soft winter wheat development has not been thoroughly studied and needs further investigation. Key words : Triticum aestivum L.; phenology; water stress; climate; cultivars.

Meiotic Chromosome Stability and Suppression of Crossover Between Non-homologous Chromosomes in xBrassicoraphanus, an Intergeneric Allotetraploid Derived From a Cross Between Brassica rapa and Raphanus sativus.

Hybridization and polyploidization are major driving forces in plant evolution. Allopolyploids can be occasionally formed from a cross between distantly related species but often suffer from chromosome instability and infertility. xBrassicoraphanus is an intergeneric allotetraploid (AARR; 2n = 38) derived from a cross between Brassica rapa (AA; 2n = 20) and Raphanus sativus (RR; 2n = 18). xBrassicoraphanus is fertile and genetically stable, while retaining complete sets of both B. rapa and R. sativus chromosomes. Precise control of meiotic recombination is essential for the production of balanced gametes, and crossovers (COs) must occur exclusively between homologous chromosomes. Many interspecific hybrids have problems with meiotic division at early generations, in which interactions between non-homologous chromosomes often bring about aneuploidy and unbalanced gamete formation. We analyzed meiotic chromosome behaviors in pollen mother cells (PMCs) of allotetraploid and allodiploid F1 individuals of newly synthesized xBrassicoraphanus. Allotetraploid xBrassicoraphanus PMCs showed a normal diploid-like meiotic behavior. By contrast, allodiploid xBrassicoraphanus PMCs displayed abnormal segregation of chromosomes mainly due to the absence of homologous pairs. Notably, during early stages of meiosis I many of allodiploid xBrassicoraphanus chromosomes behave independently with few interactions between B. rapa and R. sativus chromosomes, forming many univalent chromosomes before segregation. Chromosomes were randomly assorted at later stages of meiosis, and tetrads with unequal numbers of chromosomes were formed at completion of meiosis. Immunolocalization of HEI10 protein mediating meiotic recombination revealed that COs were more frequent in synthetic allotetraploid xBrassicoraphanus than in allodiploid, but less than in the stabilized line. These findings suggest that structural dissimilarity between B. rapa and R. sativus chromosomes prevents non-homologous interactions between the parental chromosomes in allotetraploid xBrassicoraphanus, allowing normal diploid-like meiosis when homologous pairing partners are present. This study also suggests that CO suppression between non-homologous chromosomes is required for correct meiotic progression in newly synthesized allopolyploids, which is important for the formation of viable gametes and reproductive success in the hybrid progeny.

Diversity and Postzygotic Evolution of the Mitochondrial Genome in Hybrids of Saccharomyces Species Isolated by Double Sterility Barrier.

Eukaryotic species are reproductively isolated by sterility barriers that prevent interspecies fertilization (prezygotic sterility barrier) or the fertilization results in infertile offspring (postzygotic sterility barrier). The Saccharomyces species are isolated by postzygotic sterility barriers. Their allodiploid hybrids form no viable gametes (ascospores) and the viable ascospores of the allotetraploids cannot fertilize (conjugate). Our previous work revealed that this mechanism of reproductive isolation differs from those operating in plants and animals and we designated it double sterility barrier (the failure of homeologous chromosomes to pair and the repression of mating by mating-type heterozygosity). Other studies implicated nucleo-mitochondrial incompatibilities in the sterility of the Saccharomyces hybrids, a mechanism assumed to play a central role in the reproductive isolation of animal species. In this project the mitochondrial genomes of 50 cevarum (S. cerevisiae × S. uvarum) hybrids were analyzed. 62% had S. cerevisiae mitotypes, 4% had S. uvarum mitotypes, and 34% had recombinant mitotypes. All but one hybrid formed viable spores indicating that they had genomes larger than allodiploid. Most of these spores were sterile (no sporulation in the clone of vegetative descendants; a feature characteristic of allodiploids). But regardless of their mitotypes, most hybrids could also form fertile alloaneuploid spore clones at low frequencies upon the loss of the MAT-carrying chromosome of the S. uvarum subgenome during meiosis. Hence, the cevarum alloploid nuclear genome is compatible with both parental mitochondrial genomes as well as with their recombinants, and the sterility of the hybrids is maintained by the double sterility barrier (determined in the nuclear genome) rather than by nucleo-mitochondrial incompatibilities. During allotetraploid sporulation both the nuclear and the mitochondrial genomes of the hybrids could segregate but no correlation was observed between the sterility or the fertility of the spore clones and their mitotypes. Nucleo-mitochondrial incompatibility was manifested as respiration deficiency in certain meiotic segregants. As respiration is required for meiosis-sporulation but not for fertilization (conjugation), these segregants were deficient only in sporulation. Thus, the nucleo-mitochondrial incompatibility affects the sexual processes only indirectly through the inactivation of respiration and causes only partial sterility in certain segregant spore clones.

Rapid evolution of piRNA-mediated silencing of an invading transposable element was driven by abundant de novo mutations.

The regulation of transposable element (TE) activity by small RNAs is a ubiquitous feature of germlines. However, despite the obvious benefits to the host in terms of ensuring the production of viable gametes and maintaining the integrity of the genomes they carry, it remains controversial whether TE regulation evolves adaptively. We examined the emergence and evolutionary dynamics of repressor alleles after P-elements invaded the Drosophila melanogaster genome in the mid-twentieth century. In many animals including Drosophila, repressor alleles are produced by transpositional insertions into piRNA clusters, genomic regions encoding the Piwi-interacting RNAs (piRNAs) that regulate TEs. We discovered that ∼94% of recently collected isofemale lines in the Drosophila melanogaster Genetic Reference Panel (DGRP) contain at least one P-element insertion in a piRNA cluster, indicating that repressor alleles are produced by de novo insertion at an exceptional rate. Furthermore, in our sample of approximately 200 genomes, we uncovered no fewer than 80 unique P-element insertion alleles in at least 15 different piRNA clusters. Finally, we observe no footprint of positive selection on P-element insertions in piRNA clusters, suggesting that the rapid evolution of piRNA-mediated repression in D. melanogaster was driven primarily by mutation. Our results reveal for the first time how the unique genetic architecture of piRNA production, in which numerous piRNA clusters can encode regulatory small RNAs upon transpositional insertion, facilitates the nonadaptive rapid evolution of repression.

Dramatically diverse Schizosaccharomyces pombe wtf meiotic drivers all display high gamete-killing efficiency.

Meiotic drivers are selfish alleles that can force their transmission into more than 50% of the viable gametes made by heterozygotes. Meiotic drivers are known to cause infertility in a diverse range of eukaryotes and are predicted to affect the evolution of genome structure and meiosis. The wtf gene family of Schizosaccharomyces pombe includes both meiotic drivers and drive suppressors and thus offers a tractable model organism to study drive systems. Currently, only a handful of wtf genes have been functionally characterized and those genes only partially reflect the diversity of the wtf gene family. In this work, we functionally test 22 additional wtf genes for meiotic drive phenotypes. We identify eight new drivers that share between 30–90% amino acid identity with previously characterized drivers. Despite the vast divergence between these genes, they generally drive into >85% of gametes when heterozygous. We also identify three wtf genes that suppress other wtf drivers, including two that also act as autonomous drivers. Additionally, we find that wtf genes do not underlie a weak (64% allele transmission) meiotic driver on chromosome 1. Finally, we find that some Wtf proteins have expression or localization patterns that are distinct from the poison and antidote proteins encoded by drivers and suppressors, suggesting some wtf genes may have non-meiotic drive functions. Overall, this work expands our understanding of the wtf gene family and the burden selfish driver genes impose on S. pombe.

State-of-the-art in reproductive bench science: Hurdles and new technological solutions

Infertility is a growing issue in modern society, being the fifth highest serious global disability according to the World Health Organization. To study infertility and other reproductive system complications, bench science still relies on 2D and animal studies, which regularly have been criticized due to their inability to mimic the human body. Particular challenges in 2D studies include the inability to mimic fluid dynamics, gametes modulation and their crosstalk, hormonal patterns as well as the low quality and viability of gametes and embryos. Animal models also present other drawbacks, namely the absence of menstruation, making it difficult to establish a reliable predictive model for the human system. Additionally, reproductive studies should not be limited to the fallopian tube as the sole responsible for most infertility cases, but instead the research spectrum should be widened to the whole reproductive system given the tight interconnectivity between each and every organ. In the last few decades, new in vitro technologies have been developed and applied to the study of reproductive system complications. These systems allow to create complex three-dimensional structures, which are therefore able to more closely resemble specific microenvironments and provide more realistic physical and biochemical cues. 3D (bio)printing, organoids and organs-on-chips are some of the dynamic technologies which are replacing conventionally employed static 2D culture. Herein, we provide an overview of the challenges found in conventional 2D and animal models of the reproductive system and present potential technological solutions for those same challenges.

Transcription-Wide Mapping of Dihydrouridine (D) Reveals that mRNA Dihydrouridylation is Essential for Meiotic Chromosome Segregation

Epitranscriptomic has emerged as a fundamental control of gene expression. Nevertheless, the determination of the transcriptome-wide occupancy of RNA modifications remains challenging. We have developed Rho-seq, an integrated pipeline detecting a range of modifications through differential modification-dependent Rhodamine labeling. Using Rho-seq, we confirm that the reduction of uridine to dihydrouridine targets tRNAs in E. coli. Unexpectedly, we find that the D modification expands to mRNAs in fission yeast.The modified mRNAs are enriched for cytoskeleton-related encoding proteins. We show that the α-tubulin encoding mRNA nda2 undergoes dihydrouridylation, which affects its protein expression level. The absence of the modification onto the nda2 mRNA impacts meiosis by inducing a metaphase delay or by completely preventing the formation of spindles during meiosis I and meiosis II, resulting in low gamete viability. Collectively these data show that the codon-specific reduction of uridine within specific mRNA is required for proper meiotic chromosome segregation and gamete viability.

Diets for Macrobrachium acanthurus breeders: Sperm evaluation

Abstract The application of biotechnology in animal reproduction has enabled the production of young forms in both quantity and quality. Increasing the number of viable gametes produced by reproducers, among other factors, through an ideal diet, can ensure higher production. Therefore, the aim of this study was to determine the influence of three diets on the sperm survival of Macrobrachium acanthurus. To this end, 24 M. acanthurus males were used, distributed randomly and equally among treatments. Their diets were composed of 100% fresh food (fish and squid muscle - 14% protein), 100% dry feed (commercial feed - 50% protein) and a mixture of these diets containing 30% protein. Spermatophores were extracted through electrical stimulation every 15 days, and the controls consisted of spermatophores obtained directly from nature. No significant difference between diets was observed comparing shrimp and spermatophore weights. The 100% fresh diet provided the best sperm survival performance.

ATR function is indispensable to allow proper mammalian follicle development.

Mammalian female fertility relies on the proper development of follicles. Right after birth in the mouse, oocytes associate with somatic ovarian cells to form follicles. These follicles grow during the adult lifetime to produce viable gametes. In this study, we analyzed the role of the ATM and rad3-related (ATR) kinase in mouse oogenesis and folliculogenesis using a hypomorphic mutation of the Atr gene (Murga et al. 2009). Female mice homozygotes for this allele have been reported to be sterile. Our data show that female meiotic prophase is not grossly altered when ATR levels are reduced. However, follicle development is substantially compromised, since Atr mutant ovaries present a decrease of growing follicles. Comprehensive analysis of follicular cell death and proliferation suggest that wild-type levels of ATR are required to achieve optimal follicular development. Altogether, these findings suggest that reduced ATR expression causes sterility due to defects in follicular progression rather than in meiotic recombination. We discuss the implications of these findings for the use of ATR inhibitors such as anti-cancer drugs and its possible side-effects on female fertility.